Immunoenzymatic detection of the new proliferation associated protein p100 by means of a cellular ELISA: specific detection of cells in cell cycle phases S, G2 and M.

In vitro proliferation assays are widely used in biomedical research. We describe the immunoenzymatic (ELISA) detection of a recently described proliferation associated protein (p100) by means of a new monoclonal mouse IgG1 antibody (Ki-S2). P100 is a 100 kDa nuclear protein that is specifically detected during the cell cycle phases S, G2 and M. Comparative studies on lectin-stimulated leukocytes using 3H-thymidine labelling and Ki-67 antibodies revealed a statistically significant positive correlation. Since p100 is absent in GO and G1 cells, its detection permits the precise and specific measurement of actual cell cycle events under culture conditions.

Improved ELISA proliferation assay (EPA) for the detection of in vitro cell proliferation by a new Ki-67-antigen directed monoclonal antibody (Ki-S3).

We describe a simplified and improved proliferation assay based on a conventional ELISA system for the in vitro measurement of cellular proliferation (ELISA proliferation assay = EPA). The assay is based on a new monoclonal antibody (Ki-S3) to the proliferation-specific Ki-67-antigen and is carried out in 96-well microtiter plates using conventional immunoenzymatic methods. Ki-S3 is an immunoprecipitating monoclonal mouse IgG1 antibody, which recognizes a formalin-resistant epitope of the Ki-67 antigen. It can be used to measure proliferating cells in the cell cycle phases G1, S, G2 and M. In phytohemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMC) the absorbance values obtained with the EPA show a statistically significant correlation to the number of Ki-S3 positive cells in simultaneously immunostained cytospin slides (r = 0.88). A direct comparison with [3H]thymidine labeling reveals the test to be an equally sensitive method for monitoring cellular proliferation (r = 0.91). This assay is an improved ELISA proliferation assay, which is easy to perform, does not require time-consuming pretreatments and avoids the hazards of radioactive isotopes.