A flow cytometric assay for the determination of cell proliferation with a monoclonal antibody directed against DNA-methyltransferase.

Abstract

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.