A new quantitative test method for cell proliferation based on detection of the Ki-67 protein

A quantitative method to assess cell proliferation is one essential prerequ isite for testing biomaterial cytocompatibility in vitro. Currently used me thods, e.g. bromodeoxyuridine incorporation, show serious disadvantages con cerning either sensitivity, specificity or handling. A new enzyme linked im munosorbent assay (ELISA) system for the quantification of cell proliferati on based on detection of the Ki-67 protein is described. This protein has t urned out to be strictly correlated with the active parts of the cell cycle but to be absent in G(0). The measurement of Ki-67 expression by different human cell types, e.g. endothelial cells and HeLa cells, was evaluated in order to answer the question of whether the data obtained using the Ki-67 E LISA method correlate with the proliferation measured with flow cytometrica l DNA analysis and microscopical evaluation. Methods currently used for the evaluation of cell proliferation were compared to the new Ki-67 ELISA meth od. In addition, the functionality of adherent endothelial cells, and the v iability and morphology of the cells were investigated. Cells were treated with standard culture medium with or without the transcription inhibitor, a ctinomycin D, or growth factors, e.g. endothelial cell growth factor (ECGF) , and were exposed to metal ion standard solutions. These solutions were in a cytotoxic-non-cytotoxic range. Ki-67 ELISA was found to be a reliable qu antitative method to assess proliferation of cultured human cells in vitro. It has advantages over methods that are currently being used. It is easy t o perform and corresponds to the requirements for a test to be selected for biomaterial testing according to ISO standard 10 993. (C) 2000 Kluwer Acad emic Publishers.