Semen evaluation methodology is complex and difficult to standardize. Rigorously standardized laboratory protocols and strict quality control (QC) are essential for meaningful comparison of data from multiple sites. We describe the methods used for determination of semen volume, sperm concentration, and percent sperm motility in the Study for Future Families, a multicenter study of semen quality in the United States. Each of these 3 semen parameters was assessed using 2 techniques, which provided the opportunity to compare precision and assess suitability for multicenter studies. Detailed protocols were used, and technicians were centrally trained. A total of 509 semen evaluations were performed. Semen volume measured by weight was greater (P <.0001) than that determined by pipetting (3.7 +/- 1.6 mL vs 3.2 +/- 1.6 mL). Sperm concentration determined using hemacytometer chambers was consistently higher (P <.001) than that using disposable MicroCell chambers (81.0 x 10(6)/mL vs 65.9 x 10(6)/mL). Precision was slightly greater for the MicroCell chamber. The percentage of motile sperm was assessed by a simple counting technique as well as by the World Health Organization categorical method that assigns individual motile sperm to "a," "b," and "c" categories on the basis of progression. When these 3 categories were collapsed, the methods provided values that were not statistically different (P >.05), although the collapsed values tended to be higher (58.1% vs 51.6%) and less precise (CV 7.7% vs 4.1%) for the categorical method than for motility determined using the simple method. The data obtained in this study demonstrate the critical need for rigorous standardization of protocols and techniques for multicenter studies.