Total number of particles in a bounded region estimated directly with the nucleator: Granulosa cell number in ovarian follicles

Abstract

Our objective was to evaluate the precision of the "nucleator" with regard to estimation of the number of granulosa cells in individual follicles and the average number of granulosa cells per follicle in a group of follicles. Nine murine ovarian follicles embedded in plastic were oriented isotropically with the "orientator" by Mattfeldt, cut into 20 microns serial sections, and stained with hematoxylin. In each follicle the number of granulosa cells was estimated in two ways, both unbiased: (1) with the nucleator principle in the section that contains the nucleolus of the oocyte and (2) in the complete set of follicle sections with the "fractionator" principle. Estimates with the fractionator were interpreted as "true" values. Therefore the precision (CE) of the nucleator estimates could be calculated. The nucleator provides precise estimates of the number of granulosa cells in one complete follicle: CE = 0.12. The amount of effort is minimal, because estimation is based on roughly 50 granulosa cells. The precision of the procedure with regard to estimation of the average number of granulosa cells in a group of follicles depends mainly on the biologic variation (CVbiol = 0.71).