UNIT 8.6 Dynamic Proliferation Assessment in Flow Cytometry
Published Online: 1 SEP 2010
Copyright © 2010 by John Wiley & Sons, Inc.
Lab Protocol Title
Current Protocols in Cell Biology
How to Cite
Diermeier-Daucher, S. and Brockhoff, G. 2010. Dynamic Proliferation Assessment in Flow Cytometry. Current Protocols in Cell Biology. 48:8.6:8.6.1–8.6.23.
- Published Online: 1 SEP 2010
- Published Print: SEP 2010
Dynamic proliferation assessment via flow cytometry is legitimately supposed to be the most powerful tool for recording cell cycle kinetics in-vitro. The preeminent feature is a single cell–based multi-informative analysis by temporal high-resolution. Flow cytometric approaches are based on labeling of proliferating cells via thymidine substitution by a base analog (e.g., 5-bromo-2′-deoxyuridine, BrdU) that is added to cell cultures either for a short period of time (pulse labeling) or continuously until cell harvesting. This unit describes the alternative use of the thymidine analog 5-ethynyl-2′-deoxyuridine (EdU) in place of BrdU for three different applications: (1) dynamic proliferation assessment by EdU pulse cell labeling; (2) the same approach as (1) but in combination with live/dead cell discrimination; and (3) dynamic cell cycle analysis based on continuous cell labeling with EdU and Hoechst fluorochrome quenching. In contrast to the detection of BrdU incorporation, EdU-positive cells can be identified by taking advantage of click chemistry, which facilitates a simplified and fast cell preparation. Further analysis options but also limitations of the utilization of EdU are discussed. Curr. Protoc. Cell Biol. 48:8.6.1-8.6.23. © 2010 by John Wiley & Sons, Inc.
- cell proliferation;
- flow cytometry;
- cell cycle kinetics