Zinc fixation preserves flow cytometry scatter and fluorescence parameters and allows simultaneous analysis of DNA content and synthesis, and intracellular and surface epitopes.

Cytometry. Part a : the Journal of the International Society for Analytical Cytology
Uffe Birk JensenRikke Christensen

Abstract

Zinc salt-based fixation (ZBF) has proved advantageous in histochemical analyses conducted on intact tissues but has not been exploited in flow cytometry procedures that focus on quantitative analysis of individual cells. Here, we show that ZBF performs equally well to paraformaldehyde in the preservation of surface epitope labeling and forward and side scatter parameters as measured by flow cytometry. ZBF-fixed mouse epithelial keratinocytes exhibit a staining pattern for the surface markers Sca-1, CD34 and alpha6 integrin that is highly analogous to live cells. Furthermore, ZBF also preserves DNA allowing subsequent quantitative PCR analysis or labeling for incorporation of the thymidine analog EdU following surface and intracellular epitope staining. Finally, ZBF treatment allows for long-term storage of labeled cells with little change in these parameters. Thus, we present a protocol for zinc salt fixation of cells that allows for the simultaneous analysis of DNA and intracellular and cell surface proteins by flow cytometry.

References

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Citations

May 20, 2011·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Hong ZhaoZbigniew Darzynkiewicz
Aug 30, 2012·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Yanli SunJinming Li
Mar 23, 2011·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Zbigniew DarzynkiewiczJiangwei Li
Jul 28, 2016·Stem Cell Research·Anders Patrik GunnarssonUffe Birk Jensen
Feb 13, 2013·Laboratory Investigation; a Journal of Technical Methods and Pathology·Robert D CardiffAlexander D Borowsky
May 28, 2019·Frontiers in Cell and Developmental Biology·Ashley WaldronAlicia Ebert
Jul 7, 2011·Current Protocols in Cytometry·Rikke ChristensenUffe Birk Jensen
Jul 12, 2017·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Jacqueline M GertzBeth A Bouchard
Oct 5, 2021·Bioanalysis·Anusree Nair, Sonal M Manohar

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