Description, Validation, and Performance Characteristics of a New Computer-Automated Sperm Motility Analysis System1

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Abstract
A computer-automated sperm motility assay (CASMA) system has been developed that provides a rapid and accurate analysis of multiple sperm movement parameters and a new measure of linearity, the linear deviation angle. CASMA provides objective, unbiased sampling and accurate quantitation of the movement characteristics of 200 sperm cells in 20 min. It consists of a microscope-mounted video camera, a high-resolution video disk recorder, a video digitizer/memory board mounted in an IBM 9000 microcomputer, and newly developed software. After manual recording, at 60 frames/s, of the video sequences (takes) of sperm suspensions, each take is automatically played back frame by frame, digitized, and stored in video memory. The software searches each frame, recognizes sperm cells, randomly selects a preset number for analysis, and traces each cell through the sequence to generate sperm "tracks" that are then stored in disk memory. This process is repeated for each take. Analysis of the stored tracks of each take yields the mean +/- SEM of the standard sperm motility parameters: percent motile (%M), curvilinear velocity (VC), net velocity (Vn), position-averaged velocity (Va), linear index (Vn/Va), progressiveness ration (Vn/Vc), and curvilinear progressiveness ratio (Va/Vc). Additionally, CASMA allows measurement of the linear deviation angle, a more direct measure of the linearity of sperm movement. For statistical comparisons, multiple takes can be considered either as replicates or separate experimental determinations. Finally, for more detailed analysis, each individually stored track, with its associated parameters, or histogram distributions of all sperm for each parameter can be displayed and printed. The performance of CASMA was evaluated by comparison of CASMA-determined movement parameters with manually determined values derived from the same sperm cells in the same video sequence and by comparison with published values determined using microcinematographic techniques. In each case, the CASMA values were essentially identical to those determined by manual measurements. Finally, CASMA accurately quantitates the linearity of sperm movement, a characteristic previously determined only by much more time-consuming methods. CASMA is a rapid and accurate system for measuring washed bull sperm motility and has reliably analyzed monkey and elephant sperm. The system has the potential to quantitate motility equally well with sperm from any species that have similar sperm head size.