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When Phase Contrast Fails: ChainTracer and NucTracer, Two ImageJ Methods for Semi-Automated Single Cell Analysis Using Membrane or DNA Staining

Lookup NU author(s): Simon Syvertsson, Dr Leendert Hamoen

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This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).


Abstract

Within bacterial populations, genetically identical cells often behave differently. Single-cell measurement methods are required to observe this heterogeneity. Flow cytometry and fluorescence light microscopy are the primary methods to do this. However, flow cytometry requires reasonably strong fluorescence signals and is impractical when bacteria grow in cell chains. Therefore fluorescence light microscopy is often used to measure population heterogeneity in bacteria. Automatic microscopy image analysis programs typically use phase contrast images to identify cells. However, many bacteria divide by forming a cross-wall that is not detectable by phase contrast. We have developed 'ChainTracer', a method based on the ImageJ plugin ObjectJ. It can automatically identify individual cells stained by fluorescent membrane dyes, and measure fluorescence intensity, chain length, cell length, and cell diameter. As a complementary analysis method we developed 'NucTracer', which uses DAPI stained nucleoids as a proxy for single cells. The latter method is especially useful when dealing with crowded images. The methods were tested with Bacillus subtilis and Lactococcus lactis cells expressing a GFP-reporter. In conclusion, ChainTracer and NucTracer are useful single cell measurement methods when bacterial cells are difficult to distinguish with phase contrast.


Publication metadata

Author(s): Syvertsson S, Vischer NOE, Gao YQ, Hamoen LW

Publication type: Article

Publication status: Published

Journal: PLoS One

Year: 2016

Volume: 11

Issue: 3

Online publication date: 23/03/2016

Acceptance date: 25/02/2016

Date deposited: 14/06/2016

ISSN (electronic): 1932-6203

Publisher: Public Library of Science

URL: http://dx.doi.org/10.1371/journal.pone.0151267

DOI: 10.1371/journal.pone.0151267


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Funding

Funder referenceFunder name
Biotechnology and Biological Sciences Research Council (BBSRC)
12128Stichting voor de Technische Wetenschappen (STW) Vici grant

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