ANALYSIS OF CELL CYCLE-RELATED KI-67 AND P120 EXPRESSION BY FLOW CYTOMETRIC BRDURD-HOECHST 7AAD AND IMMUNOLABELING TECHNIQUE/

Flow cytometric multiparameter analysis of two proliferation associate d. antigens, Ki-67 and p120, was combined with cell cycle kinetic anal ysis, achieved by continuous labeling with 5-Bromodeoxy-uridine (BrdUr d), followed by staining with Hoechst 33258 and 7-Aminoactinomycin D ( 7AAD). Exponential and plateau phase monolayer cultures of the human b ladder carcinoma cell Line J82 were examined, Resting cells, character ized by their absent BrdUrd incorporation, showed no reactivity with t he MIB1 antibody, which was used for the detection of the Ki-67 antige n, Proliferating cells revealed a cell cycle phase dependent Ki-67 sta ining intensity, which was partially related to the time period spent in G1 after mitosis, In contrast to the Ki-67 antigen expression, no d ecrease in p120 immunofluorescence staining intensity of non-cycling c ells could be observed. We could demonstrate that a dissection of the history of cell replication, obtained by the BrdUrd/Hoechst technique combined with a simultaneous immunofluorescence staining reveals detai led information, on a single. cell level, about time dependent express ion of proliferation associated antigens in all cell cycle compartment s. (C) 1997 Wiley-Liss, Inc.