Single-section counting error when distinguishing between primordial and early primary follicles in sections of rat ovary of different thickness

The number of primordial follicles within an ovary is frequently determined by counting 5, 7 or 10 mu m thick sections and multiplying by the fraction of sections counted and a correction factor to adjust for duplicate counts . The objectives of the present study were: (i) to evaluate the accuracy of the correction factor developed by Abercrombie (1946); (ii) to evaluate th e accuracy of the classification of primordial follicles from single tissue sections; and (iii) to determine the incorporation rate of 5-bromo-2-deoxy uridine into primordial follicles. Ln Expt 1, rat ovaries were sectioned at a thickness of 5, 7 or 10 mu m. Primordial follicles were counted and clas sified across ten adjacent ovarian sections. The percentage of primordial f ollicles from single sections that were counted twice was 10, 9 and 2% in 5 , 7 and 10 mu m sections, respectively. This was lower than predicted by Ab ercrombie's method. The major error in counting from single sections was cl assification of early primary follicles as primordial follicles (55, 33 and 3% in 5, 7 and 10 mu m sections, respectively). Ln Expt 2, a mean of 12 +/ - 7% of primordial follicles incorporated 5-bromo-2-deoxyuridine after infu sion for 7 days (four of seven rats had no labelled primordial follicles). In conclusion: (i) Abercrombie's correction factor should not be used for a djusting counts of follicles; (ii) evaluation of primordial follicles from single sections gives inaccurate counts and incorrect classification is of greater importance than duplicate counting, particularly in thinner section s; (iii) for evaluation of the number of follicles, 10 mu m is the optimal thickness; and (iv) primordial follicles incorporated 5-bromo-2-deoxyuridin e infrequently.