Abstract
This report describes an improved immunochemical procedure to stain cells in suspension for incorporated bromodeoxyuridine (BrdUrd) and total DNA content. The procedure consists of five steps: (1) chromatin proteins are extracted by treating with 0.1 M HCl and 0.7% Triton X‐100 to facilitate DNA denaturation and to minimize nonspecific staining; (2) cellular DNA is denatured by heating to 100°C in distilled water; (3) BrdUrd in single‐stranded DNA (ssDNA) is stained using an immunochemical procedure; (4) autofluorescence is reduced using sodium borohydride (NaBH4); and (5) DNA is stained with the fluorescent dye propidium iodide. With this procedure, the BrdUrd incorporated by CHO cells during periods as short as a few seconds can be detected using flow cytometry. In addition, the stoichiometry of the immunofluorescent staining procedure is high.
Original language | English (US) |
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Pages (from-to) | 235-239 |
Number of pages | 5 |
Journal | Cytometry |
Volume | 8 |
Issue number | 2 |
DOIs | |
State | Published - Mar 1987 |
Externally published | Yes |
Keywords
- Anti‐bromodeoxyuridine
- DNA
- flow cytometry
- stoichiometry
- thermal denaturation
ASJC Scopus subject areas
- Pathology and Forensic Medicine
- Biophysics
- Hematology
- Endocrinology
- Cell Biology