Flow cytometric analysis of the cell cycle in transfected cells without cell fixation

Dimitri G. Pestov, Marina Polonskaia, Lester F. Lau

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

We describe a simple and rapid protocol for the flow cytometric analysis of the cell cycle in transfected cells using a green fluorescent protein anchored in the intracellular membranes of the endoplasmic reticulum (ER- GFP) as a transfection marker. The transfected cells are analyzed by dual- parameter flow cytometry after a brief incubation with digitonin, followed by staining with propidium iodide. Treatment of cells with digitonin efficiently preserves the ER-GFP fluorescence and allows reproducible and quantitative DNA staining, thus obviating the need for cell fixation before flow cytometry. The digitonin-based protocol is faster and easier to perform than conventional cell fixation and is illustrated herein by cell cycle analyses of U2OS and NIH 3T3 cells.

Original languageEnglish (US)
Pages (from-to)102-106
Number of pages5
JournalBioTechniques
Volume26
Issue number1
DOIs
StatePublished - 1999
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Biochemistry, Genetics and Molecular Biology(all)

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