Segmentation and analysis of mother machine data: SAM

Time-lapse imaging of bacteria growing in micro-channels in a controlled environment has been instrumental in studying the single cell dynamics of bacterial growth. This kind of a microfluidic setup with growth chambers is popularly known as mother machine [1]. In a typical experiment with such a set-up, bacterial growth can be studied for numerous generations with high resolution and temporal precision using image processing. However, as in any other experiment involving imaging, the image data from a typical mother machine experiment has considerable intensity fluctuations, cell intrusion, cell overlapping, filamentation etc. The large amount of data produced in such experiments makes it hard for manual analysis and correction of such unwanted aberrations. We have developed a modular code for segmentation and analysis of mother machine data (SAM) for rod shaped bacteria where we can detect such aberrations and correctly treat them without manual supervision. We track cumulative cell size and use an adaptive segmentation method to avoid faulty detection of cell division. SAM is currently written and compiled using MATLAB. It is fast (∼ 15 min/GB of image) and can be efficiently coupled with shell scripting to process large amount of data with systematic creation of output file structures and graphical results. It has been tested for many different experimental data and is publicly available in Github.

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Time-lapse imaging: the state of the art†

Biology of Reproduction ◽

10.1093/biolre/ioz035 ◽

2019 ◽

Vol 101 (6) ◽

pp. 1146-1154 ◽

Cited By ~ 4

Author(s):

Raquel Del Gallego ◽

José Remohí ◽

Marcos Meseguer

Keyword(s):

Embryo Development ◽

Clinical Success ◽

Time Lapse ◽

Embryo Cell ◽

Cell Dynamics ◽

Vitro Fertilization ◽

Cycle Lengths ◽

Novel Concept ◽

Time Lapse Imaging

Abstract The introduction of time-lapse imaging to clinical in vitro fertilization practice enabled the undisturbed monitoring of embryos throughout the entire culture period. Initially, the main objective was to achieve a better embryo development. However, this technology also provided an insight into the novel concept of morphokinetics, parameters regarding embryo cell dynamics. The vast amount of data obtained defined the optimal ranges in the cell-cycle lengths at different stages of embryo development. This added valuable information to embryo assessment prior to transfer. Kinetic markers became part of embryo evaluation strategies with the potential to increase the chances of clinical success. However, none of them has been established as an international standard. The present work aims at describing new approaches into time-lapse: progress to date, challenges, and possible future directions.

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An immobilization technique for long-term time-lapse imaging of explanted Drosophila tissues

10.1101/2020.08.04.234864 ◽

2020 ◽

Author(s):

Matthew P. Bostock ◽

Anadika R. Prasad ◽

Rita Chaouni ◽

Alice C. Yuen ◽

Rita Sousa-Nunes ◽

...

Keyword(s):

Time Lapse ◽

Tissue Level ◽

Anisotropic Pressure ◽

Biological Processes ◽

Effective Technique ◽

Cell Dynamics ◽

First Instar ◽

Stress Damage ◽

Time Lapse Imaging

AbstractTime-lapse imaging is an essential tool to study dynamic biological processes that cannot be discerned from fixed samples alone. However, imaging cell- and tissue-level processes in intact animals poses numerous challenges if the organism is opaque and/or motile. Explant cultures of intact tissues circumvent some of these challenges, but sample drift remains a considerable obstacle. We employed a simple yet effective technique to immobilize tissues in medium-bathed agarose. We applied this technique to study multiple Drosophila tissues from first-instar larvae to adult stages in various orientations and with no evidence of anisotropic pressure or stress damage. Using this method, we were able to image fine features for up to 18 hours and make novel observations. Specifically, we report that fibers characteristic of quiescent neuroblasts are inherited by their basal daughters during reactivation; that the lamina in the developing visual system is assembled roughly 2-3 columns at a time; that lamina glia positions are dynamic during development; and that the nuclear envelopes of adult testis cyst stem cells do not break down completely during mitosis. In all, we demonstrate that our protocol is well-suited for tissue immobilization and long-term live imaging, enabling new insights into tissue and cell dynamics in Drosophila.

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Multiplexed End-point Microfluidic Chemotaxis Assay using Centrifugal Alignment

10.1101/2020.07.09.195644 ◽

2020 ◽

Author(s):

Sampath Satti ◽

Pan Deng ◽

Kerryn Matthews ◽

Simon P. Duffy ◽

Hongshen Ma

Keyword(s):

Fluid Flow ◽

Drug Testing ◽

Time Lapse ◽

Cell Alignment ◽

Cell Geometry ◽

End Point ◽

Starting Point ◽

Correct Location ◽

Set Up ◽

Time Lapse Imaging

AbstractA fundamental challenge to multiplexing microfluidic chemotaxis assays at scale is the requirement for time-lapse imaging to continuously track migrating cells. Drug testing and drug screening applications require the ability to perform hundreds of experiments in parallel, which is not feasible for assays that require continuous imaging. To address this limitation, end-point chemotaxis assays have been developed using fluid flow to align cells in traps or sieves prior to cell migration. However, these methods require precisely controlled fluid flow to transport cells to the correct location without undesirable mechanical stress, which introduce significant set up time and design complexity. Here, we describe a microfluidic device that eliminates the need for precise flow control by using centrifugation to align cells at a common starting point. A chemoattractant gradient is then formed using passive diffusion prior to chemotaxis in an incubated environment. This approach provides a simple and scalable approach to multiplexed chemotaxis assays. Centrifugal alignment is also insensitive to cell geometry, enabling this approach to be compatible with primary cell samples that are often heterogeneous. We demonstrate the capability of this approach by assessing chemotaxis of primary neutrophils in response to an fMLP (N-formyl-met-leu-phe) gradient. Our results show that cell alignment by centrifugation offers a potential avenue to develop scalable end-point multiplexed microfluidic chemotaxis assays.

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Image and video analysis techniques for cellular microscopy

10.32469/10355/46443 ◽

2014 ◽

Author(s):

◽

Ilker Ersoy

Keyword(s):

Level Set ◽

Image Data ◽

Time Lapse ◽

Image Features ◽

Biological Knowledge ◽

Cell Detection ◽

Automated Classification ◽

Stepping Stones ◽

Cell Dynamics ◽

Microscopy Imaging

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] Advances in automated digital microscopy imaging made it possible to produce multi-dimensional image data that can capture dynamic characteristics of sub-cellular and cellular structures. Biologists routinely produce large volumes of bioimage time lapse data that necessitates automated algorithms for unbiased and repeatable quantitative analysis. These algorithms are the stepping stones in bioimage informatics to turn the image data into biological knowledge. Unique challenges posed by different imaging modalities and cell dynamics require a combination of accurate detection, segmentation, classification and tracking approaches tailored to address and exploit particular image characteristics. In this dissertation, we present algorithms for the analysis of microscopy image sequences to address these challenges. We propose a level set active contour approach to address accurate segmentation in phase-contrast as well as brightfield microscopy imaging that utilizes edge profiles. Our approach significantly outperforms traditional level set approaches. We show the applications of our approach to cell spreading analysis and red blood cell analysis with robust solutions for cell detection to delineate clustered cells. We also present two studies for automated classification of cells in fluorescence microscopy emphasizing the importance of choosing image features for the specific problem. Lastly, we present a fully automated cell detection and tracking approach tailored for muscle satellite cells that enables efficient and unbiased analysis of factors that promote cell motility.

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Cleavage of Human Embryos: Options and Diversity

Acta Naturae ◽

10.32607/20758251-2016-8-3-88-96 ◽

2016 ◽

Vol 8 (3) ◽

pp. 88-96

Author(s):

Yu. K. Doronin ◽

I. V. Senechkin ◽

L. V. Hilkevich ◽

M. A. Kurcer

Keyword(s):

Time Lapse ◽

Reproductive Technologies ◽

Human Embryos ◽

Imaging Data ◽

Noninvasive Methods ◽

Topographic Features ◽

Time Parameters ◽

Embryo Cleavage ◽

Time Lapse Imaging ◽

Developmental Dynamics

In order to estimate the diversity of embryo cleavage relatives to embryo progress (blastocyst formation), time-lapse imaging data of preimplantation human embryo development were used. This retrospective study is focused on the topographic features and time parameters of the cleavages, with particular emphasis on the lengths of cleavage cycles and the genealogy of blastomeres in 2- to 8-cell human embryos. We have found that all 4-cell human embryos have four developmental variants that are based on the sequence of appearance and orientation of cleavage planes during embryo cleavage from 2 to 4 blastomeres. Each variant of cleavage shows a strong correlation with further developmental dynamics of the embryos (different cleavage cycle characteristics as well as lengths of blastomere cycles). An analysis of the sequence of human blastomere divisions allowed us to postulate that the effects of zygotic determinants are eliminated as a result of cleavage, and that, thereafter, blastomeres acquire the ability of own syntheses, regulation, polarization, formation of functional contacts, and, finally, of specific differentiation. This data on the early development of human embryos obtained using noninvasive methods complements and extend our understanding of the embryogenesis of eutherian mammals and may be applied in the practice of reproductive technologies.

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