Quantitation of AgNORs by flow versus image cytometry

AgNORs are nucleolar proteins that interact specifically with silver salts. The size of silver precipitates measured by image analysis (ICM) in cyclin g cells proved to be inversely proportional to the cell cycle time and prov ided a significant correlation with prognosis for a large spectrum of cance rs. Because ICM is time-consuming and poorly reproducible among laboratorie s using different imaging settings, this article presents a new approach to AgNOR quantitation based on flow cytometry (FCM). We report that silver pr ecipitates caused a great decrease in the forward scattered light and that this effect was correlated with the AgNOR's relative area as measured by IC M. These results were confirmed by measuring cell lines having different ce ll cycle durations. Moreover, double staining using APase-Fast red fluoresc ence to reveal the Ki-67/MIB 1 antigen of cycling cells and silver nitrate to stain the AgNORs was successfully analyzed by FCM. The procedure makes i t possible, for the first time, to validly and rapidly compare the growth f raction and cycling speed of partially proliferating cell populations, such as tumors.