Image analysis in nonlinear microscopy

Hagmar, Jonas; Brackmann, Christian; Gustavsson, Tomas; Enejder, Annika

doi:10.1364/josaa.25.002195

Cited by 15 publications

(9 citation statements)

References 37 publications

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“…5. In order to obtain quantitative data from the large set of CARS microscopy images, a segmentation algorithm of Local thresholding, which has been proven particularly useful for nonlinear microscopy, [46] was employed for converting the z-stacks to binary images (areas identified as organelles −1, background −0), segmenting the lipid droplets from the background signal generated by the yeast www.interscience.wiley.com/journal/jrs cell bodies and the surrounding medium. After segmentation, the particles in the binary image stacks were analyzed using a 3D objects counter developed for ImageJ.…”

Section: Data Acquisition and Image Evaluationmentioning

confidence: 99%

CARS microscopy of lipid stores in yeast: the impact of nutritional state and genetic background

Brackmann

Norbeck

Åkeson

et al. 2009

J Raman Spectroscopy

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We have developed a protocol for sub-micrometer resolved and chemically specific imaging of lipid storage in vivo employing coherent anti-Stokes Raman scattering (CARS) microscopy of one of the most important model organisms Saccharomyces cerevisiae -the yeast cell. By probing the carbon-hydrogen vibration using the nonlinear process of CARS, lipid droplets in the yeast cells clearly appear, as confirmed by comparative studies on relevant labeled organelles using two-photon fluorescence microscopy. From the images, unique quantitative data can be deduced with high three-dimensional resolution, such as the volume, shape, number, and intracellular location of the neutral lipid stores. We exemplify the strength and usability of the method for two cases: the impact on lipid storage of the nutritional condition (starvation and type of carbon source available) as well as of genetic modification of two fundamental metabolic regulation pathways involving carbohydrate and lipid storage (BCY1 and DGA1, LRO1, ARE1/2 deletions), respectively. While the impact of carbon source on the total cellular lipid volume was minimal, long-term starvation induces a significant accumulation of lipid droplets. We also confirm that the lipid-storage-deficient mutant is indeed unable to synthesize lipid droplets, and that the inability of the bcy1-mutant to store carbohydrates is compensated by a two-fold increase in stored neutral lipids. We note that there is a significant cell-to-cell variability in neutral lipid storage in general, i.e. that there is a correspondence to the noise found for gene expression also in lipidomics.

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Section: Data Acquisition and Image Evaluationmentioning

confidence: 99%

CARS microscopy of lipid stores in yeast: the impact of nutritional state and genetic background

Brackmann

Norbeck

Åkeson

et al. 2009

J Raman Spectroscopy

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“…A further relevant topic in improving the information content to be gained from the multimodal images is the development of image processing algorithms [52][53][54][55][56][57] optimized to address particular targets. Here we argue for the benefit of such approaches by turning the attention collagen structural alterations upon formation of keloids as shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

Towards multimodal nonlinear optical tomography - experimental methodology

et al. 2011

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All-optical microspectroscopic and tomographic tools reveal great potential for clinical dermatologic diagnostics, i.e., investigation of human skin and skin diseases. While optical-coherence tomography has been complemented by twophoton fluorescence tomography and second-harmonic generation tomography, a joint study of various nonlinear optical microspectroscopies, i.e., application of the recently developed multimodal imaging approach, to sizable human-tissue samples has not been evaluated up to now. Here, we present such multimodal approach combining different nonlinear optical contrast mechanisms for imaging, namely two-photon excited fluorescence (TPF), second-harmonic generation (SHG), and coherent anti-Stokes Raman scattering (CARS) into a joint microscopic experiment. We show the potential of imaging large skin areas and discuss the information obtained in a case study comparing normal skin and keloid tissue.

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“…In particular it is employed to estimate the sizes of the fat droplets observed in the CARS images. First, the recognition of the fat-containing cells has been automized by the application of a global thresholding approach using the watershed algorithm [48]. Assuming circular shapes a mean diameter of the fat-containing cells of 55 mm has been estimated in the high-contrast region of the image.…”

Section: Resultsmentioning

confidence: 99%

Multimodal imaging to study the morphochemistry of basal cell carcinoma

Vogler

Meyer

Akimov

et al. 2010

Journal of Biophotonics

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Basal cell carcinoma is the most abundant malignant neoplasm in humans, the pathology of which is characterized by an abnormal proliferation of basal cells. Basal cell carcinoma can show a variety of different morphologies, which are based on different cellular biology. Furthermore, the carcinoma often grows invisibly to the eye imbedded in the surrounding skin. Therefore, in some cases its clinical detection is challenging. Thus, our work aims at establishing an unsupervised tissue classification method based on multimodal imaging and the application of chemometrics to discriminate basal cell carcinoma from non‐diseased tissue. A case study applying multimodal imaging to ex‐vivo sections of basal cell carcinoma is presented. In doing so, we apply a combination of various linear and non‐linear imaging modalities, i.e. fluorescence, Raman and second‐harmonic generation microscopy, to study the morphochemistry of basal cell carcinoma. The joint information content obtained by such multimodal approach in studying various aspects of the malignant tissue alterations associated with basal cell carcinoma is discussed. (© 2010 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

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Image analysis in nonlinear microscopy

Cited by 15 publications

References 37 publications

CARS microscopy of lipid stores in yeast: the impact of nutritional state and genetic background

CARS microscopy of lipid stores in yeast: the impact of nutritional state and genetic background

Towards multimodal nonlinear optical tomography - experimental methodology

Multimodal imaging to study the morphochemistry of basal cell carcinoma

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