A flow cytometric assay for the determination of cell proliferation with a monoclonal antibody directed against DNA-methyltransferase.
The enzyme DNA-methyltransferase is responsible for the methylation of a newly synthesized DNA-strand. A monoclonal antibody directed against DNA-methyltransferase was used to determine cell proliferation by means of flow cytometry. The reactivity of DNA-methyltransferase antibody was compared with the known proliferation markers transferrin-receptor and Ki67. All three methods showed comparable reactivity with the erythroblastic cell line K562 (86%, 81%, 76% respectively). In a second set of experiments peripheral blood mononuclear cells were stimulated with phytohaemagglutinin; in three experiments, a mean of 63% of the cells reacted with DNA-methyltransferase antibody after 72 h of culture as compared to a mean of 6% in the case of unstimulated control cells. HL60 cells were incubated with DMSO and harvested on day 5 of culture. The results obtained show that in differentiated cells the fraction positive with DNA-methyltransferase antibody decreased to levels below 10%. It is concluded that the technique described is a fast and easy method for the flow cytometric determination of cellular proliferation.
Universitätsklinikum Rudolf-Virchow, Abteilung Innere Medizin, Berlin, F.R.G.
A
A
Neubauer
S
S
Serke
W
W
Siegert
W
W
Kroll
R
R
Musch
D
D
Huhn
D000911:Antibodies, Monoclonal; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000378:metabolism*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D010835:Phytohemagglutinins / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000187:drug effects
D000911:Antibodies, Monoclonal; D010835:Phytohemagglutinins; D004248:DNA (Cytosine-5-)-Methyltransferases; D004121:Dimethyl Sulfoxide