MedWorm.com provides a medical RSS filtering service. Over 6000 RSS medical sources are combined and output via different filters. This feed contains the latest items from the 'Cytometry Part A' source. Mon, 06 Feb 2012 16:52:04 +0100 http://www.medworm.com/index.php?rid=5643842&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22020 This article assesses several autofocus algorithms applied in the study of fluorescence‐labeled tuberculosis bacteria. The goal of this work was to find the optimal algorithm in order to build an automatic real‐time system for diagnosing sputum smear samples, where both accuracy and computational time are important. We analyzed 13 focusing methods, ranging from well‐known algorithms to the most recently proposed functions. We took into consideration criteria that are inherent to the autofocus function, such as accuracy, computational cost, and robustness to noise and to illumination changes. We also analyzed the additional benefit provided by preprocessing techniques based on morphological operators and image projection profiling. © 2012 International Society for Advancement of Cyto... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5643842 Tue, 31 Jan 2012 14:46:32 +0100 5643842 http://www.medworm.com/index.php?rid=5643844&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22023 AbstractProper illumination is essential for light microscopy. Whereas in early years incandescent light was the only illumination, today, more and more specialized light sources, such as lasers or arc lamps are used. Because of the high efficiency and brightness that light‐emitting diodes (LED) have reached today, they have become a serious alternative for almost all kinds of illumination in light microscopy. LED have a high durability, do not need expensive electronics, and they can be switched in nanoseconds. Besides this, they are available throughout the UV/Vis/NIR‐spectrum with a narrow bandwidth. This makes them ideal light sources for fluorescence microscopy. The white LED, with a color temperature ranging from 2,600 up to 5,000 K is an excellent choice for bright‐field illum... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5643844 Mon, 30 Jan 2012 05:00:00 +0100 5643844 http://www.medworm.com/index.php?rid=5643843&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22006 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5643843 Mon, 30 Jan 2012 05:00:00 +0100 5643843 http://www.medworm.com/index.php?rid=5635416&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22018 AbstractThe Flow Cytometry Standard (FCS) format was developed back in 1984. Since then, FCS became the standard file format supported by all flow cytometry software and hardware vendors. Over the years, updates were incorporated to adapt to technological advancements in both flow cytometry and computing technologies. However, flexibility in how data may be stored in FCS has led to implementation difficulties for instrument vendors and third party software developers. In this technical note, we are providing implementation guidance and examples related to FCS 3.1, the latest version of the standard. By publishing this text, we intend to prevent potential compatibility issues that could be faced when implementing the FCS spillover and preferred display keywords that have arisen during discu... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5635416 Fri, 27 Jan 2012 21:40:46 +0100 5635416 http://www.medworm.com/index.php?rid=5635417&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22016 AbstractThe development of polychromatic cytometry has contributed to significant progress in the field of human immunology. Although numerous functional studies of rare cell populations have been performed using this technology, here we used polychromatic cytometry to explore the dynamics of complex cellular systems implicated in innate immunity. We used PBMC stimulated with live influenza virus as an experimental model. We studied the time course of activation of PBMC, which contain DC, monocytes, and NK cells, all of which are, in addition to their innate immune properties, susceptible to Flu infection. We developed 12 color panels to investigate intracellular expression of IFN‐α, TNF‐α, IL‐12, IL‐6, IFN‐γ, CD107, and influenza virus nucleoprotein simultaneously in these ce... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5635417 Wed, 25 Jan 2012 05:00:00 +0100 5635417 http://www.medworm.com/index.php?rid=5624791&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22015 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5624791 Tue, 24 Jan 2012 22:02:15 +0100 5624791 http://www.medworm.com/index.php?rid=5624790&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22124 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5624790 Tue, 24 Jan 2012 22:02:13 +0100 5624790 http://www.medworm.com/index.php?rid=5624789&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22123 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5624789 Tue, 24 Jan 2012 22:02:12 +0100 5624789 http://www.medworm.com/index.php?rid=5624788&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22122 AbstractOn the cover: Zebrafish vasculature portrayed in a whole new light on the cover of this issue. The source work by Zhang and coworkers in the current issue describes an integrated optical system, combining laser scanning confocal microscopy and in vivo flow cytometry, to track cells (both stationary and circulating) in zebrafish.Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5624788 Tue, 24 Jan 2012 22:02:11 +0100 5624788 http://www.medworm.com/index.php?rid=5617027&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22013 AbstractAcute promyelocytic leukemia (APL) is a hematological emergency in which a rapid diagnosis is essential for early administration of appropriate therapy, including all‐trans retinoic acid before the onset of fatal coagulopathy. Currently, the following methodologies are widely used for rapid initial diagnosis of APL: 1) identification of hypergranular leukemic promyelocytes by using classical morphology; 2) identification of cells with diffuse promyelocytic leukemia (PML) protein distribution by immunofluorescence microscopy; 3) evidence of aberrant promyelocyte surface immunophenotype by conventional flow cytometry (FCM). Here, we show a method for immunofluorescent detection of PML localization using ImageStream FCM. This technique provides objective per‐cell quantitative imag... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5617027 Sun, 01 Jan 2012 05:00:00 +0100 5617027 http://www.medworm.com/index.php?rid=5603993&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22014 AbstractIn cerebrospinal fluid (CSF) analysis, hematology analyzers (HAs) Sysmex® XT‐4000i and XE‐5000, equipped with flow cytometry (FCM), were used to count cells and differentiate leukocytes into mononuclear and polymorphonuclear cells (MNCs, PMCs) applying body fluid mode. FCM was evaluated with 20 DGKL CSF controls containing viable human leukocytes and erythrocytes. HA values were compared with reference values by Passing/Bablok regression analysis to reveal conformity. Conformity of white blood cells (WBCs) was obtained with native leukocytes, counted in calibrated Fuchs–Rosenthal chamber as reference; red blood cell counts proved inaccurate. CV <40% with WBC counts <20 per μL impairs accuracy. Reference WBC differentiation was assayed using FACS Canto II™ and FC‐5... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5603993 Sun, 01 Jan 2012 05:00:00 +0100 5603993 http://www.medworm.com/index.php?rid=5568853&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22003 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5568853 Sun, 01 Jan 2012 05:00:00 +0100 5568853 http://www.medworm.com/index.php?rid=5556786&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22012 AbstractCurrently, there is no standardized panel for immunophenotyping myeloid cells in mouse spleen using flow cytometry. Markers such as CD11b, CD11c, F4/80, Gr‐1, Ly6C, and Ly6G have long been used to identify various splenic cell myeloid populations. Flow cytometry and fluorescence‐activated cell sorting (FACS) analysis demonstrated that Ly6G/Ly6C markers are superior to Gr‐1 for identifying splenic neutrophils, eosinophils, and subsets of monocytes/macrophages. Moreover, these experiments showed that F4/80 is not required for identifying these myeloid subsets and that many of the commercially available preparations of anti‐F4/80 antibodies stain poorly for this antigen in spleen. Taken together, we have now developed an informative flow cytometry panel that can be combined wi... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5556786 Sun, 01 Jan 2012 05:00:00 +0100 5556786 http://www.medworm.com/index.php?rid=5549798&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22007 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5549798 Thu, 29 Dec 2011 15:09:39 +0100 5549798 http://www.medworm.com/index.php?rid=5556787&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22010 AbstractCardiovascular disease (CVD) is now the largest killer in western society, and the importance of interactions between vascular endothelium and circulating blood components in disease pathogenesis is well established. Microparticles are a heterogeneous population of <1 μm blood borne particles that arise from blebbing or shedding of cell membranes. The microparticle population includes several classes of apoptotic bodies; however, increased numbers of procoagulant microparticles have been described in plasma from people with CVD. We have previously demonstrated that interactions of monocytes and platelets with isolated inflamed endothelial cells lead to production of pro‐coagulant tissue factor bearing microparticles under laminar flow conditions. Here we have investigated mic... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5556787 Thu, 29 Dec 2011 05:00:00 +0100 5556787 http://www.medworm.com/index.php?rid=5549800&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22009 AbstractAmyloid beta (Aβ) oligomers and phosphorylated tau (p‐tau) aggregates are increasingly identified as potential toxic intermediates in Alzheimer's disease (AD). In cortical AD synapses, p‐tau co‐localizes with Aβ, but the Aβ and p‐tau peptide species responsible for synaptic dysfunction and demise remains unclear. The present experiments were designed to use high‐speed cell sorting techniques to purify synaptosome population based on size, and then extend the method to physically isolate Aβ‐positive synaptosomes with the goal of understanding the nature of Aβ and tau pathology in AD synapses. To examine the purity of size‐gated synaptosomes, samples were first gated on size; particles with sizes between 0.5 and 1.5 microns were collected. Electron microscopy docum... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5549800 Wed, 28 Dec 2011 05:00:00 +0100 5549800 http://www.medworm.com/index.php?rid=5549799&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22011 AbstractThe combination of color‐coded microspheres as carriers and flow cytometry as a detection platform provides new opportunities for multiplexed measurement of biomolecules. Here, we developed a software tool capable of automated gating of color‐coded microspheres, automatic extraction of statistics from all subsets and validation, normalization, and cross‐sample analysis. The approach presented in this article enabled us to harness the power of high‐content cellular proteomics. In size exclusion chromatography‐resolved microsphere‐based affinity proteomics (Size‐MAP), antibody‐coupled microspheres are used to measure biotinylated proteins that have been separated by size exclusion chromatography. The captured proteins are labeled with streptavidin phycoerythrin and de... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5549799 Wed, 28 Dec 2011 05:00:00 +0100 5549799 http://www.medworm.com/index.php?rid=5520363&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22004 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5520363 Tue, 20 Dec 2011 09:34:43 +0100 5520363 http://www.medworm.com/index.php?rid=5520362&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22000 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5520362 Tue, 20 Dec 2011 09:34:42 +0100 5520362 http://www.medworm.com/index.php?rid=5520361&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22121 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5520361 Tue, 20 Dec 2011 09:34:41 +0100 5520361 http://www.medworm.com/index.php?rid=5520360&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22120 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5520360 Tue, 20 Dec 2011 09:34:40 +0100 5520360 http://www.medworm.com/index.php?rid=5520359&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22119 AbstractOn the cover: Represented on the cover are the gene expression profiles that reveal the distinct and specific signature of these profiles in two cell types (adipose‐derived myogenic side population cells and musclederivedside population cells). See Andersen et al. in this issue for the complete story.Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5520359 Tue, 20 Dec 2011 09:34:38 +0100 5520359 http://www.medworm.com/index.php?rid=5501100&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21171 In this study, we have employed the novel technique of multispectral imaging flow cytometry to compare the induction and repair of γ‐H2AX foci in three human cell types with different capacities for the repair of DNA double strand breaks (DSB). A repair normal fibroblast cell line MRC5‐SV1, a DSB repair defective ataxia telangiectasia (AT5BIVA) cell line, and a DNA‐PKcs deficient cell line XP14BRneo17 were exposed to 2 Gy gamma radiation from a 60Cobalt source. Thirty minutes following exposure, we observed a dramatic induction of foci in the nuclei of these cells. After 24 hrs, there was a predictable reduction on the number of foci in the MRC5‐SV1 cells, consistent with the repair of DNA DSB. In the AT5BIVA cells, persistence of the foci over a 24‐hr period was due to the fail... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5501100 Tue, 13 Dec 2011 05:00:00 +0100 5501100 http://www.medworm.com/index.php?rid=5501099&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21178 This study is an extension of our previous finding to explore the extracellular bacterial killing, source of DNA in the expelled NETs, their ability to induce proinflammatory cytokines release from platelets/THP‐1 cells, and assessment of NO‐mediated free radical formation by using a consistent NO donor, DETA‐NONOate. NO‐mediated NETs exhibited extracellular bacterial killing as determined by colony forming units. NO‐mediated NETs formation was due to the activation of NADPH oxidase and myeloperoxidase. NO‐ or PMA‐mediated NETs were positive for both nuclear and mitochondrial DNA as well as proteolytic enzymes. Incubation of NETs with human platelets enhanced the release of IL‐1β and IL‐8, while with THP‐1 cells, release of IL‐1β, IL‐8, and TNFα was observed. Thi... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5501099 Tue, 13 Dec 2011 05:00:00 +0100 5501099 http://www.medworm.com/index.php?rid=5501098&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22005 AbstractErythroid biology research involving rhesus macaques has been applied to several topics including malaria, hemoglobinopathy and gene therapy research. However, analyses of the rhesus red blood cells are limited by the inability to identify and sort those cells in research blood samples using flow cytometry. Here it is reported that the BRIC 6 hybridoma clone raised to the human erythroid surface molecule (referred to as CD233, Band 3, AE1, or SLC4A1) produces cross‐reactive and erythroid‐specific antibodies for flow cytometric detection and sorting of rhesus macaque erythrocytes. © 2011 International Society for Advancement of Cytometry (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5501098 Tue, 13 Dec 2011 05:00:00 +0100 5501098 http://www.medworm.com/index.php?rid=5501097&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.22002 AbstractPresence of circulating tumor cells (CTC), as detected by the CellSearch® System, in patients with metastatic carcinomas is associated with poor survival prospects. CellTracks TDI, a dedicated image cytometer, was developed to improve the enumeration of these rare CTC. The CellSearch System was used to enumerate CTC in 7.5 mL blood of 68 patients with cancer and 9 healthy controls. Cartridges containing the fluorescently labeled CTC from this system were reanalyzed using the image cytometer, which acquires images with a TDI camera using a 40×/0.6 NA objective and lasers as light source. Automated classification of events was performed by the Random Forest method using Matlab. An automated classifier was developed to classify events into CTC, apoptotic CTC, CTC debris, leukocytes,... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5501097 Thu, 01 Dec 2011 05:00:00 +0100 5501097 http://www.medworm.com/index.php?rid=5483094&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21167 AbstractAdult zebrafish are being increasingly used as a model in cancer and stem cell research. Here we describe an integrated optical system that combines a laser scanning confocal microscope (LSCM) and an in vivo flow cytometer (IVFC) for simultaneous visualization and cell quantification. The system is set up specifically for non‐invasive tracking of both stationary and circulating cells in adult zebrafish (casper) that have been engineered to be optically transparent. Confocal imaging in this instrument serves the dual purpose of visualizing fish tissue microstructure and an imaging‐based guide to locate a suitable vessel for quantitative analysis of circulating cells by IVFC. We demonstrate initial testing of this novel instrument by imaging the transparent adult zebrafish casper... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5483094 Thu, 01 Dec 2011 05:00:00 +0100 5483094 http://www.medworm.com/index.php?rid=5473658&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21172 In this study, using 5‐ethynyl‐2′deoxyuridine (EdU) as a DNA precursor, we directly assessed the relationship between DNA replication and induction of DSBs revealed as γH2AX foci in A549 cells treated with Top1 inhibitors topotecan (Tpt) or camptothecin (Cpt) and Top2 inhibitors mitoxantrone (Mxt) and etoposide (Etp). Analysis of cells by multiparameter laser scanning cytometry following treatment with Tpt or Cpt revealed that only DNA replicating cells showed induction of γH2AX and a strong correlation between DNA replication and formation of DSBs (r = 0.86). In cells treated with Mxt or Etp, the correlation was weaker (r = 0.52 and 0.64). In addition, both Mtx and Etp caused induction of γH2AX in cells not replicating DNA. Confocal imaging of nuclei of cells treated with Tpt rev... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5473658 Thu, 01 Dec 2011 05:00:00 +0100 5473658 http://www.medworm.com/index.php?rid=5465176&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21173 AbstractClusters of MHCI, ICAM‐1, CD44, CD59, IL‐2R, and IL‐15R molecules have been studied on the surface of CD4+ T‐cells from peripheral blood and lymph nodes of patients in Crohn's disease and healthy individuals as controls by using a dual‐laser flow cytometric fluorescence resonance energy transfer (FRET) technique and fluorescently stained Fabs. When cells from patients in Crohn's disease are compared to those of controls, the surface expression level for the MHCI reduced by ∼45%, for CD44 enhanced by ∼100%, and for IL‐2Rα, IL‐15Rα, and common γc enhanced by ∼50%, ∼70%, and ∼130%, respectively. Efficiencies of FRET monitoring homoassociation for the MHCI and CD44 reduced, that for IL‐2Rα enhanced. While efficiencies of FRET monitoring the association of ... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5465176 Thu, 01 Dec 2011 05:00:00 +0100 5465176 http://www.medworm.com/index.php?rid=5437253&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21175 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5437253 Wed, 23 Nov 2011 09:50:20 +0100 5437253 http://www.medworm.com/index.php?rid=5437252&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21174 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5437252 Wed, 23 Nov 2011 09:50:19 +0100 5437252 http://www.medworm.com/index.php?rid=5437251&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21176 AbstractOn the cover: The cover image depicts the multi‐layered effect generated after the uptake of layer‐by‐layer microcarriers by polymorphonuclear leukocytes. Read the complete article by Rathmann and coworkers in this issue.Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5437251 Wed, 23 Nov 2011 09:50:17 +0100 5437251 http://www.medworm.com/index.php?rid=5396805&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21150 AbstractFluorescence resonance energy transfer (FRET) between fluorescent proteins (FPs) is a powerful method to visualize and quantify protein–protein interaction in living cells. Unfortunately, the emission bleed‐through of FPs limits the usage of this complex technique. To circumvent undesirable excitation of the acceptor fluorophore, using two‐photon excitation, we searched for FRET pairs that show selective excitation of the donor but not of the acceptor fluorescent molecule. We found this property in the fluorescent cyan fluorescent protein (CFP)/yellow fluorescent protein (YFP) and YFP/mCherry FRET pairs and performed two‐photon excited FRET spectral imaging to quantify protein interactions on the later pair that shows better spectral discrimination. Applying non‐negative ... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5396805 Thu, 10 Nov 2011 05:00:00 +0100 5396805 http://www.medworm.com/index.php?rid=5396804&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21165 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5396804 Thu, 10 Nov 2011 05:00:00 +0100 5396804 http://www.medworm.com/index.php?rid=5396808&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21164 AbstractDrosophila embryogenesis is an established model to investigate mechanisms and genes related to cell divisions in an intact multicellular organism. Progression through the cell cycle phases can be monitored in vivo using fluorescently labeled fusion proteins andtime‐lapse microscopy. To measure cellular properties in microscopic images, accurate and fast image segmentation methods are a critical prerequisite. To quantify static and dynamic features of interphase nuclei and mitotic chromosomes, we developed a three‐dimensional (3D) segmentation method based on multiple level sets. We tested our method on 3D time‐series images of live embryos expressing histone‐2Av‐green fluorescence protein. Our method is robust to low signal‐to‐noise ratios inherent to high‐speed im... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5396808 Tue, 08 Nov 2011 05:00:00 +0100 5396808 http://www.medworm.com/index.php?rid=5396807&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21168 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5396807 Tue, 08 Nov 2011 05:00:00 +0100 5396807 http://www.medworm.com/index.php?rid=5396806&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21169 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5396806 Tue, 08 Nov 2011 05:00:00 +0100 5396806 http://www.medworm.com/index.php?rid=5396809&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21157 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5396809 Mon, 07 Nov 2011 05:00:00 +0100 5396809 http://www.medworm.com/index.php?rid=5375331&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21154 In this study, an optimization of calyculin A exposure on chromosome morphology and PCC induction frequency was investigated using a human peripheral blood lymphocyte (PBL) ex vivo irradiation (60Co‐γ rays; ∼0.6 Gy/min; 0–30 Gy) model. Treatment with calyculin A (50 nM) for 15 and 30 min resulted in 11.3 ± 2.7 and 9.9 ± 1.6‐fold increases in the frequency of G2/M‐PCC cells with extended length chromosomes compared with the 60‐min treated group over a broad dose range (0 to 20 Gy), respectively. The G2/M‐PCC scoring index per PCC in 15‐ and 30‐min treated groups was increased by 1.9 ± 0.2 (P = 0.001) and 1.8 ± 0.2 (P = 0.001) compared with the 60‐min treated group over 0–20 Gy, respectively. The G2/M‐PCC efficiency of 30‐min treated group was highest in the th... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5375331 Thu, 03 Nov 2011 04:00:00 +0100 5375331 http://www.medworm.com/index.php?rid=5375330&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21166 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5375330 Thu, 03 Nov 2011 04:00:00 +0100 5375330 http://www.medworm.com/index.php?rid=5375329&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21163 In this study, an aerodynamic particle sizer was used to determine the concentration and aerodynamic diameter (AD) of aerosols produced by a FACS Aria II cell sorter under various conditions. Aerosol containment and evacuation were also evaluated using this novel methodology. The results showed that high concentrations of aerosols in the range of 1–3 μm can be produced in fail mode and that with decreased sheath pressure, aerosol concentration decreased and AD increased. Although the engineering controls of the FACS Aria II for containment were effective, sort chamber evacuation of aerosols following a simulated nozzle obstruction was ineffective. However, simple modifications to the FACS Aria II are described that greatly improved sort chamber aerosol evacuation. The results of this st... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5375329 Thu, 03 Nov 2011 04:00:00 +0100 5375329 http://www.medworm.com/index.php?rid=5375326&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21158 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5375326 Thu, 03 Nov 2011 04:00:00 +0100 5375326 http://www.medworm.com/index.php?rid=5396803&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21170 AbstractMicrobial adhesion is a field of recognized relevance and, as such, an impressive array of tools has been developed to understand its molecular mechanisms and ultimately for its quantification. Some of the major limitations found within these methodologies concern the incubation time, the small number of cells analyzed, and the operator's subjectivity. To overcome these aspects, we have developed a quantitative method to measure yeast cells' adhesion through flow cytometry. In this methodology, a suspension of yeast cells is mixed with green fluorescent polystyrene microspheres (uncoated or coated with host proteins). Within 2 h, an adhesion profile is obtained based on two parameters: percentage and cells‐microsphere population's distribution pattern. This flow cytometry protoco... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5396803 Tue, 01 Nov 2011 04:00:00 +0100 5396803 http://www.medworm.com/index.php?rid=5375325&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21162 AbstractMinimal residual disease (MRD) has emerged as a major prognostic factor for monitoring patients with B‐lineage acute lymphoblastic leukemia (B‐ALL). The quantification of MRD by flow cytometry (FC) is based on the identification of a leukemia‐associated phenotype (LAP). Because phenotypic switch is common during treatment, multiple LAPs must be available and used for MRD detection over time. We evaluated the potential usefulness of CD304 as a new marker for monitoring MRD. CD304 was expressed in 48% of B‐ALL (24/50) with discriminative fluorescence intensity compared with CD304‐negative normal B‐cell precursors (n = 15). The sensitivity of CD304‐based MRD detection reached 10−4, as with some of established LAPs. The stability of CD304 expression evaluated during the... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5375325 Tue, 01 Nov 2011 04:00:00 +0100 5375325 http://www.medworm.com/index.php?rid=5343510&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21156 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5343510 Mon, 24 Oct 2011 01:36:23 +0100 5343510 http://www.medworm.com/index.php?rid=5343509&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21160 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5343509 Mon, 24 Oct 2011 01:36:21 +0100 5343509 http://www.medworm.com/index.php?rid=5343508&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21159 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5343508 Mon, 24 Oct 2011 01:36:20 +0100 5343508 http://www.medworm.com/index.php?rid=5343507&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21161 AbstractOn the cover: Various protist species are major threats to human health. Cytometry can support our battle against them. Attila Tárnok's Editorial highlights the specific approaches from the work of Fischer and Korzeniewski, Aldebert et al., Schuck et al. and Gerena et al. in this issue.Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5343507 Mon, 24 Oct 2011 01:36:18 +0100 5343507 http://www.medworm.com/index.php?rid=5330642&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21155 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5330642 Wed, 19 Oct 2011 20:39:03 +0100 5330642 http://www.medworm.com/index.php?rid=5321512&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21141 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5321512 Thu, 13 Oct 2011 04:00:00 +0100 5321512 http://www.medworm.com/index.php?rid=5310751&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21121 AbstractThe skeletal muscle‐derived side population (mSP) which highly excludes Hoechst 33342 is composed of CD45+ and CD45− subpopulations; yet, rareness of mSP cells in general has complicated extensive quantitative analysis of gene expression profiles in primarily isolated mSP cells. Here, we describe the isolation of adult mouse normal skeletal muscle residing SPCD45+ and SPCD45− cells from a parent mononuclear muscle‐derived cell (MDC) population. Relative quantitative real time PCR (RT‐PCR) of 64 genes revealed that mSPCD45− compared with mSPCD45+ was enriched for cells expressing transcripts associated with endothelial cells, Notch signaling and myogenic precursors. By comparing the mRNA signatures of mSPs with those of adipose tissue‐derived SP populations, a common e... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5310751 Tue, 11 Oct 2011 04:00:00 +0100 5310751 http://www.medworm.com/index.php?rid=5310750&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21149 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5310750 Tue, 11 Oct 2011 04:00:00 +0100 5310750 http://www.medworm.com/index.php?rid=5310749&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21139 We report a new, small particle lyophilized immunogold reagent that maintains activity after 42°C storage for a year and can be rapidly dissolved into stable liquid suspension for use in labelling cells with larger particle aggregates that have enhanced scattering cross section. Labeling requires less than 1 min at 20°C, which is ∼30 times faster than customary fluorescent antibody labeling. The labeling step involves neutralizing the surface charge of immunogold and creating specifically bound aggregates of gold on the cell surface. This process provides distinct side‐scatter cluster separation with blue laser light at 488 nm, which is further improved by using red laser light at 640 nm. Similar comparisons using LED light sources showed less improvement with red light, thereby indi... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5310749 Tue, 11 Oct 2011 04:00:00 +0100 5310749 http://www.medworm.com/index.php?rid=5310748&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21145 In this study, the capability of functionalized CaCO3 microcarriers as AIS transporter system targeted at PMNs is investigated. The time‐dependent interaction of protamine sulfate and dextran sulfate multilayer‐coated 5 μm ± 1 μm CaCO3 carriers with PMNs, in comparison with the usage of SiO2 carriers as monodisperse model system of defined sizes (1, 3, and 5 μm), reveals a sufficient carrier/cell interaction and uptake for coincubation periods between 2 and 24 h. Furthermore, the microcarriers are exposed to an environment simulating primary granule/phagosomal conditions after phagocytosis by means of PMN stimulation. The incubation of CaCO3 microcarriers with cell supernatant demonstrates a partial multilayer decomposition (three to five layers) within 24 h, allowing the gradual r... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5310748 Tue, 11 Oct 2011 04:00:00 +0100 5310748 http://www.medworm.com/index.php?rid=5310747&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21153 AbstractSET/I2PP2A is a nuclear protein that was initially identified as an oncogene in human undifferentiated acute myeloid leukemia, fused to the nuclear porin Nup‐214. In addition, SET is a potent inhibitior of the phosphatase PP2A. Previously, we proposed a model in which the small GTPase Rac1 recruits SET from the nucleus to the plasma membrane to promote cell migration. This event represents an entirely novel concept in the field of cell migration. Now, fluorescent versions of the SET protein are generated to analyze its nucleo‐cytoplasmic shuttling in live cells. Our studies showed that under steady‐state conditions a fraction of the SET protein, which is primarily localized in the nucleus, translocates to the cytosol in an apparently random fashion. SET exiting the nucleus wa... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5310747 Tue, 11 Oct 2011 04:00:00 +0100 5310747 http://www.medworm.com/index.php?rid=5310746&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21151 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5310746 Tue, 11 Oct 2011 04:00:00 +0100 5310746 http://www.medworm.com/index.php?rid=5310745&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21147 AbstractTissue factor (TF)‐positive microparticles (MPs) are highly procoagulant, and linked to thrombosis in sepsis and cancer. MP‐associated TF may be assayed by immunological or functional methods. Several reports have demonstrated discrepancies between TF‐protein and TF‐activity, which have been explained by antibody binding to “encrypted” or degraded forms of inactive TF‐protein. Our goal was to evaluate the possible interference of fluorescent antibody aggregates in solutions containing antibodies against TF and CD14 in flow cytometric analysis. Using monocyte‐derived microparticles (MPs) released from human monocytes, incubated with or without lipopolysaccharides (LPS) in vitro, we measured MP‐associated TF‐protein (flow cytometry) and TF‐activity (clot formati... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5310745 Tue, 11 Oct 2011 04:00:00 +0100 5310745 http://www.medworm.com/index.php?rid=5310744&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21148 AbstractFlow cytometry is a valuable tool in research and diagnostics including minimal residual disease (MRD) monitoring of hematologic malignancies. However, its gradual advancement toward increasing numbers of fluorescent parameters leads to information rich datasets, which are challenging to analyze by standard gating and do not reflect the multidimensionality of the data.We have developed a novel method to analyze complex flow cytometry data, based on hierarchical clustering analysis (HCA) but with a new underlying algorithm, using Mahalanobis distance measure. HCA is scalable to analyze complex multiparameter datasets (here demonstrated on up to 12 color flow cytometry and on a 20‐parameter synthetic dataset).We have validated this method by comparison with standard gating approach... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5310744 Tue, 11 Oct 2011 04:00:00 +0100 5310744 http://www.medworm.com/index.php?rid=5321511&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21122 In this study, several image segmentation methods were therefore compared and evaluated in order to identify the most appropriate segmentation schemes that are usable with little new parameterization and robustly with different types of fluorescence‐stained cells for various biological and biomedical tasks. The study investigated, compared, and enhanced four different methods for segmentation of cultured epithelial cells. The maximum‐intensity linking (MIL) method, an improved MIL, a watershed method, and an improved watershed method based on morphological reconstruction were used. Three manually annotated datasets consisting of 261, 817, and 1,333 HeLa or L929 cells were used to compare the different algorithms. The comparisons and evaluations showed that the segmentation performance ... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5321511 Sat, 01 Oct 2011 04:00:00 +0100 5321511 http://www.medworm.com/index.php?rid=5310743&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21152 AbstractPericyte perivascular cells, believed to originate mesenchymal stem cells (MSC), are characterized by their capability to differentiate into various phenotypes and participate in tissue reconstruction of different organs, including the brain. We show that these cells can be induced to differentiation into neural‐like phenotypes. For these studies, pericytes were obtained from aorta ex‐plants of Sprague–Dawley rats and differentiated into neural cells following induction with trans retinoic acid (RA) in serum‐free defined media or differentiation media containing nerve growth and brain‐derived neuronal factor, B27, N2, and IBMX. When induced to differentiation with RA, cells express the pluripotency marker protein stage‐specific embryonic antigen‐1, neural‐specific p... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5310743 Sat, 01 Oct 2011 04:00:00 +0100 5310743 http://www.medworm.com/index.php?rid=5268627&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21144 AbstractThe analysis of protein‐protein interactions is a key focus of proteomics efforts. The yeast two‐hybrid system (Y2H) has been the most commonly used method in genome‐wide searches for protein interaction partners. However, the throughput of the current yeast two‐hybrid array approach is hampered by the involvement of the time‐consuming LacZ assay and/or the incompatibility of liquid handling automation due to the requirement for selection of colonies/diploids on agar plates. To facilitate large‐scale Y2H assays, we report a novel array approach by coupling a GFP reporter based Y2H system with high throughput flow cytometry that enables the processing of a 96‐well plate in as little as 3 min. In this approach, the yEGFP reporter has been established in both AH109 (MATa... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5268627 Fri, 30 Sep 2011 11:49:22 +0100 5268627 http://www.medworm.com/index.php?rid=5246318&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21146 AbstractThe schematic (principle) of photoacoustic flow cytometry, in which laser‐induced acoustic waves in single circulation tumor cells targeted by nanoparticles directly in the bloodstream are detected with an ultrasound transducer attached to the skin.Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5246318 Fri, 23 Sep 2011 10:34:56 +0100 5246318 http://www.medworm.com/index.php?rid=5233444&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21134 AbstractIn metastasis, the cancer cells that travel through the body are capable of establishing new tumors in locations remote from the site of the original disease. To metastasize, a cancer cell must break away from its tumor and invade either the circulatory or lymphatic system, which will carry it to a new location, and establish itself in the new site. Once in the blood stream, the cancer cells now have access to every portion of the body. Here, we have used the “in vivo flow cytometer” to study if there is any relationship between metastatic potential and depletion kinetics of circulating tumor cells. The in vivo flow cytometer has the capability to detect and quantify continuously the number and flow characteristics of fluorescently labelled cells in vivo. We have improved the c... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5233444 Tue, 13 Sep 2011 04:00:00 +0100 5233444 http://www.medworm.com/index.php?rid=5233443&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21143 AbstractFlow cytometry (FCM) has been a fundamental tool of biological discovery for many years. Invasive extraction of cells from a living organism, however, may lead to changes in cell properties and prevents studying cells in their native environment. These problems can be overcome by use of in vivo FCM, which provides detection and imaging of circulating normal and abnormal cells directly in blood or lymph flow. The goal of this review is to provide a brief history, features, and challenges of this new generation of FCM methods and instruments. Spectrum of possibilities of in vivo FCM in biological science (e.g., cell metabolism, immune function, or apoptosis) and medical fields (e.g., cancer, infection, and cardiovascular disorder) including integrated photoacoustic‐photothermal the... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5233443 Tue, 13 Sep 2011 04:00:00 +0100 5233443 http://www.medworm.com/index.php?rid=5233442&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21124 AbstractInvestigations of rare cell types in peripheral blood samples, such as tumor, fetal, and endothelial cells, represent an emerging field with several potentially valuable medical applications. Peripheral blood is a particularly attractive body fluid for the detection of rare cells as its collection is minimally invasive and can be repeated throughout the course of the disease. Because the number of rare cells in mononuclear cells can be very low (1 in 10 million), a large number of cells must be quickly screened, which places demanding requirements on the screening technology. While enrichment technology has shown promise in managing metastatic disease, enrichment can cause distortions of cell morphology that limit pathological identification, and the enrichment targeting adds addit... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5233442 Tue, 13 Sep 2011 04:00:00 +0100 5233442 http://www.medworm.com/index.php?rid=5204173&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21125 AbstractWe provide an overview of the methods used to label circulating cells for fluorescence detection by in vivo flow cytometry. These methods are useful for cell tracking in small animals without the need to draw blood samples and are particularly useful for the detection of circulating cancer cells and quantification of circulating immune cells. © 2011 International Society for Advancement of Cytometry. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5204173 Thu, 08 Sep 2011 04:00:00 +0100 5204173 http://www.medworm.com/index.php?rid=5204172&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21127 AbstractRecently, photoacoustic (PA) flow cytometry (PAFC) has been developed for in vivo detection of circulating tumor cells and bacteria targeted by nanoparticles. Here, we propose multispectral PAFC with multiple dyes having distinctive absorption spectra as multicolor PA contrast agents. As a first step of our proof‐of‐concept, we characterized high‐speed PAFC capability to monitor the clearance of three dyes (Indocyanine Green [ICG], Methylene Blue [MB], and Trypan Blue [TB]) in an animal model in vivo and in real time. We observed strong dynamic PA signal fluctuations, which can be associated with interactions of dyes with circulating blood cells and plasma proteins. PAFC demonstrated enumeration of circulating red and white blood cells labeled with ICG and MB, respectively, a... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5204172 Thu, 08 Sep 2011 04:00:00 +0100 5204172 http://www.medworm.com/index.php?rid=5204171&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21137 This study was designed to further explore a correlation between these events and DNA replication. Toward this end, we utilized 5‐ethynyl‐2′deoxyuridine (EdU) and the “click chemistry” approach to label DNA during replication, followed by exposure of A549 cells to H2O2. Multiparameter laser scanning cytometric analysis of these cells made it possible to identify DNA replicating cells and directly correlate H2O2‐induced ATM activation and induction of γH2AX with DNA replication on a cell by cell basis. After pulse‐labeling with EdU and exposure to H2O2, confocal microscopy was also used to examine the localization of DNA replication sites (“replication factories”) versus the H2AX phosphorylation sites (γH2AX foci) in nuclear chromatin in an attempt to observe the absence... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5204171 Thu, 08 Sep 2011 04:00:00 +0100 5204171 http://www.medworm.com/index.php?rid=5204170&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21128 AbstractIn vivo flow cytometry has facilitated advances in the ultrasensitive detection of tumor cells, bacteria, nanoparticles, dyes, and other normal and abnormal objects directly in blood and lymph circulatory systems. Here, we propose in vivo plant flow cytometry for the real‐time noninvasive study of nanomaterial transport in xylem and phloem plant vascular systems. As a proof of this concept, we demonstrate in vivo real‐time photoacoustic monitoring of quantum dot‐carbon nanotube conjugates uptake by roots and spreading through stem to leaves in a tomato plant. In addition, in vivo scanning cytometry using multimodal photoacoustic, photothermal, and fluorescent detection schematics provided multiplex detection and identification of nanoparticles accumulated in plant leaves in t... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5204170 Thu, 08 Sep 2011 04:00:00 +0100 5204170 http://www.medworm.com/index.php?rid=5204169&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21140 AbstractStandard multiphoton laser scanning microscopy (MPLSM) has revolutionized our view of physiologic and pathologic processes in living organisms by enlightening different aspects of cellular choreography in immune responses, that is, cellular motility and co‐localization. To understand cellular communication on a molecular level, novel transgenic reporter mice have been generated. In parallel, MPLSM systems have been developed, which make it possible for this technique to be more widely used to address crucial immunological questions. Here, we review the latest progress concerning transgenic mouse technology and multiphoton imaging capacities and discuss further developments which will enable us to visualize all around monitoring and quantification of cellular function at a molecul... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5204169 Thu, 08 Sep 2011 04:00:00 +0100 5204169 http://www.medworm.com/index.php?rid=5204168&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21135 AbstractP‐glycoprotein (P‐gp) is a transmembrane protein that mediates the efflux of innumerous structurally unrelated compounds. It was initially found over‐expressed in tumor cells, associated to a multidrug resistance phenotype (MDR). Then, P‐gp was found constitutively expressed in excretory cells/tissues and in circulating cells, such as lymphocytes. Considering the importance of this transporter in the establishment of therapeutic protocols and the existence of contradictory results, this study aimed at evaluating the influence of aging in the expression and function of P‐gp in human lymphocytes, comparing two different methodologies to assess both parameters. P‐gp activity and expression were evaluated in lymphocytes isolated from whole blood samples of 65 healthy caucas... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5204168 Thu, 08 Sep 2011 04:00:00 +0100 5204168 http://www.medworm.com/index.php?rid=5204167&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21142 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5204167 Thu, 08 Sep 2011 04:00:00 +0100 5204167 http://www.medworm.com/index.php?rid=5233441&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21118 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5233441 Thu, 01 Sep 2011 04:00:00 +0100 5233441 http://www.medworm.com/index.php?rid=5204166&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21138 AbstractToxplasma is a protozoan parasite, which forms persistent cysts in tissues of chronically infected animals and humans. Cysts can reactivate leading to severe pathologies. They also contribute to the transmission of Toxoplasma infection in humans by ingestion of undercooked meat. Classically, the quantification of cyst burden in tissues uses microscopy methods, which are laborious and time consuming. Here, we have developed automated protocols to quantify cysts, based on flow cytometry or high‐throughput microscopy. Brains of rodents infected with cysts of Prugniaud strain were incubated with the FITC‐Dolichos biflorus lectin and analyzed by flow cytometry and high‐throughput epifluorescence microscopy. The comparison of cyst counts by manual epifluorescence microscopy to flow... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5204166 Thu, 01 Sep 2011 04:00:00 +0100 5204166 http://www.medworm.com/index.php?rid=5179214&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21136 AbstractMelatonin and its derivatives modulate the Plasmodium falciparum and Plasmodium chabaudi cell cycle. Flow cytometry was employed together with the nucleic acid dye YOYO‐1 allowing precise discrimination between mono‐ and multinucleated forms of P. falciparum‐infected red blood cell. The use of YOYO‐1 permitted excellent discrimination between uninfected and infected red blood cells as well as between early and late parasite stages. Fluorescence intensities of schizont‐stage parasites were about 10‐fold greater than those of ring‐trophozoite form parasites. Melatonin and related indolic compounds including serotonin, N‐acetyl‐serotonin and tryptamine induced an increase in the percentage of multinucleated forms compared to non‐treated control cultures. YOYO‐1 s... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5179214 Wed, 31 Aug 2011 03:17:46 +0100 5179214 http://www.medworm.com/index.php?rid=5179218&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21120 This study describes the key components and principles involved in building a next‐generation flow cytometer based on a 32‐channel PMT array detector, a phase‐volume holographic grating, and a fast electronic board. The system is capable of full‐spectral data collection and spectral analysis at the single‐cell level. As demonstrated using fluorescent microspheres and lymphocytes labeled with a cocktail of antibodies (CD45/FITC, CD4/PE, CD8/ECD, and CD3/Cy5), the presented technology is able to simultaneously collect 32 narrow bands of fluorescence from single particles flowing across the laser beam in <5 μs. These 32 discrete values provide a proxy of the full fluorescence emission spectrum for each single particle (cell). Advanced statistical analysis has then been performed... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5179218 Mon, 29 Aug 2011 23:00:00 +0100 5179218 http://www.medworm.com/index.php?rid=5179217&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21132 AbstractA number of apoptotic stimuli produce a different response by CD4+ regulatory and effector lymphocytes. So far, little is known concerning the sensitivity of CD4+ regulatory T cells (Treg) to genotoxic agents. Observations from a mouse model suggest that Treg are more resistant to DNA damage compared to CD4+ T effector cells (Teff). By flow cytometry we analysed the apoptotic response to genotoxic stimuli in culture, comparing Treg and Teff. CD4+ regulatory lymphocytes appeared to be more resistant than CD4+ effector lymphocytes. Results of costaining experiments for CD45RA suggest that this dissimilarity is not related to the differentiation to a CD45RA negative phenotype. Further, neither the antiapoptotic protein Bcl‐2 nor Bcl‐xL were found to be expressed in greater amounts... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5179217 Mon, 29 Aug 2011 23:00:00 +0100 5179217 http://www.medworm.com/index.php?rid=5179216&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21126 AbstractElongation measurements of red cells subjected to simple shear flow are usually performed using a single suspending medium (viscosity η0) and varying the mean shear rate . Such data are often plotted versus the shear stress suggesting that the elongation scales with τ. In this work, normal blood samples were tested in a rheoscope varying both η0 and . The ranges of were chosen to restrict the elongation of the red cells to low values where the behavior is dominated by their intrinsic properties. It was found that the elongation scales with with s decreasing from two at η0 = 20 mPas to unity at η0 = 70 mPas. Above η0 = 70 mPas, the elongation is therefore essentially determined by the membrane elasticity alone. A side observation was a large variation of the elongation both... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5179216 Mon, 29 Aug 2011 23:00:00 +0100 5179216 http://www.medworm.com/index.php?rid=5179215&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21133 We present ex vivo and in vivo experimental verifications involving high‐speed PT imaging of RBCs, identification of sickle cells in a mouse model of human sickle cell disease and in vivo monitoring of complex hemorheological changes (e.g., RBC deformability, hematocrit and RBC aggregation). The multi‐parameter platform that integrates PT, PA, and conventional optical techniques has potential for translation to clinical applications using safe, portable, laser‐based medical devices for point‐of‐care screening of disease progression and therapy efficiency. © 2011 International Society for Advancement of Cytometry (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5179215 Mon, 29 Aug 2011 23:00:00 +0100 5179215 http://www.medworm.com/index.php?rid=5168876&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21114 We report the results of the evaluation of a microfabricated filtration device for blood preparation that separates erythrocytes from leukocytes based on their size and mechanical properties. The microfabricated filter evaluated here requires a rapid and simple procedure and results in high leukocytes recovery without introducing bias among the leukocyte subpopulations. The filter removes erythrocytes, platelets, plasma proteins, and unbound staining reagent. This gentle filtration process produces very clean stained leukocytes for cytometric analysis without any apparent damage to leukocytes. © 2011 International Society for Advancement of Cytometry (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5168876 Sun, 28 Aug 2011 01:38:23 +0100 5168876 http://www.medworm.com/index.php?rid=5155623&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21123 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5155623 Thu, 25 Aug 2011 07:38:56 +0100 5155623 http://www.medworm.com/index.php?rid=5155622&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21130 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5155622 Thu, 25 Aug 2011 07:38:55 +0100 5155622 http://www.medworm.com/index.php?rid=5155621&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21129 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5155621 Thu, 25 Aug 2011 07:38:55 +0100 5155621 http://www.medworm.com/index.php?rid=5155620&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21131 AbstractOn the cover: Flow cytometry is of high importance in cancer research, therapy optimization, and individualized diagnosis. It helps to unravel novel signaling cascades among other functions. The authors examine, for example, how cyclopamine modulates ABC transporters expressed in stem cells, indicating the role those transporters play in preserving and protecting stem cells from outside forces (such as oxysterols). The cover depicts how cyclopamine interacts with and effects the various signaling pathways. (See the article by Balbuena et al. in this issue).Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5155620 Thu, 25 Aug 2011 07:38:52 +0100 5155620 http://www.medworm.com/index.php?rid=5168878&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21113 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5168878 Wed, 24 Aug 2011 23:00:00 +0100 5168878 http://www.medworm.com/index.php?rid=5168877&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21119 This study demonstrates that red laser‐based flow cytometry using R800 or LD700 can be used for effective quantification of parasitemia levels in Plasmodium infected red blood cells. Furthermore, this method has the advantage that it does not require RNAse pretreatment and allows for a greater amount of cells to be analyzed for parasite burden than otherwise measured by conventional microscopy. © 2011 International Society for Advancement of Cytometry (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5168877 Wed, 24 Aug 2011 23:00:00 +0100 5168877 http://www.medworm.com/index.php?rid=5137821&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21106 AbstractConventional photothermal (PT) and photoacousic (PA) imaging, spectroscopy, and cytometry are preferentially based on positive PT/PA effects, when signals are above background. Here, we introduce PT/PA technique based on detection of negative signals below background. Among various new applications, we propose label‐free in vivo flow cytometry of circulating clots. No method has been developed for the early detection of clots of different compositions as a source of thromboembolism including ischemia at strokes and myocardial infarction. When a low‐absorbing, platelet‐rich clot passes a laser‐irradiated vessel volume, a transient decrease in local absorption results in an ultrasharp negative PA hole in blood background. Using this phenomenon alone or in combination with pos... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5137821 Sun, 31 Jul 2011 23:00:00 +0100 5137821 http://www.medworm.com/index.php?rid=5068587&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21112 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5068587 Wed, 27 Jul 2011 15:41:32 +0100 5068587 http://www.medworm.com/index.php?rid=5068589&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21110 AbstractA handful of research teams around the world have recently begun to utilize multiphoton techniques in cytometry, especially for in vivo applications. These approaches offer similar enhancements to flow cytometry as the multiphoton phenomenon brought to the field of microscopy at the turn of the 20th century, with at least six advantages over single‐photon excitation. Here, we review the published literature on multiphoton cytometry in vivo or in vitro from the initial experiments in 1999 to present. Multiphoton cytometry instrumentation set‐ups vary from adapted multiphoton microscopy to a dedicated system, with laser pulse power and repetition rate serving as important variables. Two‐beam geometry enables quantitation of cell size. Labeling strategies include conjugated fluo... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5068589 Tue, 26 Jul 2011 23:00:00 +0100 5068589 http://www.medworm.com/index.php?rid=5068588&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21111 AbstractBy using imaging spectrophotometry with paired images in the 200‐ to 280‐nm wavelength range, we have directly mapped intracellular nucleic acid and protein distributions across a population of Chinese hamster ovary (CHO‐K1) cells. A broadband 100× objective with a numerical aperture of 1.2 NA (glycerin immersion) and a novel laser‐induced‐plasma point source generated high‐contrast images with short (∼100 ms) exposures and a lateral resolution nearing 200 nm that easily resolves internal organelles. In a population of 420 CHO‐K1 cells and 477 nuclei, we found a G1 whole‐cell nucleic acid peak at 26.6 pg, a nuclear‐isolated total nucleic acid peak at 11.4 pg, and, as inferred by RNase treatment, a G1 total DNA mass of 7.4 pg. At the G1 peak, we found a whole‐... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5068588 Tue, 26 Jul 2011 23:00:00 +0100 5068588 http://www.medworm.com/index.php?rid=5057249&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21109 AbstractWhile previous studies have investigated the utility of Luminex technology in comparison to other standard techniques, there have been few studies directly comparing different bead‐based assays. A key barrier to establishing Luminex technology in research or clinical laboratories is the apparent need to purchase not only encoded bead sets but also the Luminex instrument. However, as flow cytometry instrumentation continues to improve in sensitivity and in the number and diversity of detection parameters, a diverse range of bead‐based assays is likely to emerge. Human papillomavirus (HPV) genotyping requires multiplexed analysis of 10–100 individual genotypes per sample, which is well suited to bead‐based assays whilst technically challenging and costly for related technolog... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5057249 Sun, 24 Jul 2011 04:35:17 +0100 5057249 http://www.medworm.com/index.php?rid=5048298&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21108 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5048298 Thu, 21 Jul 2011 23:45:41 +0100 5048298 http://www.medworm.com/index.php?rid=5048297&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21116 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5048297 Thu, 21 Jul 2011 23:45:41 +0100 5048297 http://www.medworm.com/index.php?rid=5048296&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21115 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5048296 Thu, 21 Jul 2011 23:45:40 +0100 5048296 http://www.medworm.com/index.php?rid=5048295&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21117 AbstractOn the cover: Like the rare and difficult to cultivate tropical flower Rafflesia arnoldii, the largest flower in the world, so are endothelial progenitor cells extremely rare and difficult to grow. They form one of the largest organs in our bodies: the blood vessels. Now Paprocka and colleagues (in this issue) were successful in generating two stable endothelial progenitor cell lines that provide useful and easy to handle model systems for the study of endothelial differentiation and function.Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5048295 Thu, 21 Jul 2011 23:45:39 +0100 5048295 http://www.medworm.com/index.php?rid=5048293&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21104 AbstractExploring the possibilities offered by flow cytometric microbead analyses for the detection of genetic alterations, an assay based on the dependence of the melting point of double‐stranded DNA molecules on their length has been developed, making use of PCR products carrying biotin and fluorescent moiety on their two ends. The samples of different length PCR products immobilized on streptavidine coated microbeads are heat‐treated in the presence of formamide at temperatures between the melting point of the longer and that of the shorter PCR product, when the mean fluorescence intensity of the beads carrying the shorter molecules decreases as a result of denaturation, as opposed to the sample containing the longer product. The efficacy and sensitivity of the method is demonstrate... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5048293 Thu, 21 Jul 2011 23:45:34 +0100 5048293 http://www.medworm.com/index.php?rid=5057252&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21102 AbstractThe circulating tumor cells (CTCs) appear to be a marker of metastasis development, especially, for highly aggressive and epidemically growing melanoma malignancy that is often metastatic at early stages. Recently, we introduced in vivo photoacoustic (PA) flow cytometry (PAFC) for label‐free detection of mouse B16F10 CTCs in melanoma‐bearing mice using melanin as an intrinsic marker. Here, we significantly improve thespeed of PAFC by using a high‐pulse repetition rate laser operating at 820 and 1064 nm wavelengths. This platform was used in preclinical studies for label‐free PA detection of low‐pigmented human CTCs. Demonstrated label‐free PAFC detection, low level of background signals, and favorable safety standards for near‐infrared irradiation suggest that a fiber... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5057252 Thu, 21 Jul 2011 23:00:00 +0100 5057252 http://www.medworm.com/index.php?rid=5057251&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21105 AbstractDifferential white blood cell count (dWBC) is a frequently used diagnostic tool. For most patient samples an automated blood counter produces a five‐part differential count. If this dWBC does not meet pre‐set criteria, microscopic dWBC is performed. Microscopy is labor intensive and requires sustained training of technicians. Inter‐observer variation and statistical variation are significant, due to limited numbers of cells counted. Flow cytometry is a candidate reference method for dWBC. Advantages are immunological definitions and large number of measured cells. Our goal was to replace (part of) the microscopic dWBC by a flow cytometric dWBC, that gives additional information on blasts, myeloid precursors, and lymphocyte subsets. We designed a cocktail of antibodies (CD4, C... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5057251 Thu, 21 Jul 2011 23:00:00 +0100 5057251 http://www.medworm.com/index.php?rid=5057250&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21107 AbstractOver the last decade, a number of monoclonal antibodies and small molecule inhibitors emerged as potent therapeutic agents in the treatment of Her2/neu overexpressing breast cancer. Numerous patients, however, do not adequately respond to anti‐epidermal growth factor receptor (EGFR)/Her2 receptor targeting. Receptor‐ and, in turn, growth‐stimulating effects, which potentially hamper antiproliferative cell treatment, have barely been investigated. BT474 and SK‐BR‐3 breast cancer cell lines were treated with Trastuzumab, Pertuzumab, and Lapatinib alone using different combinations and concentrations. Moreover, epidermal growth factor (EGF) or heregulin (HRG) was added to reveal potential growth factor‐mediated compensatory effects. Receptor and intracellular signaling wer... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5057250 Thu, 21 Jul 2011 23:00:00 +0100 5057250 http://www.medworm.com/index.php?rid=5048294&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21103 AbstractThe Sonic Hedgehog (Hh) pathway has been implicated in the maintenance of stem or progenitor cells in many adult tissues. Importantly, abnormal Hh pathway activation is also associated with initiation of neoplasia, but its role in tumor growth is still unclear. Here, we demonstrate that cyclopamine, a plant‐derived alkaloid product used to inhibit the Hh signaling pathway, reduces the Side Population (SP) obtained by Hoechst 33342 (Ho342) dye measurements. In addition, cyclopamine is able to modulate, along with oxysterols and other products, the ABCG2 transporter by increasing Ho342 and mitoxantrone uptake. Therefore, if the SP is solely measured as a Ho342 dye extruding fraction, this may be significantly modulated by the inhibition of ABCG2 transport fraction, independently fr... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5048294 Mon, 18 Jul 2011 23:00:00 +0100 5048294 http://www.medworm.com/index.php?rid=5017815&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21095 In this study, we report on the design and performance of a confocal laser based system built to collect backscattered light over a range of 26° at 405, 488, and 633 nm to discriminate leukemic cells from normal red blood cells (RBC) and white blood cells (WBC). The design of the system is based on the spectral differences observed from spectroscopy measurements with a similar system designed with a white light source. Significant differences are observed in the intensity and wavelength dependence of leukemic cells from normal RBC and WBC. Specifically, the distinct light scattering of RBC is due to hemoglobin absorption, allowing for its discrimination from leukemic cells, mononuclear, and polymorphonuclear WBC particularly at certain wavelengths. Meanwhile, the high scattering intensiti... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5017815 Thu, 07 Jul 2011 23:00:00 +0100 5017815 http://www.medworm.com/index.php?rid=5006341&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21073 AbstractMesenchymal stromal cells (MSCs) do not express a unique definite epitope or marker gene. As such, minimal criteria were recently established for defining multipotent MSC. These criteria include expression of CD73, CD90, CD105, and a lack of hematopoietic marker expression. However, we detected binding of a CD14 antibody on bone marrow‐ and placenta‐derived MSC and investigated the staining of CD14 antibodies on these MSC in more detail. The MSC were isolated from human bone marrow and placenta tissue, expanded, characterized by quantitative RT‐PCR, flow cytometry, and immunocytochemistry and differentiated to generate osteoblasts, chondrocytes, and adipocytes. The CD14‐cross‐reactive MSCs were enriched by cell sorting. Human peripheral blood mononuclear cells, fibroblast... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5006341 Tue, 05 Jul 2011 23:00:00 +0100 5006341 http://www.medworm.com/index.php?rid=5017814&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21101 AbstractIntravital microscopy is a valuable tool for research into sickle cell disease with studies being carried out on transgenic mice and human volunteers. The method has helped to develop an explanation for sickle crises based on cell adhesion to the vascular endothelium followed by logjamming of rigid sickle cells and has stimulated much research into new treatments. In recent years there have been numerous new optical techniques developed for imaging the microcirculation and understanding the circulation of cells within the body, many of which have been further developed into in vivo flow cytometry techniques. This brief review highlights some of the progress made to date in the understanding of sickle cell disease using intravital microscopy. New techniques for imaging the microcirc... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5017814 Thu, 30 Jun 2011 23:00:00 +0100 5017814 http://www.medworm.com/index.php?rid=5006340&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21084 AbstractWe have developed a high‐throughput platform to detect the presence of HIV‐1 and SIV‐specific ADCC‐mediating antibody responses. The assay is based on the hydrolysis of a cell‐permeable fluorogenic peptide substrate containing a sequence recognized by the serine protease, Granzyme B (GzB). GzB is delivered into target cells by cytotoxic effector cells as a result of antigen (Ag)‐specific Ab‐Fcγ receptor interactions. Within the target cells, effector cell‐derived GzB hydrolyzes the substrate, generating a fluorescent signal that allows individual target cells that have received a lethal hit to be identified by flow cytometry. Results are reported as the percentage of target cells with GzB activity (%GzB). Freshly isolated or cryopreserved PBMC and/or NK cells can b... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=5006340 Thu, 30 Jun 2011 23:00:00 +0100 5006340 http://www.medworm.com/index.php?rid=4974668&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21097 AbstractZebrafish somitogenesis is governed by a segmentation clock that generates oscillations of gene expression in the zebrafish presomitic mesoderm (PSM) cells. The segmentation clock causes cells to undergo repeated cycles of transcriptional activation and repression, which can be divided into eight phases based on their distinct mRNA co‐localizations. Recognizing different gene oscillation phases of cells is important in zebrafish research, but manual analysis is time‐consuming and difficult. In this article, an effective automated gene oscillation phase classification framework is established for zebrafish PSM cell images. The framework consists of three major steps: (1) identify the individual cells by a two‐stage segmentation procedure; (2) extract multiple features on each ... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4974668 Tue, 28 Jun 2011 18:26:15 +0100 4974668 http://www.medworm.com/index.php?rid=4974670&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21092 AbstractEndothelial progenitor cells (EPCs) modulate postnatal vascularization and contribute to vessel regeneration in adults. Stem cells and progenitor cells were found in umbilical cord blood, bone marrow, and mobilized peripheral blood cells, from where they were isolated and cultured. However, the yield of progenitor cells is usually not sufficient for clinical application and the quality of progenitor cells varies. The aim of the study was the immortalization of early progenitor cells with high proliferative potential, capable to differentiate to EPCs and, further, toward endothelial cells. Two cell lines, namely HEPC‐CB.1 and HEPC‐CB.2 (human endothelial progenitor cells—cord blood) were isolated. As assessed by specific antibody labeling and flow cytometric analysis, they exp... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4974670 Sun, 26 Jun 2011 23:00:00 +0100 4974670 http://www.medworm.com/index.php?rid=4974669&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21094 In conclusion, the two methods were comparable in sensitivity in the studies of IRES mutations and host cell types. We discussed the significance of our findings and potential advantage of the cytometric assay: application to the molecular study of the HCV translation and to screening anti‐IRES drugs. © 2011 International Society for Advancement of Cytometry (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4974669 Sun, 26 Jun 2011 23:00:00 +0100 4974669 http://www.medworm.com/index.php?rid=4967581&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21099 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4967581 Sun, 26 Jun 2011 17:11:29 +0100 4967581 http://www.medworm.com/index.php?rid=4967580&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21098 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4967580 Sun, 26 Jun 2011 17:11:28 +0100 4967580 http://www.medworm.com/index.php?rid=4967579&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21100 AbstractOn the cover: This image of a synthetic three‐dimensional HeLa cell shows the nuclear membrane in red, the cell membrane in blue, and lysosomal membranes in green. It was randomly generated from a model learned from real 3D images using the approach described by Peng and Murphy, Cytometry Part A 79A:383–391, 2011. The generative modeling approach provides a means of aligning data from different sources into a common simulation framework. The presentation generated here provides an artistic interpretation: the computers' eye “sees” the cell in its own way and translates that perception to the scientist.Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4967579 Sun, 26 Jun 2011 17:11:28 +0100 4967579 http://www.medworm.com/index.php?rid=4932255&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21079 This study compares segmentation results from nine different segmentation techniques applied to two different cell lines and five different sets of imaging conditions. Significant variability in the results of segmentation was observed that was due solely to differences in imaging conditions or applications of different algorithms. We quantified and compared the results with a novel bivariate similarity index metric that evaluates the degree of underestimating or overestimating a cell object. The results show that commonly used threshold‐based segmentation techniques are less accurate than k‐means clustering with multiple clusters. Segmentation accuracy varies with imaging conditions that determine the sharpness of cell edges and with geometric features of a cell. Based on this observa... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4932255 Mon, 13 Jun 2011 23:00:00 +0100 4932255 http://www.medworm.com/index.php?rid=4910377&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21076 AbstractCultured mammalian cells [e.g., murine hybridomas, Chinese hamster ovary (CHO) cells] used to produce therapeutic and diagnostic proteins often exhibit increased specific productivity under osmotic stress. This increase in specific productivity is accompanied by a number of physiological changes, including cell size variation. Investigating the cell size variation of hyperosmotically stressed cultures may reveal, in part, the basis for increased specific productivity as well as an understanding of some of the cellular defense responses that occur under hyperosmotic conditions. The regulation of cell volume is a critical function maintained in animal cells. Although these cells are highly permeable to water, they are significantly less permeable to ionic solutes. Appropriate cell–... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4910377 Mon, 06 Jun 2011 23:00:00 +0100 4910377 http://www.medworm.com/index.php?rid=4891572&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21085 AbstractToxicity of engineered nanomaterials (ENMs), such as metal oxides, has been of concern among environmental and health scientists. For ecotoxicity studies of ENMs, it is important to assess nanoparticle uptake and correlate it with the cellular response. However, due to nonavailability of adequate methods for assessing cellular uptake of ENMs, there is a lack of information in this important area. In the present study, a method has been developed using flow cytometry, which allows for rapid detection of ENM internalization in live bacteria under different experimental conditions for several generations. Our data demonstrate significant internalization of Zinc oxide (ZnO) and Titanium (IV) oxide (TiO2) nanoparticles (NPs) in Escherichia coli in a dose‐dependent manner. ZnO NPs trea... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4891572 Wed, 01 Jun 2011 23:00:00 +0100 4891572 http://www.medworm.com/index.php?rid=4891571&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21086 AbstractThe prognostic value of assessing minimal residual disease (MRD) in leukemia has been established with advancements in flow cytometry and PCR. Nonetheless, these techniques are limited by high equipment costs, complex, and costly cell processing and the need for highly trained personnel. Here, we demonstrate the potential of exploiting differences in the relative intensities of backscattered light at three wavelengths to detect the presence of leukemic cells in samples containing varying mixtures of white blood cells (WBCs) and leukemic cells flowing through microfluidic channels. Using 405, 488, and 633 nm illumination, we identify distinct light scattering intensity distributions for Nalm‐6 leukemic cells, normal mononuclear (PBMC) and polymorphonuclear (PMN) white blood cells ... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4891571 Wed, 01 Jun 2011 23:00:00 +0100 4891571 http://www.medworm.com/index.php?rid=4951343&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21096 AbstractFollowing activation by antigen, T cells enter the cell cycle in stages that can be defined with flow cytometric markers. We show that these markers include the increase of forward light scatter width (FSC‐W) signal and the ratio of FSC area/peak. This change in light scatter precedes the first cell division and may reflect blast transformation. We show that the FSC‐W parameter can be used, alone or in combination with other activation markers, to monitor the relative and absolute numbers of T cells responding to a proliferative stimulus. In contrast to dye dilution assays, the FSC‐W method does not allow discrimination between consecutive cell divisions, but it has several advantages and could complement the dye dilution assay. Our findings also show that the routine elimina... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4951343 Tue, 31 May 2011 23:00:00 +0100 4951343 http://www.medworm.com/index.php?rid=4932254&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21093 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4932254 Tue, 31 May 2011 23:00:00 +0100 4932254 http://www.medworm.com/index.php?rid=4920000&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21087 AbstractSpecific single‐molecule detection opens new possibilities in genomics and proteomics, and automated image analysis is needed for accurate quantification. This work presents image analysis methods for the detection and classification of single molecules and single‐molecule interactions detected using padlock probes or proximity ligation. We use simple, widespread, and cost‐efficient wide‐field microscopy and increase detection multiplexity by labeling detection events with combinations of fluorescence dyes. The mathematical model presented herein can classify the resulting point‐like signals in dual‐channel images by spectral angles without discriminating between low and high intensity. We evaluate the methods on experiments with known signal classes and compare to clas... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4920000 Tue, 31 May 2011 23:00:00 +0100 4920000 http://www.medworm.com/index.php?rid=4910376&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21075 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4910376 Tue, 31 May 2011 23:00:00 +0100 4910376 http://www.medworm.com/index.php?rid=4891570&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21091 AbstractAsymmetric cell division is an important mechanism for generating cellular diversity, however, techniques for measuring the distribution of fate‐regulating molecules during mitosis have been hampered by a lack of objectivity, quantitation, and statistical robustness. Here we describe a novel imaging flow cytometric approach that is able to report a cells proliferative history and cell cycle position using dye dilution, pH3, and PI staining to then measure the spatial distribution of fluorescent signals during mitosis using CCD‐derived imagery. Using Jurkat cells, resolution of the fluorescently labeled populations was comparable to traditional PMT based cytometers thus eliminating the need to sort cells with specific division histories for microscopy. Subdividing mitotic stages... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4891570 Tue, 31 May 2011 23:00:00 +0100 4891570 http://www.medworm.com/index.php?rid=4861225&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21083 AbstractIn this article, we demonstrate the potential of a microfluidic chip for the differentiation of immunologically stained blood cells. To this end, white blood cells stained with antibodies typically applied for the determination of the immune status were measured in the micro‐device. Relative concentrations of lymphocytes and subpopulations of lymphocytes are compared to those obtained with a conventional flow cytometer. The stability of the hydrodynamic focusing and the optical setup was determined by measuring the variation of the signal pulse height of fluorescence calibration beads, being about 2% for the micro‐device. This value and the overall performance of the micro‐device are similar to conventional flow cytometers. It follows from our results that such microfluidic s... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4861225 Wed, 25 May 2011 22:57:11 +0100 4861225 http://www.medworm.com/index.php?rid=4839680&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21061 AbstractMitochondria are most important organelles in the survival of eukaryotic aerobic cells because they are the primary producers of ATP, regulators of ion homeostasis or redox state, and producers of free radicals. The key role of mitochondria in the generation of primordial ATP for the survival and proliferation of eukaryotic cells has been proven by extensive biochemical studies. In this context, it is crucial to understand the complexity of the mitochondrial compartment and its functionality and to develop experimental tools allowing the assessment of its nature and its function and metabolism. This review covers the role of the mitochondria in the cell, focusing on its structure, the mechanism of the mitochondrial respiratory chain, the maintenance of the transmembrane potential a... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4839680 Thu, 19 May 2011 19:59:40 +0100 4839680 http://www.medworm.com/index.php?rid=4839679&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21081 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4839679 Thu, 19 May 2011 19:59:38 +0100 4839679 http://www.medworm.com/index.php?rid=4839678&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21089 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4839678 Thu, 19 May 2011 19:59:37 +0100 4839678 http://www.medworm.com/index.php?rid=4839677&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21088 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4839677 Thu, 19 May 2011 19:59:35 +0100 4839677 http://www.medworm.com/index.php?rid=4839676&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21090 AbstractThe cover image is in the style of a painting by the 19th century artist Paul Klee “Fire in the evening” (1929). Somatic cells need for genes to be reprogrammed to iPCS. However, only certain stoichiometric combinations of gene expression (symbolized by the color and size of the bars that represent one of the four different genes) render high yield of pluripotent stem cells. See the accompanying article by Tiemann et al. in this issue.Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4839676 Thu, 19 May 2011 19:59:34 +0100 4839676 http://www.medworm.com/index.php?rid=4839675&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21082 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4839675 Thu, 19 May 2011 19:59:24 +0100 4839675 http://www.medworm.com/index.php?rid=4819659&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21054 AbstractFluorescence Lifetime Imaging Microscopy (FLIM) is a powerful technique to investigate the local environment of fluorophores in living cells. To correctly estimate all lifetime parameters, time domain FLIM imaging requires a high number of photons and consequently long laser exposure times. This is an issue because long exposure times are incompatible with the observation of dynamic molecular events and induce cellular stress. To minimize exposure time, we have developed an original approach that statistically inflates the number of collected photons. Our approach, called Adaptive Monte Carlo Data Inflation (AMDI), combines the well‐known bootstrap technique with an adaptive Parzen kernel. We here demonstrate using both Monte Carlo simulations and live cells that our robust metho... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4819659 Wed, 11 May 2011 23:00:00 +0100 4819659 http://www.medworm.com/index.php?rid=4819658&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21078 AbstractAlthough the frequency and consequence of sperm chromosomal abnormalities are considerable, few epidemiologic studies in large samples have been conducted to investigate etiologic risk factors. This is, in part, attributable to the labor intensive demands of manual sperm fluorescence in situ hybridization (FISH) scoring. As part of an epidemiologic study investigating environmental risk factors for aneuploidy among men attending a hospital‐based fertility clinic, a semi‐automated method of slide scoring was further validated and used to estimate sex chromosome sperm disomy frequency in a large number of samples. Multiprobe FISH for chromosomes X, Y, and 18 was used to determine sex chromosome disomy in sperm nuclei. Semi‐automated scoring methods were used to quantify X disom... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4819658 Wed, 11 May 2011 23:00:00 +0100 4819658 http://www.medworm.com/index.php?rid=4788146&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21070 AbstractSmall multicellular organisms such as nematodes, fruit flies, clawed frogs, and zebrafish are emerging models for an increasing number of biomedical and environmental studies. They offer substantial advantages over cell lines and isolated tissues, providing analysis under normal physiological milieu of the whole organism. Many bioassays performed on these alternative animal models mirror with a high level of accuracy those performed on inherently low‐throughput, costly, and ethically controversial mammalian models of human disease. Analysis of small model organisms in a high‐throughput and high‐content manner is, however, still a challenging task not easily susceptible to laboratory automation. In this context, recent advances in photonics, electronics, as well as material sc... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4788146 Tue, 03 May 2011 23:00:00 +0100 4788146 http://www.medworm.com/index.php?rid=4788145&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21074 AbstractWe instrumentally, theoretically, and experimentally demonstrate a new approach for characterization of nonspherical individual particles from light scattering. Unlike the original optical scheme of the scanning flow cytometer that measures an angle‐resolved scattering corresponding in general to S11 element of the light‐scattering matrix, the modernized instrument allows us to measure the polarized light‐scattering profile of individual particles simultaneously. Theoretically, the polarized profile is expressed by the combination of a few light‐scattering matrix elements. This approach supports us with additional independent data to characterize a particle with a complex shape and an internal structure. Applicability of the new method was demonstrated from analysis of poly... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4788145 Tue, 03 May 2011 23:00:00 +0100 4788145 http://www.medworm.com/index.php?rid=4819657&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21077 AbstractTrypanosoma cruzi is the protozoan that causes Chagas disease. It divides in the insect vector gut or in the cytosol of an infected mammalian cell. T. cruzi has one mitochondrion, one Golgi complex, one flagellum, and one cytostome. Here, we provide three‐dimensional (3D) models of this protozoan based on images obtained from serial sections on electron microscopy at different stages of the cell cycle. Ultrathin serial sections were obtained from Epon™ embedded parasites, photographed in a transmission electron microscope, and 3D models were generated using Reconstruct and Blender 3D modeling softwares. The localization and distribution of organelles was evaluated and attributed to specific morphological patterns and deduced by distribution of specific markers by immunofluoresc... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4819657 Sat, 30 Apr 2011 23:00:00 +0100 4819657 http://www.medworm.com/index.php?rid=4788144&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21072 AbstractSomatic cells can be reprogrammed toward pluripotency by overexpression of a set of transcription factors, yielding induced pluripotent stem cells (iPSCs) with features similar to embryonic stem cells. Little is known to date about stoichiometric requirements of the individual reprogramming factors (RFs) for efficient reprogramming and especially about whether stoichiometry also influences the quality of derived iPSCs. To address this important issue, we chose bicistronic lentiviral vectors coexpressing fluorescent reporters (eGFP, dTomato, Cerulean, or Venus) along with the canonical RFs to transduce a bulk of murine embryonic fibroblasts (MEFs). Using a flow cytometric approach, we were able to independently and proportionally quantify all fluorophores in multiple‐infected MEFs... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4788144 Sat, 30 Apr 2011 23:00:00 +0100 4788144 http://www.medworm.com/index.php?rid=4767503&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21071 AbstractCell phenotyping and cell cycle analysis are two commonly used assays in both clinical diagnosis and biomedical research. Cell phenotyping by identifying different biomarkers is essential for the diagnosis of hematologic malignancy, sub‐classifying diseases, monitoring response to treatment, predicting prognosis, detecting rare cell populations and residual malignant cells. Cell cycle analysis distinguishes cells in different phases of cell cycle and is often used to determine the cellular response to drugs and biological stimulations. These assays have been traditionally carried out by sensitive fluorescence detection methods such as flow cytometry and laser scanning cytometry for fluorescence‐based cell population analysis. However, these instruments remain relatively expensi... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4767503 Sat, 30 Apr 2011 11:11:39 +0100 4767503 http://www.medworm.com/index.php?rid=4752929&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21068 AbstractThe nuclear factor kappa B (NF‐κB) pathway, which regulates many cellular processes including proliferation, apoptosis, and survival, has emerged as an important therapeutic target in cancer. Activation of the NF‐κB transcription factor is associated with nuclear translocation of the p65 component of the complex. Conventional methods employed to determine nuclear translocation of NF‐κB either lack statistical robustness (microscopy) or the ability to discern heterogeneity within the sampled populations (Western blotting and Gel Shift assays). The ImageStream platform combines the high image content information of microscopy with the high throughput and multiparameter analysis of flow cytometry which overcomes the aforementioned limitations of conventional assays. It is dem... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4752929 Wed, 27 Apr 2011 00:21:29 +0100 4752929 http://www.medworm.com/index.php?rid=4752930&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21067 AbstractMammalian spermatogenesis is still nowadays poorly understood at the molecular level. Testis cellular heterogeneity is a major drawback for spermatogenic gene expression studies, especially when research is focused on stages that are usually very short and poorly represented at the cellular level such as initial meiotic prophase I (i.e., leptotene [L] and zygotene [Z]). Presumably, genes whose products are involved in critical meiotic events such as alignment, pairing and recombination of homologous chromosomes are expressed during the short stages of early meiotic prophase. Aiming to characterize mammalian early meiotic gene expression, we have found the guinea pig (Cavia porcellus) as an especially attractive model. A detailed analysis of its first spermatogenic wave by flow cyto... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4752930 Sun, 24 Apr 2011 23:00:00 +0100 4752930 http://www.medworm.com/index.php?rid=4744638&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21069 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4744638 Sun, 24 Apr 2011 04:38:18 +0100 4744638 http://www.medworm.com/index.php?rid=4744637&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21080 AbstractOn the cover: A top down view of The City of Firsts, where the red flag designates the riches of cytometry and related technologies from signaling in single cells, multiphoton flow cytometry, and nanocytometry to image analysis, microfabrication devices for cellular analysis, and stem cell processing or identification—that will provide even greater richness to the Charm City during CYTO 2011.Cover design by Bärbel Beran [www.beran‐design.de]. (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4744637 Sun, 24 Apr 2011 04:38:13 +0100 4744637 http://www.medworm.com/index.php?rid=4682843&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21056 (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4682843 Tue, 05 Apr 2011 23:00:00 +0100 4682843 http://www.medworm.com/index.php?rid=4682842&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21066 AbstractGiven the importance of subcellular location to protein function, computational simulations of cell behaviors will ultimately require the ability to model the distributions of proteins within organelles and other structures. Toward this end, statistical learning methods have previously been used to build models of sets of two‐dimensional microscope images, where each set contains multiple images for a single subcellular location pattern. The model learned from each set of images not only represents the pattern but also captures the variation in that pattern from cell to cell. The models consist of sub‐models for nuclear shape, cell shape, organelle size and shape, and organelle distribution relative to nuclear and cell boundaries, and allow synthesis of images with the expectat... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4682842 Tue, 05 Apr 2011 23:00:00 +0100 4682842 http://www.medworm.com/index.php?rid=4713992&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21064 AbstractImproving access to CD4 testing in resource‐limited settings can be achieved through both centralized and decentralized testing networks. Decentralized testing models are more suitable for countries where the HIV epidemic affects a large portion of rural populations. Timely access to accurate CD4 results is crucial at the primary level of the health system. For the past 7 years, the Institute of Human Virology of the University of Maryland School of Medicine has implemented a flexible and sustainable three‐phase model: (1) site assessment and improvement, (2) appropriate technology selection with capacity building through practical training and laboratory mentoring, and (3) quality management system strengthening and monitoring, to support accessibility to reliable CD4 counting... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4713992 Thu, 31 Mar 2011 23:00:00 +0100 4713992 http://www.medworm.com/index.php?rid=4708659&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21063 AbstractDevelopment of a microfluidic device is generally based on fabrication‐design‐fabrication loop, as, unlike the microelectronics design, there is no rigorous simulation‐based verification of the chip before fabrication. This usually results in extremely long, and hence expensive, product development cycle if micro/nano fabrication facilities are used from the beginning of the cycle. Here, we illustrate a novel approach of device prototyping that is fast, cheap, reliable, and most importantly, this technique can be adopted even if no state‐of‐the‐art microfabrication facility is available. A water‐jet machine is used to cut the desired microfluidic channels into a thin steel plate which is then used as a template to cut the channels into a thin sheet of a transparent an... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4708659 Thu, 31 Mar 2011 23:00:00 +0100 4708659 http://www.medworm.com/index.php?rid=4703548&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21062 AbstractConventional compensation of flow cytometry (FMC) data of an N‐stained sample requires additional data sets, of N single‐stained control samples, to estimate the spillover coefficients. Single‐stained controls however are the least rigorous controls because any of the multi‐stained controls are closer to the N‐stained sample. In this article, a new, optimization based, compensation method has been developed that is able to use not only single‐ but also multi‐stained controls to improve estimates of the spillover coefficients. The method is demonstrated on a data set from five‐stained dentritic cells (DCs) with five single‐stained and eight multi‐stained controls. This approach is practical and leads to significant improvements in FCM compensation. © 2011 Intern... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4703548 Thu, 31 Mar 2011 23:00:00 +0100 4703548 http://www.medworm.com/index.php?rid=4682841&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21049 AbstractThe aim of this study was to evaluate the diagnostic value of monitoring CD64 antigen upregulation on polymorphonuclear neutrophils (PMN) for the identification of infectious complications in the postoperative course of solid organ transplanted patients. Twenty‐five kidney, 13 liver, and four pancreas–kidney transplanted patients were included. Beginning with preoperative values up to postoperative values after 3 months for each patient, the PMN CD64 Index, HLA‐DR on monocytes, NKp44+ NK and NK/T cells, CXCR3+ NK cells, CXCR3+ T helper cells, CXCR3+ NK/T cells, and CD4/CD8 ratio were measured by flow cytometry. Subsequently they were correlated with confirmed postoperative complications. Measuring the PMN CD64 Index reached a sensitivity of 89% and a specificity of 65% in the... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4682841 Thu, 31 Mar 2011 23:00:00 +0100 4682841 http://www.medworm.com/index.php?rid=4676014&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21065 In conclusion, CK7 targeting raises diagnostic efficiency of Urovysion, and could be an ideal tool for identifying tumor cells in ambiguous cases or when other tumors are present. Statistical evaluation produces accuracy comparable with results of conventional evaluation, and with laboratories setting cut‐offs individually but according harmonized protocol, it could aid method standardization. Furthermore, by providing additional quantitative information about tumor characteristics, is likely to have therapy relevant value in the future. © 2011 International Society for Advancement of Cytometry (Source: Cytometry Part A) Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4676014 Thu, 31 Mar 2011 23:00:00 +0100 4676014 http://www.medworm.com/index.php?rid=4669555&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21045 We report a novel strategy of time‐gated luminescent scanning for accurate counting of rare‐event cells, which exploits the large difference in luminescence lifetimes between the lanthanide biolabels, >100 μs, and the autofluorescence backgrounds, <0.1 μs, to render background autofluorescence invisible to the detector. Rather than having to resort to sophisticated imaging analysis, the background‐free feature allows a single‐element photomultiplier to locate rare‐event cells, so that requirements for data storage and analysis are minimized to the level of image confirmation only at the final step. We have evaluated this concept in a prototype instrument using a 2D scanning stage and applied it to rare‐event Giardia detection labeled by a europium complex. For a slide a... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4669555 Thu, 31 Mar 2011 23:00:00 +0100 4669555 http://www.medworm.com/index.php?rid=4669554&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21050 AbstractFluorescent labeled monoclonal antibodies (mAbs) against CD36 are routinely used as monocyte, erythroid, or platelet markers in clinical cytometry. CD36 has recently been proposed by various authors as a valuable marker helping to enumerate leukocyte's subpopulations by flow cytometry. However, it is known that binding of CD36 may induce platelet activation and formation of platelet's rosettes on leukocytes, resulting in false expression of platelet markers on white blood cells. To study this phenomenon, we have combined classical flow cytometry and a new quantitative flow imaging technique with the ImageStream® analyzer. We show that CD36 ligation induces activation of platelets with CD62 expression and their adhesion on leukocytes due to CD62 and CD162 interactions. Preincubatio... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4669554 Thu, 31 Mar 2011 23:00:00 +0100 4669554 http://www.medworm.com/index.php?rid=4669553&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21060 AbstractBy virtue of superior preservation of proteins and nucleic acids the zinc salt‐based fixatives (ZBF) has been proposed as an alternative to precipitants and cross‐linking fixatives in histopathology. It was recently reported that ZBF is compatible with analysis of cell surface immunophenotype and detection of intracellular epitopes by flow cytometry. The aim of this study was to explore whether ZBF is also compatible with the detection of DNA damage response assessed by phospho‐specific antibodies (Abs) detecting phosphorylation of the key proteins of that pathway. DNA damage in human pulmonary adenocarcinoma A549 cells was induced by treatment with the DNA topoisomerase I inhibitor camptothecin and phosphorylation of histone H2AX on Ser139 (γH2AX) and of ATM on Ser1981 was ... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4669553 Thu, 31 Mar 2011 23:00:00 +0100 4669553 http://www.medworm.com/index.php?rid=4651982&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21055 AbstractThe spatiotemporal dynamics of protein complexes and genome loci are functionally linked to cellular health status. To study the inherent motion of subnuclear particles, it is essential to remove any superimposed component stemming from displacement and deformation of the nucleus. In this article, we propose a mapping of the nuclear interior, which is based on the deformation of the nuclear contour and has no shape constraints. This registration procedure enabled an accurate estimation of telomere mobility in living human cells undergoing dramatic nuclear deformations. Given the large variety of pathologies and cellular processes that are associated with strong nuclear shape changes, the contour mapping algorithm has generic value for improving the accuracy of mobility measurements... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4651982 Wed, 30 Mar 2011 03:26:07 +0100 4651982 http://www.medworm.com/index.php?rid=4651984&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21044 AbstractCytometric methodologies are becoming increasingly important in veterinary andrology as means of assessing sperm function. However, as yet, flow cytometric techniques in veterinary andrology have not kept up in sophistication with those in other areas of biology and medicine. In this brief review, we consider the present state of cytometry in andrological procedures for evaluating the fertility of domestic animal sires. We outline the aspects of sperm physiology, paying particular attention to the changes that take place during the process known as capacitation, which prepares the sperm for interaction with the egg. We then examine briefly but critically the cytometric techniques that are currently in commercial use or are being established in research laboratories for testing sper... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4651984 Mon, 28 Mar 2011 23:00:00 +0100 4651984 http://www.medworm.com/index.php?rid=4651983&cid=s_33764_67_f&fid=33764&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fcyto.a.21052 AbstractLanthanide bioprobes offer a number of novel advantages for advanced cytometry, including the microsecond luminescence lifetime, sharp spectral emission, and large stokes shift. However, to date, only the europium‐based bioprobes have been broadly studied for time‐gated luminescence cell imaging, though a wide range of efficient terbium bioprobes have been synthesized and some of them are commercially available. We analyze that the bottleneck problem was due to the lack of an efficient microscope with pulsed excitation at wavelengths of 300–330 nm. We investigate a recently available 315 nm ultraviolet (UV) light emitting diode to excite an epifluorescence microscope. Substituting a commercial UV objective (40×), the 315 nm light efficiently delivered the excitation light on... Cytometry Part A journals http://www.medworm.com/rss/comments.php?id=4651983 Mon, 28 Mar 2011 23:00:00 +0100 4651983