Objective-To investigate whether and to what extent the two major AgNOR proteins, nucleolin and protein B23, are maintained after one cell division in proliferating cells.Design-Using three asynchronously growing human cancer cell lines, TG, SJNKP, and CHP 212 cells, nucleolin and protein B23 were first identified on SDS-polyacrylamide separated nucleolar proteins, transferred to nitrocellulose and silver stained for AgNOR proteins. Measurement of doubling time indicated a period very close to 24h for each of the cell lines. To quantify the percentage of nucleolin and protein B23 maintained in daughter cells after duplication, cells were labelled with [(35)S]-methionine and a 24h cold chase performed. Nucleolin and protein B23 labelling was evaluated by densitometric analysis on nitrocellulose autoradiograms.Results-The radioactivity relative to nucleolin and protein B23 bands maintained in the daughter cells was a constant fraction of that present before cell duplication. In the three cell lines the percentage of residual radioactivity measured in the nucleolin bands was 42.2, 40.6, and 41.2 and in protein B23 bands 48.0, 46.2, and 44.1.Conclusions-After one cell division the nucleolin and protein B23 quantity present in cells may be highly variable, depending on the amount of the two proteins present in the mother cell. This is important in relation to the correct utilisation of AgNOR protein quantity as an index for evaluating cell kinetics.