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Updated: Aug 10, 2026
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Assessment of Global DNA Double-Strand End Resection using BrdU-DNA Labeling coupled with Cell Cycle Discrimination Imaging
Published on: April 28, 2021
Evaluation of intranuclear BrdU detection procedures for use in multicolor flow cytometry
Kristina Rothaeusler1, Nicole Baumgarth
1Center for Comparative Medicine, University of California, Davis, USA.
Summary
Paraformaldehyde/saponin cell preparation methods effectively preserve fluorescent staining for bromodeoxyuridine (BrdU) analysis. This enables detailed cell turnover studies in rare B cell subsets, revealing new functional insights.
Area of Science:
- Immunology
- Cell Biology
- Flow Cytometry
Background:
- Multicolor flow cytometry combined with bromodeoxyuridine (BrdU) incorporation aids cell proliferation studies.
- Harsh cell preparation methods may compromise fluorescent dye staining quality, impacting analysis of cell subsets.
Purpose of the Study:
- To compare cell preparation protocols for BrdU measurement.
- To assess the impact of cell preparation on fluorescent surface staining and scatter parameters.
- To evaluate BrdU measurement in small B lymphocyte subsets.
Main Methods:
- Compared three cell preparation protocols for BrdU measurement.
- Developed a 10-color fluorescent panel to test surface staining quality.
- Assessed BrdU measurement capability on splenic B cell subsets.
Main Results:
- All tested protocols impacted fluorescent and/or scatter parameters to some extent.
- Paraformaldehyde/saponin-based methods maintained sufficient fluorescent staining for BrdU analysis in all splenic B cell subsets.
- B-1a cell turnover rates were found to be similar to immature B cells and higher than other mature B cell subsets.
Conclusions:
- Paraformaldehyde/saponin-based cell preparation supports in vivo cell turnover studies in small cell subsets.
- These methods reveal novel functional information on rare cell populations.
