Invitrogen
CellTrace™ Violet Cell Proliferation Kit, for flow cytometry
C34571
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Establishing CRISPRa Screen Conditions to Enhance T cell Resistance Against Immunosuppressive Signals.
Cell Proliferation by Flow Cytometry
N/A
Suspend each vial in 20uL of DMSO, you can pool afterward and then freeze in aliquots at -20C (double-check the protocol that they didn’t change the vial size or so). Count cells, spin down the amount needed (calculate to lose 20% of cells by staining with CTV), resuspend at 1M/ml in PBS, add CTV (1:1,000 dilution), mix carefully, 20min 37C (incubator, exactly, CTV becomes toxic otherwise), fill up with cell culture media (at least 5x the volume, must contain FCS!) to stop the reaction, put at 37C for 5min, then spin down, remove sup completely, resuspend in assay media, count again. They should now be 100% CTV positive (it makes sense to check that when it’s your first time doing this protocol, so just compare some unstained vs CTV-stained cells by flow).
Here, T cell proliferation was assessed by CTV staining in presence of different concentrations of glucose. Primary human T cells cultured in absence of glucose proliferated the least. A dose-dependent proliferation was observed.
Reliable and good signal
Expensive
Great signal, highly recommended.