Progressive motility is a vital functional characteristic of ejaculated human spermatozoa that governs their ability to penetrate into, and migrate through, both cervical mucus and the oocyte vestments, and ultimately fertilize the oocyte. A detailed protocol, based on traditional manual/visual methods, is provided for performing an accurate four-category differential count including the reliable identification of rapid progressive (grade "a") spermatozoa-the most biologically, and hence clinically, important subpopulation. Thorough prior training and the use of a microscope fitted with a heated stage are both essential requirements for achieving accuracy and an acceptable uncertainty of measurement of no more than ±10%.