Journal of Biophotonics via MedWorm.com

Cell morphology and intracellular ionic homeostasis explored with a multimodal approach combining epifluorescence and digital holographic microscopy

Journal of Biophotonics — Fri, 19 Mar 2010 00:00:00 +0100

The authors have developed a live-cell multimodality microscope combining epifluorescence with digital holographic microscopy; it has been implemented with a decoupling procedure allowing to separately measure from the quantitative phase important cell parameters including absolute volume, shape and integral intracellular refractive index. In combination with the numerous different specific fluorescent cellular probes, this multimodality microscopy can address important issues in cell biology. This is demonstrated by the study of intracellular calcium homeostasis associated with the change in cell volume, which play a critical role in the excitotoxicity-induced neuronal death. (© 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Real time characterization of hydrodynamics in optically trapped networks of micro-particles

Journal of Biophotonics — Thu, 18 Mar 2010 00:00:00 +0100

The hydrodynamic interactions of micro-silica spheres trapped in a variety of networks using holographic optical tweezers are measured and characterized in terms of their predicted eigenmodes. The characteristic eigenmodes of the networks are distinguishable within 20-40 seconds of acquisition time. Three different multi-particle networks are considered; an eight-particle linear chain, a nine-particle square grid and, finally, an eight-particle ring. The eigenmodes and their decay rates are shown to behave as predicted by the Oseen tensor and the Langevin equation, respectively. Finally, we demonstrate the potential of using our micro-ring as a non-invasive sensor to the local environmental viscosity, by showing the distortion of the eigenmode spectrum due to the proximity of a planar boun...

Optical micromanipulations in the non-diffractive regime

Journal of Biophotonics — Wed, 17 Mar 2010 00:00:00 +0100

The ever-evolving topic of optical micromanipulation has established itself as a discipline over the last three decades, and is of much interest to a wide research community due to constantly emerging new applications across the various key disciplines. Performing optical manipulation using evanescent waves is termed near-field optical manipulation, which is essentially the manipulation of particles in the non-diffractive regime. The concept of the breaking of diffraction limit is the spur driving near-field optics studies, as opposed to all far field optical applications where light cannot be focused to a spot smaller than the diffraction limited value, which is about half the wavelength of light in the medium. The authors present a review of the various near-field optical manipulation te...

Manipulating CD4+ T cells by optical tweezers for the initiation of cell-cell transfer of HIV-1

Journal of Biophotonics — Wed, 17 Mar 2010 00:00:00 +0100

Cell-cell interactions through direct contact are very important for cellular communication and coordination - especially for immune cells. The human immunodeficiency virus type I (HIV-1) induces immune cell interactions between CD4+ cells to shuttle between T cells via a virological synapse. A goal to understand the process of cell-cell transmission through virological synapses is to determine the cellular states that allow a chance encounter between cells to become a stable cell-cell adhesion. We demonstrate the use of optical tweezers to manipulate uninfected primary CD4+ T cells near HIV Gag-iGFP transfected Jurkat T cells to probe the determinants that induce stable adhesion. When combined with fast 4D confocal fluorescence microscopy, optical tweezers can be utilized not only to faci...

Optofluidic chip for single cell trapping and stretching fabricated by a femtosecond laser

Journal of Biophotonics — Wed, 17 Mar 2010 00:00:00 +0100

The authors present the design and optimization of an optofluidic monolithic chip, able to provide optical trapping and controlled stretching of single cells. The chip is fabricated in a fused silica glass substrate by femtosecond laser micromachining which can produce both optical waveguides and microfluidic channels with great accuracy. A new fabrication procedure adopted in this work allows the demonstration of microchannels with a square cross-section, thus guaranteeing an improved quality of the trapped cell images. Femtosecond laser micromachining emerges as a promising technique for the development of multifunctional integrated biophotonic devices that can be easily coupled to a microscope platform, thus enabling a complete characterization of the cells under test. (© 2010 WILEY-VC...

Comparison of broadband and ultrabroadband pulses at MHz and GHz pulse-repetition rates for nonlinear femtosecond-laser scanning microscopy

Journal of Biophotonics — Thu, 11 Mar 2010 00:00:00 +0100

Nonlinear optical imaging of human skin and of polychromatic microspheres was carried out to compare and evaluate the imaging properties of three different excitation femtosecond lasers: a spectrally tunable 80 MHz Ti: sapphire oscillator that produced 100 fs pulses (spectral width [sim]10 nm) and two ultrabroadband Ti: sapphire oscillators with repetition rates of 85 MHz and 1 GHz. The latter of these two and the 100 fs laser were combined with a laser scanning microscope (TauMap). The intensities of images of the polychromatic microsphere samples obtained with both lasers are in accordance with the usual dependence of two-photon processes on laser pulse parameters, i.e. the intensity is proportional to the square of the mean laser power and the reciprocal pulse duration. In contrast to t...

A Telemetric light delivery system for metronomic photodynamic therapy (mPDT) in rats

Journal of Biophotonics — Wed, 10 Mar 2010 00:00:00 +0100

Light delivery and monitoring during photodynamic therapy (PDT) is often limited by the need for a physical link between the light source, detectors and the treatment volume. This paper reports on the first in vivo experiments performed with a fully implantable telemetric system, designed for a rat glioblastoma model. In this system, light delivery is performed using a solid state optode containing 2 LEDs, and 4 photodiodes which will be used to monitor light delivery in future experiments. Powering and communication is achieved by means of an inductive link. The implant may remain in the animal for extended time periods, making it particularly interesting for performing metronomic PDT. In this paper, we demonstrate the feasibility of in vivo light delivery and biocompatibility of the devi...

Explicit dosimetry for photodynamic therapy: macroscopic singlet oxygen modeling

Journal of Biophotonics — Wed, 10 Mar 2010 00:00:00 +0100

Singlet oxygen (1O2) is the major cytotoxic agent responsible for cell killing for type-II photodynamic therapy (PDT). An empirical four-parameter macroscopic model is proposed to calculate the "apparent reacted 1O2 concentration", [1O2]rx, as a clinical PDT dosimetry quantity. This model incorporates light diffusion equation and a set of PDT kinetics equations, which can be applied in any clinical treatment geometry. We demonstrate that by introducing a fitting quantity "apparent singlet oxygen threshold concentration" [1O2]rx, sd, it is feasible to determine the model parameters by fitting the computed [1O2]rx to the Photofrin-mediated PDT-induced necrotic distance using interstitially-measured Photofrin concentration and optical properties within each mouse. After determining the model ...

A helpful technology - the luminescence detection of singlet oxygen to investigate photodynamic inactivation of bacteria (PDIB)

Journal of Biophotonics — Wed, 10 Mar 2010 00:00:00 +0100

Photodynamic inactivation of bacteria (PDIB) is considered a new approach for the struggle against multiresistant bacteria. To achieve a sufficient level of bacteria killing, the photosensitizer must attach to and/or penetrate the bacteria and generate a sufficiently high amount of singlet oxygen. To optimize PDIB, the direct detection and quantification of singlet oxygen in bacteria is a helpful tool. Singlet-oxygen luminescence is a very weak signal, in particular in living bacteria. We first performed experiments in aqueous photosensitizer solution to optimize the luminescence system. We eliminated non-singlet-oxygen photons, which is important for the quantification of singlet oxygen and its rise and decay rates. This procedure is even more important when the laser excitation beam is s...

Pollen characterization and identification by elastically scattered light

Journal of Biophotonics — Sat, 06 Mar 2010 00:00:00 +0100

The authors recorded the elastic light-scattering pattern of pollen over a large spatial angle range to investigate the potential light scattering for pollen identification. The scattering from elm, hazel, birch, chestnut, willow, sunflower, ragweed and pine was measured. The scattering patterns show distinct differences that can be used for the classification of pollen with simple algorithms. (© 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

High-resolution tomographic diffractive microscopy of biological samples

Journal of Biophotonics — Fri, 05 Mar 2010 00:00:00 +0100

The authors have developed a tomographic diffractive microscope that combines microholography with illumination from an angular synthetic aperture. It images specimens relative to their complex index of refraction distribution (index and absorption) and permits imaging of unlabelled specimens, with high lateral resolution. The authors now study its use for biological applications, and imaged several preparations with fluorescence confocal microscopy and tomographic diffractive microscopy. The results highlight some interesting features of this instrument, which should attract the interest of biologists for this new technique. (© 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Quantitative cell bioimaging using gold-nanoshell conjugates and phage antibodies

Journal of Biophotonics — Mon, 01 Mar 2010 00:00:00 +0100

The authors describe a quantitative evaluation of the efficacy of cell labeling with plasmon-resonant light-scattering nanoparticles used as contrast agents for dark-field microscopy imaging. The experimental model is based on the biospecific labeling of pig embryo kidney (SPEV) cells with primary phage antibodies, followed by the dark-field microscopic visualization using conjugates of silica/gold nanoshells with secondary rabbit antiphage antibodies. To quantify nanoparticle binding, the authors introduce the labeling-efficacy factor (LEF) which is equal to the ratio of the bound-particle pixels per cell to the total number of pixels occupied by the cell. The LEF is calculated by an imaging-analysis algorithm based on the freely available ImageJ Java-based processing code. In terms of th...

Gold nanorods as new nanochromophores for photothermal therapies

Journal of Biophotonics — Mon, 01 Mar 2010 00:00:00 +0100

Results and perspectives on the biomedical exploitation of gold nanorods with plasmon resonances in the near infrared window are reported. The authors describe experimental studies of laser-activated nanoparticles in the direct welding of connective tissues, which may become a valuable technology in biomedicine. In particular, colloidal gold nanorods excited by diode laser radiation at 810 nm were used to mediate functional photothermal effects and weld eye's lens capsules and arteries. The preparation of biopolymeric matrices including gold nanorods is also described, as well as preliminary tests for their application in the closure of wounds in vessels and tendons. Finally, the use of these nanoparticles for future applications in the diagnosis, imaging and therapy of cancer is discussed...

Thermal model for optimization of vascular laser tissue soldering

Journal of Biophotonics — Mon, 01 Mar 2010 00:00:00 +0100

Laser tissue soldering (LTS) is a promising technique for tissue fusion based on a heat-denaturation process of proteins. Thermal damage of the fused tissue during the laser procedure has always been an important and challenging problem. Particularly in LTS of arterial blood vessels strong heating of the endothelium should be avoided to minimize the risk of thrombosis. A precise knowledge of the temperature distribution within the vessel wall during laser irradiation is inevitable. The authors developed a finite element model (FEM) to simulate the temperature distribution within blood vessels during LTS. Temperature measurements were used to verify and calibrate the model. Different parameters such as laser power, solder absorption coefficient, thickness of the solder layer, cooling of the...

Optical micro-angiography images structural and functional cerebral blood perfusion in mice with cranium left intact

Journal of Biophotonics — Tue, 23 Feb 2010 00:00:00 +0100

Alteration in regional cerebral blood flow (CBF) is the direct result of changes in neuronal activity. It is crucial to monitor the spatio-temporal characteristics of cerebro-vascular blood perfusion in the studies of cerebral diseases. Optical micro-angiography (OMAG) is a recently developed imaging technique capable of resolving 3D distribution of dynamic blood perfusion at a capillary level resolution within microcirculatory beds in vivo. The authors report the applications of OMAG in mouse ischemic stroke model. The study demonstrates that OMAG is a useful method capable of providing in vivo serial assessment of 3D cerebro-vascular pathophysiology with high sensitivity, and therefore, has the potential for use in the study of brain disorders and repairs. (© 2010 WILEY-VCH Verlag GmbH ...

Site-specific opening of the blood-brain barrier

Journal of Biophotonics — Tue, 16 Feb 2010 00:00:00 +0100

The blood-brain barrier (BBB) poses a significant impediment for the delivery of therapeutic drugs into the brain. This is particularly problematic for the treatment of malignant gliomas which are characterized by diffuse infiltration of tumor cells into normal brain where they are protected by a patent BBB. Selective disruption of the BBB, followed by administration of anti-cancer agents, represents a promising approach for the elimination of infiltrating glioma cells. A summary of the techniques (focused ultrasound, photodynamic therapy and photochemical internalization) for site-specific opening of the BBB will be discussed in this review. Each approach is capable of causing localized and transient opening of the BBB with minimal damage to surrounding normal brain as evidenced from magn...

Stretching single DNA molecules to demonstrate high-force capabilities of holographic optical tweezers

Journal of Biophotonics — Fri, 12 Feb 2010 00:00:00 +0100

The well calibrated force-extension behaviour of single double-stranded DNA molecules was used as a standard to investigate the performance of phase-only holographic optical tweezers at high forces. Specifically, the characteristic overstretch transition at 65 pN was found to appear where expected, demonstrating (1) that holographic optical trap calibration using thermal fluctuation methods is valid to high forces; (2) that the holographic optical traps are harmonic out to >250 nm of 2.1 [mu]m particle displacement; and (3) that temporal modulations in traps induced by the spatial light modulator (SLM) do not affect the ability of optical traps to hold and steer particles against high forces. These studies demonstrate a new high-force capability for holographic optical traps achievable by ...

Determination of the influence of IR radiation on the antioxidative network of the human skin

Journal of Biophotonics — Thu, 11 Feb 2010 00:00:00 +0100

Production of free radicals in the human skin subsequent to IR irradiation has been demonstrated by means of two different methods. The first technique, based on resonance Raman spectroscopy, enables the non-invasive measurements of the kinetics of cutaneous carotenoid antioxidants beta-carotene and lycopene, subsequent to IR irradiation. Obtained degradation of the cutaneous carotenoids was a hint but not evidence that IR irradiation can produce free radicals in the skin. Therefore, the direct observation sustaining the production of free radicals subsequent to IR irradiation in the skin was performed in-vitro by electron paramagnetic resonance spectroscopy. Enzymatic processes as well as heat shock-induced radicals in the human skin are presumably involved in the energy transfer from IR ...

Endovenous laser therapy - application studies and latest investigations

Journal of Biophotonics — Thu, 11 Feb 2010 00:00:00 +0100

Endovenous laser therapy (ELT) was introduced in clinical practice as a therapy for incompetent veins about ten years ago. One characteristic of ELT is the broad spectrum of different treatment protocols by means of a variety of laser systems as well as manifold application forms and dosimetry concepts are under investigations. Clinical results with effective, relatively pain-free occlusion of incompetent varicosis veins have been observed, as well as undesired side effects such as ecchymosis, phlebitis and recanalization. In recent years systematic experimental investigations and the analysis of clinical results have increased the understanding of the interrelation between the clinical and physical aspects, followed to a continuous optimization of ELT. The use of IR wavelengths and radial...

PALM imaging and cluster analysis of protein heterogeneity at the cell surface

Journal of Biophotonics — Thu, 11 Feb 2010 00:00:00 +0100

The authors employed photoactivatable localization microscopy (PALM) and direct stochastic optical reconstruction microscopy (dSTORM) imaging and image analysis based on Ripley's K -function to quantify the distribution and heterogeneity of proteins at the cell plasma membrane. The membrane targeting sequence of the N-terminal region of the T cell receptor-pathway kinase Lck fused to the photo-convertible fluorescent protein tdEos (LckN10-tdEos), clusters into sub-100 nm regions which cover [sim]7% of the cell surface. 2-channel PALM imaging of LckN10-tdEos and the N-terminus of the kinase Src (SrcN15-PS-CFP2) are demonstrated. Finally, T cell microclusters at the immune synapse are imaged at super-resolution using dSTORM, showing that conventional TIRF images contain unresolved, small clu...

Diagnosis of rheumatoid arthritis using light: correction of motion artefacts and color visualization of multispectral images

Journal of Biophotonics — Wed, 27 Jan 2010 00:00:00 +0100

State-of-the-art image-processing methods offer new possibilities for diagnosing diseases using scattered light. The optical diagnosis of rheumatism is taken as an example to show that the diagnostic sensitivity can be improved using overlapped pseudocolored images of different wavelengths, provided that multispectral images are recorded to compensate for any motion-related artefacts that occur during examination. (© 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Discrimination of normal from pre-malignant cervical tissue by Raman mapping of de-paraffinized histological tissue sections

Journal of Biophotonics — Sat, 16 Jan 2010 00:00:00 +0100

The authors present Raman cluster mapping of de-paraffinized normal cervical tissue and demonstrate the ability of this approach to differentiate between normal squamous epithelium and cervical intraepithelial neoplasia (CIN). Multivariate analysis was performed by hierarchical cluster analysis (HCA) of the Raman spectra associated with the different tissue types and Raman maps were generated using the resultant clusters. Using normal cervical tissue, squamous epithelium and the epithelial-stromal interface, a muscular artery and endocervical glands were successfully mapped. Analysis of a tissue section containing a cervical intraepithelial neoplasia (CIN) grade 2 lesion adjacent to normal squamous epithelium demonstrated that the CIN lesion clustered predominantly with the basal epithelia...

Optical assessment of cutaneous blood volume depends on the vessel size distribution: a computer simulation study

Journal of Biophotonics — Wed, 09 Dec 2009 00:00:00 +0100

This study did repeated random placement of vessels to learn the variation in R due to variable vessel placement. The coefficient of variation was about 10% due to random placement. R varies with size, even with small vessels, and does not simply apply to large vessels. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Towards an optical biopsy for the diagnosis of breast cancer in vivo by endogenous fluorescence spectroscopy

Journal of Biophotonics — Wed, 02 Dec 2009 00:00:00 +0100

The techniques of medical imaging allow the detection of suspect lesions in the breast, but they do not always evidence the malignant nature of these lesions. Breast biopsies and histological analyses are therefore implemented to establish a diagnosis. In order to reduce the number of these invasive procedures, a portable clinical system was designed based upon the excitation of Endogenous Fluorescence in vivo at 405 nm via a fiber-optics probe included in a disposable needle of small diameter ( (Source: Journal of Biophotonics)

Photodynamic therapy induces microRNA-210 and -296 expression in HeLa cells

Journal of Biophotonics — Tue, 01 Dec 2009 00:00:00 +0100

In this study, apoptosis-related miRNA levels were characterized by real-time PCR after PDT in HeLa cells. miR-210 and miR-296 expression levels increased 1 hour after PDT, although the other apoptosis-related miRNA (miR-7, miR-148a, miR-204, and miR-216) expression levels were unchanged. Since miR-210 is the most prominent miRNA consistently stimulated under hypoxic conditions and VEGF is capable of increasing miR-296 expression levels, these data suggest that hypoxia induced by PDT induces miR-210 expression, followed by an increase of VEGF expression and miR-296 expression. These results constitute the first report of an analysis of miRNA expression after PDT using talaporfin sodium as a photosensitizer, demonstrating that miR-210 and miR-296 expression levels are markers of PDT efficac...

Signal generation and Raman-resonant imaging by non-degenerate four-wave mixing under tight focusing conditions

Journal of Biophotonics — Tue, 01 Dec 2009 00:00:00 +0100

The authors demonstrate Raman-resonant imaging based on the simultaneous generation of several nonlinear frequency mixing processes resulting from a 3-color coherent anti-Stokes Raman scattering (CARS) experiment. The interaction of three coincident short-pulsed laser beams simultaneously generates both 2-color (degenerate) CARS and 3-color (non-degenerate) CARS signals, which are collected and characterized spectroscopically - allowing for resonant, doubly-resonant, and non-resonant contrast mechanisms. Images obtained from both 2-color and 3-color CARS signals are compared and found to provide complementary information. The 3-color CARS microscopy scheme provides a versatile multiplexed modality for biological imaging, which may extend the capabilities of label-free non-linear microscopy...

Interpreting diffuse reflectance for in vivo skin reactions in terms of chromophores

Journal of Biophotonics — Fri, 27 Nov 2009 00:00:00 +0100

The measurement and quantification of skin reactions to insults involves certain assumptions about the relation between intensity of color appearance of the skin and the concentration of endogenous chromophores. The underlying assumption is that the Beer-Lambert law is obeyed, i.e., that a linear relation exists between the absorbance and the concentration of each chromophore and that the total absorbance is the linear superposition of the contributions of each chromophore. In this paper the authors compiled the results from a number of interventions on human skin that result in changes in its appearance and small deviations from the homeostatic state, where the results may be accounted for by a single or multiple chromophores. The validity of the assumptions is found to hold for a limited...

In vivo skin optical clearing by glycerol solutions: mechanism

Journal of Biophotonics — Tue, 24 Nov 2009 00:00:00 +0100

In this study, rat dorsal skin was used to discuss the mechanism for in vivo optical clearing. Glycerol solutions with different concentrations were applied by dermal injection; then the skin reflectance spectrum, SHG imaging and microstructural changes were monitored. The results showed that with the skin becoming transparent, the corresponding reflectance decreased, and the thickness of dermis and diameter of collagen fibril decreased, but no collagen fiber was dissolved or fractured. Hence, it can be concluded that the thickness decrease of dermis and corresponding more regular packing of tissue fibers plays an important role in the mechanism for glycerol-induced optical clearing of skin in vivo. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

2-D mapping of skin chromophores in the spectral range 500 - 700 nm

Journal of Biophotonics — Thu, 05 Nov 2009 00:00:00 +0100

The multi-spectral imaging technique has been used for distant mapping of in-vivo skin chromophores by analyzing spectral data at each reflected image pixel and constructing 2-D maps of the relative concentrations of oxy-/deoxy-haemoglobin and melanin. Instead of using a broad visible-NIR spectral range, this study focuses on narrowed spectral band 500-700 nm, speeding-up the signal processing procedure. Regression analysis confirmed that superposition of three Gaussians is optimal analytic approximation for the oxy-haemoglobin absorption tabular spectrum in this spectral band, while superposition of two Gaussians fits well for deoxy-haemoglobin absorption and exponential function - for melanin absorption. The proposed approach was clinically tested for three types of in-vivo skin provocat...

Cultured human keratinocytes for optical transmission measurement

Journal of Biophotonics — Mon, 19 Oct 2009 23:00:00 +0100

The challenges of measuring optical properties of human tissues include the thickness of the sample, homogenization, or crystallization from freezing of the tissue. This investigation demonstrates a method to avoid these problems by growing optically thin samples of human keratinocytes as a substitute for ex vivo epidermis samples. Several methods of growth were investigated. Resulting samples were measured on a spectrophotometer for transmission between 300 nm and 2600 nm. The efficacy of the cell growth was confirmed with histological examination of several cultured keratinocyte samples. Limitations were the requirement to measure samples immediately after removal from the incubation environment, and the absence of the irregular structures of normal skin such as hair and glands. (© 2009...

Concurrent observations of astrocytic Ca2+ activity and multisite extracellular potentials from an intact cerebral cortex

Journal of Biophotonics — Thu, 15 Oct 2009 23:00:00 +0100

In basic neuroscience, the attention has been recently focused on the role played by the protoplasmic astrocytes in modulating the activity of nearby neurons or else on assisting a long-term/sustained communication between these neurons and the surrounding microvasculature. However, to understand the physiological mechanisms underlying such a multiscale interactions in space and time, novel methodologies are required. This paper reports about an experimental setting and a procedure that was developed to obtain concurrently two-photon astrocytic Ca2+ imaging and multisite large-scale extracellular potentials as recorded by a silicon-based probe. Solutions to several technical drawbacks (e.g. removal of photoelectric artifacts, the establishment of safety ranges for microinjection) are provi...

Monitoring of glucose permeability in monkey skin in vivo using Optical Coherence Tomography

Journal of Biophotonics — Mon, 12 Oct 2009 23:00:00 +0100

Topical trans-dermal delivery of drugs has proven to be a promising route for treatment of many dermatological diseases. The aim of this study is to monitor and quantify the permeability rate of glucose solutions in rhesus monkey skin noninvasively in vivo as a primate model for drug diffusion. A time-domain Optical Coherence Tomography (OCT) system was used to image the diffusion of glucose in the skin of anesthetized monkeys for which the permeability rate was calculated. From 5 experiments on 4 different monkeys, the permeability for glucose-20% was found to be (4.41 ± 0.28) 10-6 cm/sec. The results suggest that OCT might be utilized for the noninvasive study of molecular diffusion in the multilayered biological tissues in vivo. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Sou...

The potential of biophotonic techniques in stem cell tracking and monitoring of tissue regeneration applied to cardiac stem cell therapy

Journal of Biophotonics — Mon, 28 Sep 2009 23:00:00 +0100

The use of injected stem cells, leading to regeneration of ischemic heart tissue, for example, following coronary artery occlusion, has emerged as a major new option for managing 'heart attack' patients. While some clinical trials have been encouraging, there have also been failures and there is little understanding of the multiplicity of factors that lead to the outcome. In this overview paper, the opportunities and challenges in applying biophotonic techniques to regenerative medicine, exemplified by the challenge of stem cell therapy of ischemic heart disease, are considered. The focus is on optical imaging to track stem cell distribution and fate, and optical spectroscopies and/or imaging to monitor the structural remodeling of the tissue and the resulting functional changes. The scien...

A fluorescence lifetime imaging scanning confocal endomicroscope

Journal of Biophotonics — Sun, 27 Sep 2009 23:00:00 +0100

We describe a fluorescence lifetime imaging endomicroscope employing a fibre bundle probe and time correlated single photon counting. Preliminary images of stained pollen grains, eGFP-labelled cells exhibiting Förster resonant energy transfer and tissue autofluorescence are presented. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Scoring of collagen organization in healthy and diseased human dermis by multiphoton microscopy

Journal of Biophotonics — Mon, 21 Sep 2009 23:00:00 +0100

We have used nonlinear imaging to evaluate collagen organization in connective tissue ex-vivo samples. Image analysis methods were tested on healthy dermis, normal scars, and keloids. The evaluation of the second harmonic to autofluorescence aging index of dermis (SAAID) has allowed a first characterization of tissues by scoring the collagen/elastin content. Further analyses on collagen morphology in healthy dermis and keloids were performed by image-pattern analysis of SHG images. The gray-level co-occurrence matrix (GLCM) analysis method has allowed classification of different tissues based on the evaluation of geometrical arrangement of collagen fibrillar bundles, whereas a pattern analysis of the FFT images has allowed the discrimination of different tissues based on the anisotropy of ...

Quantified characterization of human cutaneous normal scar using multiphoton microscopy

Journal of Biophotonics — Fri, 18 Sep 2009 23:00:00 +0100

The morphological alterations of human cutaneous normal scar were quantitatively analyzed using multiphoton microscopy (MPM) based on two-photon excited fluorescence and second harmonic generation. High-contrast, high-resolution images of normal scar and uninjured skin were obtained for comparison. In addition, some quantitative parameters have been extracted to quantitatively discriminate between normal scar and uninjured skin. The MPM combined with quantitative method enable a better understanding of microstructual alterations of the epidermis, elastic fiber, and collagen in normal scar. It may lead the way to making know the mechanism of normal scar formation and identifying feasible therapeutic options. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotoni...

Characterization of vascular structures and skin bruises using hyperspectral imaging, image analysis and diffusion theory

Journal of Biophotonics — Tue, 08 Sep 2009 23:00:00 +0100

Hyperspectral imaging, image analysis and diffusion theory were used to visualize skin vasculature and to monitor the development of fresh skin bruises. Bruises were inflicted in a porcine model, and the development of the hemorrhage was monitored using white light hyperspectral imaging (400-1000 nm). Hyperspectral images from human volunteers were also included in the study. Statistical image analysis was used to classify bruised regions and to visualize the skin vasculature. Biopsies were collected from the animals to reveal the true depth of the bruising. A three-layer diffusion model and an analytic hemoglobin transport model were used to model the reflectance spectra from the images. The results show that hyperspectral images contain depth information, and that the approximate depth a...

Tissue viability (TiVi) imaging: temporal effects of local occlusion studies in the volar forearm

Journal of Biophotonics — Fri, 28 Aug 2009 23:00:00 +0100

Tissue Viability (TiVi) imaging is a promising new technology for the assessment of microcirculation in the upper human dermis. Although the technique is easily implemented and develops large amounts of observational data, its role in the clinical workplace awaits the development of standardised protocols required for routine clinical practice. The present study investigates the use of TiVi technology in a human, in vivo, localized, skin blood flow occlusion protocol. In this feasibility study, the response of the cutaneous microcirculation after provocation on the volar surface of the forearm was evaluated using a high temporal-low spatial resolution TiVi camera. 19 healthy subjects - 10 female and 9 male - were studied after a localized pressure was applied for 5 different time periods r...

Imaging of tumor vasculature using Twente photoacoustic systems

Journal of Biophotonics — Thu, 27 Aug 2009 23:00:00 +0100

Photoacoustic imaging is a hybrid imaging modality based on the detection of acoustic waves generated by the absorption of short laser pulses in biological tissue. It combines the advantages of excellent contrast achieved in optical techniques with the high resolution of ultrasound imaging. In this article we present a review of the work done at the University of Twente to image tumor angiogenesis in vivo using this technique. We start with a description and the technical details of the different photoacoustic systems developed in our laboratory, with their validation on phantoms. We then discuss small-animal studies with results of serial imaging of angiogenesis over a 10-day period at the site of tumor induction in a rat. Further, we present clinical results using a photoacoustic mammosc...

Optical manipulation for single-cell studies

Journal of Biophotonics — Thu, 27 Aug 2009 23:00:00 +0100

In the last decade optical manipulation has evolved from a field of interest for physicists to a versatile tool widely used within life sciences. This has been made possible in particular due to the development of a large variety of imaging techniques that allow detailed information to be gained from investigations of single cells. The use of multiple optical traps has high potential within single-cell analysis since parallel measurements provide good statistics. Multifunctional optical tweezers are, for instance, used to study cell heterogeneity in an ensemble, and force measurements are used to investigate the mechanical properties of individual cells. Investigations of molecular motors and forces on the single-molecule level have led to discoveries that would have been difficult to make...

Automated laser guidance of neuronal growth cones using a spatial light modulator

Journal of Biophotonics — Mon, 24 Aug 2009 23:00:00 +0100

The growth cone of a developing neuron can be guided using a focused infra-red (IR) laser beam [1]. In previous setups this process has required a significant amount of user intervention to adjust continuously the laser beam to guide the growing neuron. Previously, a system using an acousto-optical deflector (AOD) has been developed to steer the beam [2]. However, to enhance the controllability of this system, here we demonstrate the use of a computer controlled spatial light modulator (SLM) to steer and manipulate the shape of a laser beam for use in guided neuronal growth. This new experimental setup paves the way to enable a comprehensive investigation into beam shaping effects on neuronal growth and we show neuronal growth initiated by a Bessel light mode. This is a robust platform to ...

A two-photon europium complex as specific endoplasmic reticulum probe

Journal of Biophotonics — Thu, 13 Aug 2009 23:00:00 +0100

Highly emissive europium complexes with specific endoplasmic reticulum localization potential includes several advantages such as fast uptake, long resident lifetime, low dosage requirement, low cytotoxicity and highly emissive two-photon induced f-f emission imaging. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Resonance Raman spectroscopy as an effective tool for the determination of antioxidative stability of cosmetic formulations

Journal of Biophotonics — Thu, 13 Aug 2009 23:00:00 +0100

Carotenoids beta-carotene, lutein, lycopene and others are well-known powerful antioxidants acting as an effective neutralizer of free radicals produced in the human organism as a result of the influence of stress factors, such as UV irradiation. The protective effect of antioxidants is used in cosmetic products to increase the skin protection against the destructive action of free radicals and for the stabilization of formulations against oxidation. In the skin, the different antioxidant substances form protection chains to avoid their destruction by the interaction with the free radicals. Similar effects have to be expected also in topically applied formulations. In the present study the influence of different mixtures of antioxidants (beta-carotene, vitamins C and E) on the stability of...

Imaging engineered tissues using structural and functional optical coherence tomography

Journal of Biophotonics — Mon, 10 Aug 2009 23:00:00 +0100

As the field of tissue engineering evolves, there will be an increasingly important need to visualize and track the complex dynamic changes that occur within three-dimensional constructs. Optical coherence tomography (OCT), as an emerging imaging technology applied to biological materials, offers a number of significant advantages to visualize these changes. Structural OCT has been used to investigate the longitudinal development of engineered tissues and cell dynamics such as migration, proliferation, detachment, and cell-material interactions. Optical techniques that image functional parameters or integrate multiple imaging modalities to provide complementary contrast mechanisms have been developed, such as the integration of optical coherence microscopy with multiphoton microscopy to im...

Quantification of Mucosa oxygenation using three discrete spectral bands of visible light

Journal of Biophotonics — Mon, 10 Aug 2009 23:00:00 +0100

Quantification of the mucosa oxygenation levels during Endoscopic imaging provides useful physiological/diagnostic information. In this work a method for non-contact quantification of the oxygen saturation index during Endoscopic imaging using three discrete spectral-band in the blue, the green, and the red parts of the spectrum (RGB bands) has been investigated. The oxygen saturation index (TOI_rgb) was calculated from the three discrete RGB spectral bands using diffusion approximation modeling and least-square analysis. A parametric study performed to identify the optimum band width for each of the three spectral bands. The quantification algorithm was applied to in vivo images of the endobronchial mucosa to calculate (TOI_rgb) from selected areas within the image view. The results were ...

Clinical applications of slide-based cytometry - an update

Journal of Biophotonics — Sun, 09 Aug 2009 23:00:00 +0100

Slide-based cytometric approaches open the possibility to obtain quantitative and objective data from specimens that so far have not been accessible to this kind of analysis. In this review, we will highlight the specific advantages of slide-based cytometry (SBC) and show the applications that have been established for clinical samples. Focuses are cytomic analyses of oncological and hematological samples where the slide-based concept turned out to open new dimensions in understanding underlying cellular networks. We review the recent literature and point out future applications. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Flow cytometry with gold nanoparticles and their clusters as scattering contrast agents: FDTD simulation of light-cell interaction

Journal of Biophotonics — Sun, 09 Aug 2009 23:00:00 +0100

The formulation of the finite-difference time-domain (FDTD) approach is presented in the framework of its potential applications to in-vivo flow cytometry based on light scattering. The consideration is focused on comparison of light scattering by a single biological cell alone in controlled refractive-index matching conditions and by cells labeled by gold nanoparticles. The optical schematics including phase contrast (OPCM) microscopy as a prospective modality for in-vivo flow cytometry is also analyzed. The validation of the FDTD approach for the simulation of flow cytometry may open up a new avenue in the development of advanced cytometric techniques based on scattering effects from nanoscale targets. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Intracellular imaging of host-pathogen interactions using combined CARS and two-photon fluorescence microscopies

Journal of Biophotonics — Thu, 06 Aug 2009 23:00:00 +0100

Intracellular imaging is a key tool in the investigation of host-pathogen interactions. Advances in this area are particularly sought to understand the effect of viral infection processes on the host cell and its metabolic functions including those cases where host cell lipid metabolism is modulated as a result of infection. We demonstrate the use of combined coherent anti-Stokes Raman scattering (CARS) and two-photon fluorescence microscopies to image fibroblast cells infected by cytomegalovirus. CARS is used to image the host cell membrane, lipid droplets and morphology of the nucleus. Cell nuclei are found to expand during infection, approximately doubling in area. Some cells also show accumulations of lipid droplets at the nuclear periphery. Using a genetically modified virus strain ex...

Label-free biochemical characterization of stem cells using vibrational spectroscopy

Journal of Biophotonics — Sun, 02 Aug 2009 23:00:00 +0100

This article reviews recent developments in applying these techniques for the characterization of stem cells. A discussion of the potential for these methods to address some of the major challenges in stem cell research is presented, as well as the technological and scientific advancements that are needed to progress the knowledge in the field. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

The metabolic component of cellular refractivity and its importance for optical cytometry

Journal of Biophotonics — Thu, 30 Jul 2009 23:00:00 +0100

Initially, it has been shown that the phase thickness and refractivity (the latter interpreted as the difference of the refractivity indices of an object and surrounding milieu) depend on the functional state of mitochondria. The refractivity of various objects decreased in response to energy depletion. This dependence was then demonstrated for other biological objects such as cyanobacteria, chloroplasts and human cells. This general response brought about the hypothesis of a certain "universal" factor that links the variable (or metabolic) component of refractivity with the object's functional state. However, the origin of this phenomenon remains unknown. Our hypothesis is founded on the dependence of polarization of bound water molecules and the activity of metabolic processes. Here, we ...

Surgical applications of femtosecond lasers

Journal of Biophotonics — Tue, 14 Jul 2009 23:00:00 +0100

Femtosecond laser ablation permits non-invasive surgeries in the bulk of a sample with submicrometer resolution. We briefly review the history of optical surgery techniques and the experimental background of femtosecond laser ablation. Next, we present several clinical applications, including dental surgery and eye surgery. We then summarize research applications, encompassing cell and tissue studies, research on C. elegans, and studies in zebrafish. We conclude by discussing future trends of femtosecond laser systems and some possible application directions. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Targeted optical injection of gold nanoparticles into single mammalian cells

Journal of Biophotonics — Tue, 14 Jul 2009 23:00:00 +0100

We present an all optical technique for the targeted delivery of single 100 nm diameter gold nanoparticles into a specified region of the interior of an individual mammalian cell through a combination of optical tweezing and optical injection. The internalisation of the nanoparticle is verified by confocal laser scanning microscopy and confocal laser scanning reflectance microscopy. This represents the first time that nano sized particles have been tweezed and optically injected into mammalian cells using only light, and provides a novel methodology for internalising nanosphere based biosensors within specific intracellular regions of a mammalian cell. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Diffraction imaging of spheres and melanoma cells with a microscope objective

Journal of Biophotonics — Fri, 10 Jul 2009 23:00:00 +0100

Diffraction imaging of polystyrene spheres and B16F10 mouse melanoma cells embedded in gel has been investigated with a microscope objective. The diffraction images acquired with the objective from a sphere have been shown to be comparable to the Mie theory based projection images of the scattered light if the objective is translated to defocused positions towards the sphere. Using a confocal imaging based method to reconstruct and analyze the 3D structure, we demonstrated that genetic modifications in these cells can induce morphological changes and the modified cells can be used as an experimental model for study of the correlation between 3D morphology features and diffraction image data. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Investigation of multifocal choroiditis with panuveitis by three-dimensional high-penetration optical coherence tomography

Journal of Biophotonics — Sat, 04 Jul 2009 23:00:00 +0100

A single case of multifocal choroiditis with panuveitis (MFCPU) was investigated by a three-dimensional (3-D) high-penetration optical coherence tomography. The HP-OCT is based on a swept-source OCT technology, uses a probe beam with a center wavelength of 1060 nm, and possesses a depth resolution of 10.4 [mu] min tissue. Two eyes of an MFCPU patient were involved in this study. The eyes were also examined by color fundus photograph, fluorescein angiography (FA), and indocyanine green angiography (ICGA). Findings in these four modalities are comparatively discussed. The OCT scans revealed the following characteristic properties of the lesion sites. Thinning of the retina, destructuring of the retinal layers, and disappearance of the junction of the inner and outer segments of the photorece...

Optical coherence tomography and Raman spectroscopy of the ex-vivo retina

Journal of Biophotonics — Mon, 29 Jun 2009 23:00:00 +0100

We describe an OCT microscope for ex-vivo imaging combined with Raman spectroscopy capable of collecting morphological and molecular information about a sample simultaneously. We present our first results and discuss the challenges to further development of this dual-mode instrument and limitations for future in-vivo retinal imaging. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Recent developments in Fourier Domain Mode Locked lasers for optical coherence tomography: Imaging at 1310 nm vs. 1550 nm wavelength

Journal of Biophotonics — Mon, 29 Jun 2009 23:00:00 +0100

We report on recent progress in Fourier domain mode-locking (FDML) technology. The paper focuses on developments beyond pushing the speed of these laser sources. After an overview of improvements to FDML over the last three years, a brief analysis of OCT imaging using FDML lasers with different wavelengths is presented. For the first time, high speed, high quality FDML imaging at 1550 nm is presented and compared to a system at 1310 nm. The imaging results of human skin for both wavelengths are compared and analyzed. Sample arm optics, power on the sample, heterodyne gain, detection bandwidth, colour cut levels and sample location have been identical to identify the influence of difference in scattering and water absorption. The imaging performance at 1310 nm in human skin is only slightly...

OCT imaging of skin cancer and other dermatological diseases

Journal of Biophotonics — Thu, 25 Jun 2009 23:00:00 +0100

Optical coherence tomography (OCT) provides clinicians and researchers with micrometer-resolution, in vivo, cross-sectional images of human skin up to several millimeter depth. This review of OCT imaging applied within dermatology covers the application of OCT to normal skin, and reports on a large number of applications in the fields of non-melanoma skin cancer, malignant melanomas, psoriasis and dermatitis, infestations, bullous skin diseases, tattoos, nails, haemangiomas, and other skin diseases. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

AM-FM techniques in the analysis of optical coherence tomography signals

Journal of Biophotonics — Wed, 24 Jun 2009 23:00:00 +0100

The subtle tissue changes associated with the early stages of malignancies, such as cancer, are not clearly discernible even at the current, improved, resolution of optical coherence tomography (OCT) systems. However, these changes directly affect the spectral content of the OCT image that contains information regarding these unresolvable features. Spectral analysis of OCT signals has recently been shown to provide additional information, resulting in improved contrast, directly related to scatterer size changes. Amplitude modulation-frequency modulation (AM-FM) analysis, a fast and accurate technique for the estimation of the instantaneous frequency, phase, and amplitude of a signal, can also be applied to OCT images to extract scatterer-size information. The proposed technique could make...

A concentric three element radial scanning optical coherence tomography endoscope

Journal of Biophotonics — Mon, 15 Jun 2009 23:00:00 +0100

We have developed a 2.1 mm outer diameter optical coherence tomography endoscope that provides radial scans of luminal structures. The endoscope consists of three elements: (1) a stationary central core containing the fibers and focusing elements, (2) a rotating intermediate tube with a distal rod prism, and (3) a stationary sterilized glass cover. This design enables radial and spiral scanning and allows adjustment of the axial focal distance. Additionally, this design is capable of focusing light from multiple fibers into tissue. The performance of the endoscope was demonstrated in a study of tissue engineered blood vessels imaged at various time points during development. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Doppler calibration method for Spectral Domain OCT spectrometers

Journal of Biophotonics — Mon, 15 Jun 2009 23:00:00 +0100

We present a calibration method for SD-OCT domain spectrometers based on the M-scan of a moving mirror. This method allows determination of the wavenumber sampling increment which determines the depth axis assigned to the structural image. It also allows wavelength calibration of individual pixels which ensures correct re-sampling prior to Fast Fourier Transform. Determination of the spectrometer resolution, which determines sensitivity and resolution decay with depth, is also possible. The wavelength calibration results suggest that hardware calibration of the spectrometers is a necessary complement to the computational methods presented here. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) (Source: Journal of Biophotonics)

Histogram-based filtering for quantitative 3D retinal angiography

Journal of Biophotonics — Thu, 11 Jun 2009 23:00:00 +0100

Doppler Fourier domain optical coherence tomography (D-FDOCT) can be used to visualise and measure ocular blood flow in the retina of the human eye. This has the potential to diagnose illnesses such as macula degeneration or diabetic retinopathy early on because these medical conditions cause a pathological change in the retinal vasculature and perfusion. Here we are presenting a method to separate quantitative blood flow information from static tissue. Our approach is demonstrated on retinal D-FDOCT volume scans taken at the optic nerve head (see figure) and near the fovea with a high speed CMOS-based FDOCT system. The advantage of this method is the small post-processing effort together with the immediate availability of segmented quantitative Doppler flow maps. The performance of the fi...

Measurements of depolarization distribution in the healthy human macula by polarization sensitive OCT

Journal of Biophotonics — Thu, 11 Jun 2009 23:00:00 +0100

Polarization sensitive optical coherence tomography (PS-OCT) allows for depth resolved imaging of polarization properties of retinal structures. Different layers in the retina can be distinguished not only based on their reflectivity but also due to their birefringent, depolarizing or polarization preserving character. In contrast to other structures in the posterior segment, the retinal pigment epithelium (RPE) is depolarizing, i.e., it scrambles the polarization state of light. A spectral domain (SD) PS-OCT system was used to record 3D data sets of the retina. The depolarizing effect of the RPE was analyzed by quantitative evaluation of the spatial depolarization distribution in the macula regions of healthy eyes. Depolarization in the RPE is most pronounced close to the center of the fo...

Plasmonic nanoprobes for SERS biosensing and bioimaging

Journal of Biophotonics — Tue, 09 Jun 2009 23:00:00 +0100

This article provides an overview of the development and application of plasmonic nanoprobes developed in our laboratory for biosensing and bioimaging. We describe the use of plasmonics surface-enhanced Raman scattering (SERS) gene probes for the detection of diseases using DNA hybridization to target biospecies (HIV gene, breast cancer genes etc.). For molecular imaging, we describe a hyperspectral surface-enhanced Raman imaging (HSERI) system that combines imaging capabilities with SERS detection to identify cellular components using Raman dye-labeled silver nanoparticles in cellular systems The detection of specific target DNA sequences associated with breast cancer using "molecular sentinel" nanoprobes and the use of a plasmonic nanosensor to monitor pH in single cells are presented an...

Basics of standardization and calibration in cytometry - a review