Cell proliferation assay and cell cycle assay guide
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Learn about your options when you need a cell proliferation or cell cycle assay
These assays monitor the growth rate of a cell population, detect daughter cells in a growing population, or analyze the proportions of cells in different stages of the cell cycle.
Cell proliferation assay and cell cycle assay methods
- Incorporation of nucleoside analogs (eg BrdU and EdU) during DNA synthesis
- DNA staining dyes for cell cycle analysis
- Dye dilution assays
- Protein markers such as PCNA, Ki67 and MCM-2
- Clonogenicity assays
- Senescence assays
See below to learn about these types of assays. Our most popular assay kits include: EdU, propidium iodide, and CFSE.
Alternatively, cell proliferation can also be analyzed with cell viability assays that measure the rate of cellular metabolism, such as MTT, MTS, resazurin and similar assays, mitochondrial membrane potential dependent dyes, cellular esterase cleaved dyes, ATP and ADP assays, and assays that measure glycolytic flux and oxygen consumption.
Nucleoside analogs
Bromodeoxyuridine (BrdU) and ethynyldeoxyuridine (EdU) assays measure the incorporation of BrdU or EdU into newly synthesized DNA during DNA replication. Unlike BrdU, which is detected using antibodies, EdU can be easily directly labeled, either with a fluorescent dye or biotin for colorimetric or fluorometric detection via streptavidin-HRP. Edu staining is consistent with further antibody staining, unlike the harsher BrdU protocol.
Assay | Instrument | Assay kits |
EdU | Microscope, flow cytometry, plate reader | |
BrdU | Plate reader, microscope, flow cytometry |
DNA-staining dyes
DNA-staining dyes are commonly used in flow cytometry to measure the DNA content in cell populations and assay for cell cycle state. Propidium iodide is the mostly commonly used dye.
Dye | Instrument | Note | Assay kits |
Propidium iodide | Flow cytometer | Ex/Em 536/617 nm | |
Nuclear Green CCS1 | Ex/Em 490/525 nm | ||
Nuclear Red CCS1 | Ex/Em 490/620 nm | ||
DRAQ5TM | Ex/Em 633 & 647/665–800 nm | ||
DAPI | Ex/Em 358/461 | ||
Hoechst 33342 | Ex/Em 350/461 | ||
Hoechst 33258 | Ex/Em 350/461 | ||
7-AAD | Ex/Em 488/647 nm |
Dye dilution assays
The dyes in dye dilution assays are retained within cells over multiple generations. Daughter cells receive half of the dye of parent cells and assays are analyzed on a flow cytometer. Carboxyfluorescein succinimidyl ester (CFSE) is the longest established dye.
Assay | Instrument | Notes | Assay kits |
CFSE | Flow cytometer | Ex/Em 492/517 nm. Cytotoxic at higher concentrations. | |
CytoLabel Blue | Flow cytometer, microscope | Ex/Em 403/454 nm | |
CytoLabel Green | Ex/Em 511/525 nm | ||
CytoLabel Red | Ex/Em 628/643 nm | ||
CytoLabel Orange | Ex/Em 542/556 nm |
Protein markers
For the analysis of cell proliferation within tissue samples, or sometimes within cell culture, it is common to use antibodies to stain for the presence of Ki67, PCNA, or MCM-2.
Clonogenicity assays
Although little used at high throughput, the classical method of assaying cell proliferation is to use a clonogenic/clonogenicity assay. In this assay, cells are plated out at a low density and then the number of colonies formed is counted.
Senescence assays
The most common marker of senescent cells is the overexpression and accumulation of the endogenous lysosomal beta-galactosidase (SA-beta-gal). Beta-gal activity is detected using a colorimetric or fluorometric substrate.
Assay | Instrument | Assay kits |
Beta-gal | microscope, plate reader |
Learn more with our:
Cell viability assay guide
Measure the rate of continuing cellular activities, such as metabolism.
Cytotoxicity assay guide
Test for cell membrane damage, either by measuring the leakage of cellular enzymes or staining with membrane-impermeable dyes.
Apoptosis assay / cell death analysis guide
Measure the markers present in different types of cell death.