A method for simultaneous quantification of monoclonal antibody Ki-67 and DNA content by flow cytometry. Application to breast carcinomas.

OBJECTIVE: To seek a method assessing Ki-67 immunostaining and DNA content by flow cytometry simultaneously. STUDY DESIGN: The murine monoclonal antibody Ki-67 (Ki) identifies a nuclear protein complex expressed by all nonquiescent tumor cells. Since the antigen detected by Ki is quite labile in most fixation and embedding protocols, a new method for simultaneous quantification of nuclear Ki immunofluorescence and DNA content by flow cytometry was developed. Unfixed, solid tumor cells are permeabilized only with saponin to preserve Ki antigen. The percentage of Ki-positive cell subpopulations calculated by subtraction of the related aspecific fixation histogram gives optimal results more rapidly than by cytogram analysis. Application to breast carcinoma shows the feasibility of the method. RESULTS: Significant correlations between Ki staining and the S-phase fraction were observed. Mean Ki labelling rates of aneuploid tumors were significantly higher than those of the diploid tumors, and significant differences between histologic types were found. CONCLUSION: This technique can be considered a fast, sensitive and optimal method to evaluate the proliferative activity of breast carcinomas and possibly of other solid tumors in a department of pathology.