Description of an irnmunocytochemical evaluation technique for in vitro cell proliferation by bromodeoxiuridine incorporation

Abstract

In order to evaluate in vitro cell proliferation, an inmunocytochemistry non radioactive, easy, safe and fast technique, which detects bromodeoxiuridine taken by entering in S phase cells, was standardized. Several kinds of cells were used for peroxidase-inmunocytochemistry technique standardization. In all cases the same results were obtained. Briefly, BrdU was incubated for 48 hours. then cell cultures were processed for standard inmunocyiochemistry. An additional step for DNA denaturalization using HCI, before antibody incubation, was included, DA6 (di-amino-benzidine) or AEC (amino-ethyl-carbazol) were used As chromogens. This procedure results in a specific nuclei staining on proliferating cells and it allows quick evaluation of cell proliferation on cultures.