Showing papers in "Journal of Biophotonics in 2009"

Disease recognition by infrared and Raman spectroscopy.

Abstract: Infrared (IR) and Raman spectroscopy are emerging biophotonic tools to recognize various diseases. The current review gives an overview of the experimental techniques, data-classification algorithms and applications to assess soft tissues, hard tissues and body fluids. The methodology section presents the principles to combine vibrational spectroscopy with microscopy, lateral information and fiber-optic probes. A crucial step is the classification of spectral data by a variety of algorithms. We discuss unsupervised algorithms such as cluster analysis or principal component analysis and supervised algorithms such as linear discriminant analysis, soft independent modeling of class analogies, artificial neural networks support vector machines, Bayesian classification, partial least-squares regression and ensemble methods. The selected topics include tumors of epithelial tissue, brain tumors, prion diseases, bone diseases, atherosclerosis, kidney stones and gallstones, skin tumors, diabetes and osteoarthritis.

Surgical applications of femtosecond lasers

Abstract: 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.

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

Abstract: 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 collagen fibers distribution. This multiple scoring method represents a promising tool to be extended to other collagen disorders, as well as to be used in in-vivo skin-imaging applications.

Plasmonic nanoprobes for SERS biosensing and bioimaging.

Abstract: 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 and discussed.

OCT imaging of skin cancer and other dermatological diseases.

Abstract: 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.

Mueller matrix decomposition for polarized light assessment of biological tissues

Abstract: The Mueller matrix represents the transfer function of an optical system in its interactions with polarized light and its elements relate to specific biologically or clinically relevant properties. However, when many optical polarization effects occur simultaneously, the resulting matrix elements represent several "lumped" effects, thus hindering their unique interpretation. Currently, no methods exist to extract these individual properties in turbid media. Here, we present a novel application of a Mueller matrix decomposition methodology that achieves this objective. The methodology is validated theoretically via a novel polarized-light propagation model, and experimentally in tissue simulating phantoms. The potential of the approach is explored for two specific biomedical applications: monitoring of changes in myocardial tissues following regenerative stem cell therapy, through birefringence-induced retardation of the light's linear and circular polarizations, and non-invasive blood glucose measurements through chirality-induced rotation of the light's linear polarization. Results demonstrate potential for both applications.

Circulation and distribution of gold nanoparticles and induced alterations of tissue morphology at intravenous particle delivery.

Abstract: Kinetics, biodistribution, and histological studies were performed to evaluate the particle-size effects on the distribution of 15 nm and 50 nm PEG-coated colloidal gold (CG) particles and 160 nm silica/gold nanoshells (NSs) in rats and rabbits. The above nanoparticles (NPs) were used as a model because of their importance for current biomedical applications such as photothermal therapy, optical coherence tomography, and resonance-scattering imaging. The dynamics of NPs circulation in vivo was evaluated after intravenous administration of 15 nm CG NPs to rabbit, and the maximal concentrations of gold were observed 15-30 min after injection. Rats were injected in the tail vein with PEG-coated NPs (about 0.3 mg Au/kg rats). 24 h after injection, the accumulation of gold in different organs and blood was determined by atomic absorption spectroscopy. In accordance with the published reports, we observed 15 nm particles in all organs with rather smooth distribution over liver, spleen and blood. By contrast, the larger NSs were accumulated mainly in the liver and spleen. For rabbits, the biodistribution was similar (72 h after intravenous injection). We report also preliminary data on the light microscopy and TEM histological examination that allows evaluation of the changes in biotissues after gold NPs treatment.

In vivo skin optical clearing by glycerol solutions: mechanism.

Abstract: The penetration of light in biological tissues can be enhanced by using optical-clearing techniques. However, researches on the mechanism are limited to in vitro experiments. 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.

Nanotechnology-based molecular photoacoustic and photothermal flow cytometry platform for in vivo detection and killing of circulating cancer stem cells

Abstract: In-vivo multicolor photoacoustic (PA) flow cytometry for ultrasensitive molecular detection of the CD44+ circulating tumor cells (CTCs) is demonstrated on a mouse model of human breast cancer. Targeting of CTCs with stem-like phenotype, which are naturally shed from parent tumors, was performed with functionalized gold and magnetic nanoparticles. Results in vivo were verified in vitro with a multifunctional microscope, which integrates PA, photothermal (PT), fluorescent and transmission modules. Magnet-induced clustering of magnetic nanoparticles in individual cells significantly amplified PT and PA signals. The novel noninvasive platform, which integrates multispectral PA detection and PT therapy with a potential for multiplex targeting of many cancer biomarkers using multicolor nanoparticles, may prospectively solve grand challenges in cancer research for diagnosis and purging of undetectable yet tumor-initiating cells in circulation before they form metastasis.

Raman spectroscopy of DNA packaging in individual human sperm cells distinguishes normal from abnormal cells.

Abstract: Healthy human males produce sperm cells of which about 25-40% have abnormal head shapes. Increases in the percentage of sperm exhibiting aberrant sperm head morphologies have been correlated with male infertility, and biochemical studies of pooled sperm have suggested that sperm with abnormal shape may contain DNA that has not been properly repackaged by protamine during spermatid development. We have used micro-Raman spectroscopy to obtain Raman spectra from individual human sperm cells and examined how differences in the Raman spectra of sperm chromatin correlate with cell shape. We show that Raman spectra of individual sperm cells contain vibrational marker modes that can be used to assess the efficiency of DNA-packaging for each cell. Raman spectra obtained from sperm cells with normal shape provide evidence that DNA in these sperm is very efficiently packaged. We find, however, that the relative protein content per cell and DNA packaging efficiencies are distributed over a relatively wide range for sperm cells with both normal and abnormal shape. These findings indicate that single cell Raman spectroscopy should be a valuable tool in assessing the quality of sperm cells for in-vitro fertilization.

Towards oncological application of Raman spectroscopy

Abstract: As the possibilities in the treatment of cancer continue to evolve, its early detection and correct diagnosis are becoming increasingly important. From the early detection of cancer to the guidance of oncosurgical procedures new sensitive in vivo diagnostic tools are much needed. Many studies report the Raman spectroscopic detection of malignant and premalignant tissues in different sites of the body with high sensitivities. The great appeal of this technique lies in its potential for in vivo clinical implementation. We present an overview of the in vitro and in vivo work on the oncological application of Raman spectroscopy and discuss its potential as a new tool in the clinico-oncological practice. Opportunities for integration of Raman spectroscopy in oncological cure and care as a real-time guidance tool during diagnostic (i.e. biopsy) and therapeutic (surgical resection) modalities as well as technical shortcomings are discussed from a clinician's point of view.

In vivo fiber-based multicolor photoacoustic detection and photothermal purging of metastasis in sentinel lymph nodes targeted by nanoparticles.

Abstract: This report introduces a novel diagnostic and therapeutic platform for in vivo non-invasive detection and treatment of metastases in sentinel lymph nodes (SLNs) at single cell level using an integrated system of multicolor photoacoustic (PA) lymph flow cytometry, PA lymphography, absorption image cytometry, and photothermal (PT) therapy. A melanoma-bearing mouse model was used to demonstrate the capability of this platform for real-time lymphatic mapping, counting of disseminated tumor cells (DTCs) in prenodal lymphatics, and detecting metastasis in SLNs and its purging. The detection and ablation of non-pigmented breast cancer cells in SLNs was achieved by labeling them with nanoparticles. The association between DTC count and SLN metastasis progression supports lymphatic DTCs as a novel prognostic marker of metastasis. The fiber-based portable PA device may replace the conventional SLN(s) excision and histology-based staging. The earliest detection of DTCs in the lymphatic vessels before the establishment of nodal metastasis may prevent metastasis by well-timed ablation of DTCs.

A comparative Raman and CARS imaging study of colon tissue.

Abstract: An experimental evaluation of the information content of two complimentary techniques, linear Raman and coherent anti-Stokes Raman scattering (CARS) microscopy, is presented. CARS is a nonlinear variant of Raman spectroscopy that enables rapid acquisition of images within seconds in combination with laser scanning microscopes. CARS images were recorded from thin colon tissue sections at 2850, 1660, 1450 and 1000 cm–1 and compared with Raman images. Raman images were obtained from univariate and multivariate (k-means clustering) methods, whereas all CARS images represent univariate results. Variances within tissue sections could be visualized in chemical maps of CARS and Raman images. However, identification of tissue types and characterization of variances between different tissue sections were only possible by analysis of cluster mean spectra, obtained from k-means cluster analysis. This first comparison establishes the foundation for further development of the CARS technology to assess tissue. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

Selective targeting of green fluorescent nanodiamond conjugates to mitochondria in HeLa cells.

Abstract: Fluorescent cellular biomarkers play a prominent role in biosciences. Most of the available biomarkers have some drawbacks due to either physical and optical or cytotoxic properties. In view of this, we investigated the potential of green fluorescent nanodiamonds as biomarkers in living cells. Nanodiamonds were functionalized by attaching antibodies that target intracellular structures such as actin filaments and mitochondria. Then, the nanodiamond conjugates were transfected into HeLa cells. Transfections were mediated by 4(th)-generation dendrimers, cationic liposomes and protamine sulfate. Using fluorescence microscopy, we confirmed successful transfections of the nanodiamonds into HeLa cells. Nanodiamond fluorescence could be easily differentiated from cellular autofluorescence. Furthermore, nanodiamonds could be targeted selectively to intracellular structures. Therefore, nanodiamonds are a promising tool for intracellular assays.

Label-free biochemical characterization of stem cells using vibrational spectroscopy.

Abstract: Raman and infrared (IR) spectroscopy are two complementary vibrational spectroscopic techniques that have experienced a tremendous growth in their use in biological and biomedical research. This is, in large part, due to their unique capability of providing label-free intrinsic chemical information of living biological samples at tissue, cellular, or sub-cellular resolutions. 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)

Investigating FTIR based histopathology for the diagnosis of prostate cancer

Abstract: Prostate cancer is the most common gender specific cancer The current gold standard for diagnosis, histopathology, is subjective and limited by variation between different pathologists The diagnostic problems associated with the correct grading and staging of prostate cancer (CaP) has led to an interest in the development of spectroscopic based diagnostic techniques FTIR microspectroscopy used in combination with a Principal Component Discriminant Function Analysis (PC-DFA) was applied to investigate FTIR based histopathology for the diagnosis of CaP In this paper we report the results of a large patient study in which FTIR has been proven to grade CaP tissue specimens to a high degree of sensitivity and specificity

Towards a quantitative SERS approach--online monitoring of analytes in a microfluidic system with isotope-edited internal standards.

Abstract: In this contribution a new approach for quantitative measurements using surface-enhanced Raman spectroscopy (SERS) is presented. Combining the application of isotope-edited internal standard with the advantages of the liquid-liquid segmented-flow-based approach for flow-through SERS detection seems to be a promising means for quantitative SERS analysis. For the investigations discussed here a newly designed flow cell, tested for ideal mixing efficiency on the basis of grayscale-value measurements, is implemented. Measurements with the heteroaromatics nicotine and pyridine using their respective deuterated isotopomers as internal standards show that the integration of an isotopically labeled internal standard in the used liquid-liquid two-phase segmented flow leads to reproducible and comparable SERS spectra independent from the used colloid. With the implementation of an internal standard into the microfluidic device the influence of the properties of the colloid on the SERS activity can be compensated. Thus, the problem of a poor batch-to-batch reproducibility of the needed nanoparticle solutions is solved. To the best of our knowledge these are the first measurements combining the above mentioned concepts in order to correct for differences in the enhancement behaviour of the respective colloid.

Clinical optical coherence tomography combined with multiphoton tomography of patients with skin diseases.

Abstract: We report on the first clinical study based on optical coherence tomography (OCT) in combination with multiphoton tomography (MPT) and dermoscopy. 47 patients with a variety of skin diseases and disorders such as skin cancer, psoriasis, hemangioma, connective tissue diseases, pigmented lesions, and autoimmune bullous skin diseases have been investigated with (i) state-of-the-art OCT systems for dermatology including multibeam swept source OCT, (ii) the femtosecond laser multiphoton tomograph, and (iii) dermoscopes. Dermoscopy provides two-dimensional color images of the skin surface. OCT images reflect modifications of the intratissue refractive index whereas MPT is based on nonlinear excitation of endogenous fluorophores and second harmonic generation. A stack of cross-sectional OCT “wide field” images with a typical field of view of 5 × 2 mm2 gave fast information on the depth and the volume of the lesion. Multiphoton tomography provided 0.36 × 0.36 mm2 horizontal/diagonal optical sections within seconds of a particular region of interest with superior submicron resolution down to a tissue depth of 200 μm. The combination of OCT and MPT provides a unique powerful optical imaging modality for early detection of skin cancer and other skin diseases as well as for the evaluation of the efficiency of treatments. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

Direct analysis of clinical relevant single bacterial cells from cerebrospinal fluid during bacterial meningitis by means of micro-Raman spectroscopy.

Abstract: Bacterial meningitis is a relevant public health concern. Despite the availability of modern treatment strategies it is still a life-threatening disease that causes significant morbidity and mortality. Therefore, an initial treatment approach plays an important role. For in-time identification of specific bacterial pathogens of the cerebrospinal fluid (CSF) and emerged antimicrobial and adjunctive treatment, microbiological examination is of major importance. This contribution spotlights the potential of micro-Raman spectroscopy as a biomedical assay for direct analysis of bacteria in cerebrospinal fluid of patients with bacterial meningitis. The influence of miscellaneous artificial environments on several bacterial species present during bacterial meningitis was studied by means of Raman spectroscopy. The application of chemometric data interpretation via hierarchical cluster analysis (HCA) allows for the differentiation of in vitro cultured bacterial cells and can also be achieved on a single cell level. Moreover as proof of principle the investigation of a CSF sample obtained from a patient with meningococcal meningitis showed that the cerebrospinal fluid matrix does not mask the Raman spectrum of a bacterial cell notably since via chemometric analysis with HCA an identification of N. meningitidis cells from patients with bacterial meningitis could be achieved.

Application of Raman spectroscopy for Cervical Dysplasia Diagnosis

Abstract: Cervical cancer is the second most common malignancy among women worldwide, with over 490 000 cases diagnosed and 274 000 deaths each year. Although current screening methods have dramatically reduced cervical cancer incidence and mortality in developed countries, a “See and Treat” method would be preferred, especially in developing countries. Results from our previous work have suggested that Raman spectroscopy can be used to detect cervical precancers; however, with a classification accuracy of 88%, it was not clinically applicable. In this paper, we describe how incorporating a woman's hormonal status, particularly the point in menstrual cycle and menopausal state, into our previously developed classification algorithm improves the accuracy of our method to 94%. The results of this paper bring Raman spectroscopy one step closer to being utilized in a clinical setting to diagnose cervical dysplasia. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

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

Abstract: 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 and extent of bruises can be retrieved using a combination of statistical image analysis and diffusion theory. This technique also shows potential to visualize vascular structures in human skin.

Imaging of tumor vasculature using Twente photoacoustic systems

Abstract: 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 mammoscope of breast cancer imaging based on angiogenesis-driven optical absorption contrast

Chip scale integrated microresonator sensing systems.

Abstract: Medicine, environmental monitoring, and security are application areas for miniaturized, portable sensing systems. The emerging integration of sensors with other components (electronic, photonic, fluidic) is moving sensing toward higher levels of portability through the realization of self-contained chip scale sensing systems. Planar optical sensors, and in particular, microresonator sensors, are attractive components for chip scale integrated sensing systems because they are small, have high sensitivity, can be surface customized, and can be integrated singly or in arrays in a planar format with other components using conventional semiconductor fabrication technologies. This paper will focus on the progress and prospects for the integration of microresonator sensors at the chip scale with photonic input/output components and with sample preparation microfluidics, toward self-contained, portable sensing systems. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

Imaging engineered tissues using structural and functional optical coherence tomography.

Abstract: 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 image structural and functional information from cells in engineered tissue, optical coherence elastography to generate images or maps of strain to reflect the spatially-dependent biomechanical properties, and spectroscopic OCT to differentiate different cell types From these results, OCT demonstrates great promise for imaging and visualizing engineered tissues, and the complex cellular dynamics that directly affect their practical and clinical use

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

Abstract: 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.

Spectroscopy on the wing: Naturally inspired SERS substrates for biochemical analysis

Abstract: We show that naturally occurring chitinous nanostructures found on the wings of the Graphium butterfly can be used as substrates for surface-enhanced Raman scattering when coated with a thin film of gold or silver. The substrates were found to exhibit excellent biocompatibility and sensitivity, making them ideal for protein assaying. An assay using avidin/biotin binding showed that the substrates could be used to quantify protein binding directly from changes in the surface-enhanced Raman scattering (SERS) spectra and were sensitive over a concentration range comparable with a typical enzyme-linked immunosorbent assays (ELISA) assay. A biomimetic version of the wing nanostructures produced using a highly reproducible, large-scale fabrication process, yielded comparable enhancement factors and biocompatibility. The excellent biocompatibility of the wings and biomimetic substrates is unparalleled by other lithographically produced substrates, and this could pave the way for widespread application of ultrasensitive SERS-based bioassays.

In vivo imaging of hematopoietic stem cells and their microenvironment.

Abstract: In this review we provide a description of the basic concepts and paradigms currently constituting the foundations of adult stem cell biology, and discuss the role that live imaging techniques have in the development of the field. We focus on live imaging of hematopoietic stem cells (HSCs) as the basic biology and clinical applications of HSCs have historically been at the forefront of the stem cell field, and HSC are the first mammalian tissue stem cells to be visualized in vivo using advanced light microscopy techniques. We outline the current technical challenges that remain to be overcome before stem cells and their niche can be more fully characterized using the live imaging technology.

The potential for histological screening using a combination of rapid Raman mapping and principal component analysis.

Abstract: Rapid Raman mapping was carried out on 20 μm sections of oesophageal biopsy samples. Contiguous 7 μm sections were stained with haematoxylin and eosin (H&E) with histopathology provided by an expert pathologist. The step size and acquisition times were varied and the resulting spectra, principal component (PC) score maps and loads were compared. Overall mapping times were also compared to traditional Raman point mapping. The principal component loads for each of the maps were seen to be similar despite varying the acquisition time and number of spectra. Gross biochemical information was extracted showing good correlation with the H&E sections even for short overall mapping times (30–90 minutes for a 2 mm biopsy, 0.5 s acquisition time per 25.3 μm Raman pixel). This demonstrates that low signal to noise spectral maps are sufficient for the identification of histologically relevant biochemistry using principal component analysis as long as the spectral dataset is large enough. (© 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

Classification of fixed urological cells using Raman tweezers.

Abstract: In this paper we report on preliminary investigations into using Raman tweezers to classify urological cell lines. This builds on earlier work within the group, whereby Raman tweezer methodologies were developed, and the application of this technique to differentiate between live prostate cancer (CaP) and bladder cells lines (PC-3 and MGH-U1 respectively) was demonstrated.In this present study we analysed chemically fixed cells using two different fixative methods; SurePath (a commercial available liquid based cytology media) and 4% v/v formalin/PBS fixatives. The study has been expanded from our previous live cell study to include the androgen sensitive CaP cell line LNCaP, primary benign prostate hyperplasia (BPH) cells as well as primary urethral cells. Raman light from the cells was collected using a 514.5 nm Ar-ion laser excitation source in back-scattering configuration mode.Principal component-linear discriminate analysis (PC-LDA) models of resulting cell spectra were generated and these were validated using a blind comparison. Sensitivities and specificities of > 72% and 90% respectively, for SurePath fixed cells, and > 93% and 98% respectively for 4% v/v formalin/PBS fixed cells was achieved. The higher prediction results for the formalin fixed cells can be attributed to a better signal-to-noise ratio for spectra obtained from these cells.Following on from this work, urological cell lines were exposed to urine for up to 12 hours to determine the effect of urine on the ability to classify these cells. Results indicate that urine has no detrimental effect on prediction results.

Dual optical coherence tomography/fluorescence microscopy for monitoring of Drosophila melanogaster larval heart.

Abstract: This article demonstrates a combined instrument of two imaging modalities to acquire information on cardiac function in larval Drosophila melanogaster: optical coherence tomography (OCT) and laser scanning fluorescence microscopy (LSFM). For this purpose, a dedicated imaging instrument able to sequentially provide cross-sectional OCT and C-scan LSFM images has been developed. With this dual-imaging system, the heart can be easily located and visualized within the specimen and the change of the heart shape in a cardiac cycle can be monitored. © 2009 by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.