Dataset Information

Mathematical imaging methods for mitosis analysis in live-cell phase contrast microscopy.

ABSTRACT: In this paper we propose a workflow to detect and track mitotic cells in time-lapse microscopy image sequences. In order to avoid the requirement for cell lines expressing fluorescent markers and the associated phototoxicity, phase contrast microscopy is often preferred over fluorescence microscopy in live-cell imaging. However, common specific image characteristics complicate image processing and impede use of standard methods. Nevertheless, automated analysis is desirable due to manual analysis being subjective, biased and extremely time-consuming for large data sets. Here, we present the following workflow based on mathematical imaging methods. In the first step, mitosis detection is performed by means of the circular Hough transform. The obtained circular contour subsequently serves as an initialisation for the tracking algorithm based on variational methods. It is sub-divided into two parts: in order to determine the beginning of the whole mitosis cycle, a backwards tracking procedure is performed. After that, the cell is tracked forwards in time until the end of mitosis. As a result, the average of mitosis duration and ratios of different cell fates (cell death, no division, division into two or more daughter cells) can be measured and statistics on cell morphologies can be obtained. All of the tools are featured in the user-friendly MATLAB®Graphical User Interface MitosisAnalyser.

SUBMITTER: Grah JS

PROVIDER: S-EPMC6414815 | biostudies-literature | 2017 Feb

REPOSITORIES: biostudies-literature

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Mathematical imaging methods for mitosis analysis in live-cell phase contrast microscopy.

Grah Joana Sarah JS Harrington Jennifer Alison JA Koh Siang Boon SB Pike Jeremy Andrew JA Schreiner Alexander A Burger Martin M Schönlieb Carola-Bibiane CB Reichelt Stefanie S

Methods (San Diego, Calif.) 20170209

In this paper we propose a workflow to detect and track mitotic cells in time-lapse microscopy image sequences. In order to avoid the requirement for cell lines expressing fluorescent markers and the associated phototoxicity, phase contrast microscopy is often preferred over fluorescence microscopy in live-cell imaging. However, common specific image characteristics complicate image processing and impede use of standard methods. Nevertheless, automated analysis is desirable due to manual analysi ...[more]

PMID: 28189773

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Project description:Reliable and reproducible cell therapy strategies to treat osteoarthritis demand an improved characterization of the cell and heterogeneous cell population resident in native cartilage tissue. Using live-cell phase-contrast time-lapse imaging (PC-TLI), this study investigates the morphological attributes and biological performance of the three primary biological objects enzymatically isolated from primary human cartilage: connective tissue progenitors (CTPs), non-progenitors (NPs) and multi-cellular structures (MCSs). The authors' results demonstrated that CTPs were smaller in size in comparison to NPs (P < 0.001). NPs remained part of the adhered cell population throughout the cell culture period. Both NPs and CTP progeny on day 8 increased in size and decreased in circularity in comparison to their counterparts on day 1, although the percent change was considerably less in CTP progeny (P < 0.001). PC-TLI analyses indicated three colony types: single-CTP-derived (29%), multiple-CTP-derived (26%) and MCS-derived (45%), with large heterogeneity with respect to cell morphology, proliferation rate and cell density. On average, clonal (CL) (P = 0.009) and MCS (P = 0.001) colonies exhibited higher cell density (cells per colony area) than multi-clonal (MC) colonies; however, it is interesting to note that the behavior of CL (less cells per colony and less colony area) and MCS (high cells per colony and high colony area) colonies was quite different. Overall effective proliferation rate (EPR) of the CTPs that formed CL colonies was higher than the EPR of CTPs that formed MC colonies (P = 0.02), most likely due to CTPs with varying EPR that formed the MC colonies. Finally, the authors demonstrated that lag time before first cell division of a CTP (early attribute) could potentially help predict its proliferation rate long-term. Quantitative morphological characterization using non-invasive PC-TLI serves as a reliable and reproducible technique to understand cell heterogeneity. Size and circularity parameters can be used to distinguish CTP from NP populations. Morphological cell and colony features can also be used to reliably and reproducibly identify CTP subpopulations with preferred proliferation and differentiation potentials in an effort to improve cell manufacturing and therapeutic outcomes.

Project description:The robustness of the phase retrieval methods is of critical importance for limiting and reducing radiation doses involved in x-ray phase contrast imaging. This work is to compare the robustness of two phase retrieval methods by analyzing the phase maps retrieved from the experimental images of a phantom.Two phase retrieval methods were compared. One method is based on the transport of intensity equation (TIE) for phase contrast projections, and the TIE-based method is the most commonly used method for phase retrieval in the literature. The other is the recently developed attenuation-partition based (AP-based) phase retrieval method. The authors applied these two methods to experimental projection images of an air-bubble wrap phantom for retrieving the phase map of the bubble wrap. The retrieved phase maps obtained by using the two methods are compared.In the wrap's phase map retrieved by using the TIE-based method, no bubble is recognizable, hence, this method failed completely for phase retrieval from these bubble wrap images. Even with the help of the Tikhonov regularization, the bubbles are still hardly visible and buried in the cluttered background in the retrieved phase map. The retrieved phase values with this method are grossly erroneous. In contrast, in the wrap's phase map retrieved by using the AP-based method, the bubbles are clearly recovered. The retrieved phase values with the AP-based method are reasonably close to the estimate based on the thickness-based measurement. The authors traced these stark performance differences of the two methods to their different techniques employed to deal with the singularity problem involved in the phase retrievals.This comparison shows that the conventional TIE-based phase retrieval method, regardless if Tikhonov regularization is used or not, is unstable against the noise in the wrap's projection images, while the AP-based phase retrieval method is shown in these experiments to be superior to the TIE-based method for the robustness in performing the phase retrieval.

Dataset Information

Mathematical imaging methods for mitosis analysis in live-cell phase contrast microscopy.

Publications

Mathematical imaging methods for mitosis analysis in live-cell phase contrast microscopy.

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