FACTORS INFLUENCING HUMAN SPERM KINEMATIC MEASUREMENTS BY THE CELLTRAK COMPUTER-ASSISTED SPERM ANALYSIS SYSTEM

This study examines the effect of varying several factors, both extrin sic and technology-dependent, on the reconstruction of human sperm tra jectories and the derived kinematic measurements using videotapes and the Motion Analysis Celltrak/S instrument. In semen samples from norma l healthy men, curvilinear (VCL) and straight line velocities (VSL) we re found to increase 1.5-fold, and linearity (LIN) of trajectories and amplitude of lateral head displacement (ALH) increased 1.25-fold when the temperature of analysis was raised from 24 to 37 degrees C, Only VCL and VSL were found to increase significantly between 24 and 37 deg rees C for sperm samples selected by Percoll gradient and incubated in a capacitating medium. An analysis chamber of 20 mu m depth was found to be adequate for seminal sperm samples while for Percoll-selected s perm samples the analysis in a 50 mu m depth provided the highest prop ortions of spermatozoa with the highest VCL and the largest ALH, The g rey level detection threshold required careful adjustment: using a thr eshold lower than the optimal threshold produced spurious sperm trajec tories for seminal sperm samples and rejected some trajectories for Pe rcoll-selected sperm samples, Definition of the appropriate frame rate and maximum burst speed was critical for valid trajectory reconstruct ion and therefore adequate derived kinematic measurements. Optimal val ues of these parameters were found to be 30 Hz and 400 mu m/s for semi nal spermatozoa and 60 Hz and 700 mu m/s for selected spermatozoa, The optimal values of 'ALH path-smoothing factor' used to calculate avera ge path and ALH were 5-10 points for seminal spermatozoa analysed at 3 0 Hz and 15-20 points for selected spermatozoa analysed at 60 Hz. We p ropose a set of standard conditions for reliable kinematic analysis of human spermatozoa using the Celltrak/S system.