Dataset Information

Cell Membrane Tracking in Living Brain Tissue Using Differential Interference Contrast Microscopy.

ABSTRACT: Differential interference contrast (DIC) microscopy is widely used for observing unstained biological samples that are otherwise optically transparent. Combining this optical technique with machine vision could enable the automation of many life science experiments; however, identifying relevant features under DIC is challenging. In particular, precise tracking of cell boundaries in a thick ( ) slice of tissue has not previously been accomplished. We present a novel deconvolution algorithm that achieves the state-of-the-art performance at identifying and tracking these membrane locations. Our proposed algorithm is formulated as a regularized least squares optimization that incorporates a filtering mechanism to handle organic tissue interference and a robust edge-sparsity regularizer that integrates dynamic edge tracking capabilities. As a secondary contribution, this paper also describes new community infrastructure in the form of a MATLAB toolbox for accurately simulating DIC microscopy images of in vitro brain slices. Building on existing DIC optics modeling, our simulation framework additionally contributes an accurate representation of interference from organic tissue, neuronal cell-shapes, and tissue motion due to the action of the pipette. This simulator allows us to better understand the image statistics (to improve algorithms), as well as quantitatively test cell segmentation and tracking algorithms in scenarios, where ground truth data is fully known.

SUBMITTER: Lee J

PROVIDER: S-EPMC5839128 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

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Cell Membrane Tracking in Living Brain Tissue Using Differential Interference Contrast Microscopy.

Lee John J Kolb Ilya I Forest Craig R CR Rozell Christopher J CJ

IEEE transactions on image processing : a publication of the IEEE Signal Processing Society 20180401 4

Differential interference contrast (DIC) microscopy is widely used for observing unstained biological samples that are otherwise optically transparent. Combining this optical technique with machine vision could enable the automation of many life science experiments; however, identifying relevant features under DIC is challenging. In particular, precise tracking of cell boundaries in a thick ( ) slice of tissue has not previously been accomplished. We present a novel deconvolution algorithm that ...[more]

PMID: 29346099

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Project description:UnlabelledThe Gram-positive spore-forming anaerobe Clostridium difficile is a leading cause of nosocomial diarrhea. Spores of C. difficile initiate infection when triggered to germinate by bile salts in the gastrointestinal tract. We analyzed germination kinetics of individual C. difficile spores using Raman spectroscopy and differential interference contrast (DIC) microscopy. Similar to Bacillus spores, individual C. difficile spores germinating with taurocholate plus glycine began slow leakage of a ∼15% concentration of a chelate of Ca(2+) and dipicolinic acid (CaDPA) at a heterogeneous time T1, rapidly released CaDPA at Tlag, completed CaDPA release at Trelease, and finished peptidoglycan cortex hydrolysis at Tlysis. T1 and Tlag values for individual spores were heterogeneous, but ΔTrelease periods (Trelease - Tlag) were relatively constant. In contrast to Bacillus spores, heat treatment did not stimulate spore germination in the two C. difficile strains tested. C. difficile spores did not germinate with taurocholate or glycine alone, and different bile salts differentially promoted spore germination, with taurocholate and taurodeoxycholate being best. Transient exposure of spores to taurocholate plus glycine was sufficient to commit individual spores to germinate. C. difficile spores did not germinate with CaDPA, in contrast to B. subtilis and C. perfringens spores. However, the detergent dodecylamine induced C. difficile spore germination, and rates were increased by spore coat removal although cortex hydrolysis did not follow Trelease, in contrast with B. subtilis. C. difficile spores lacking the cortex-lytic enzyme, SleC, germinated extremely poorly, and cortex hydrolysis was not observed in the few sleC spores that partially germinated. Overall, these findings indicate that C. difficile and B. subtilis spore germination exhibit key differences.ImportanceSpores of the Gram-positive anaerobe Clostridium difficile are responsible for initiating infection by this important nosocomial pathogen. When exposed to germinants such as bile salts, C. difficile spores return to life through germination in the gastrointestinal tract and cause disease, but their germination has been studied only with population-wide measurements. In this work we used Raman spectroscopy and DIC microscopy to monitor the kinetics of germination of individual C. difficile spores, the commitment of spores to germination, and the effect of germinant type and concentration, sublethal heat shock, and spore decoating on germination. Our data suggest that the order of germination events in C. difficile spores differs from that in Bacillus spores and provide new insights into C. difficile spore germination.

Dataset Information

Cell Membrane Tracking in Living Brain Tissue Using Differential Interference Contrast Microscopy.

Publications

Cell Membrane Tracking in Living Brain Tissue Using Differential Interference Contrast Microscopy.

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