COMPARISON OF MOTILITY AND FLOW CYTOMETRIC ASSESSMENTS OF SEMINAL QUALITY IN FRESH, 24-HOUR EXTENDED AND CRYOPRESERVED HUMAN SPERMATOZOA

Functional differences among fresh 24-hour extended and cryopreserved human spermatozoa were assessed using both computer-assisted semen ana lysis (CASA) and flow cytometry. The objective was to determine if the re were interrelationships among various qualitative parameters of the fresh and treated samples when assessed by these two automated method s. Fertile donor specimens (n = 15) were split and examined for sperm motility and curvilinear velocity using CASA within 1 hour postejacula tion, after 24 hours in TEST-yolk buffer at 5 degrees C and after cryo preservation in TEST-yolk-glycerol medium. Flow cytometric analyses we re performed on 24-hour extended and cryopreserved (CP) samples after fluorescent staining with rhodamine 123 to quantify mitochondrial func tion and carboxydimethyl fluorescein diacetate and propidium iodide to assess plasma membrane integrity. The percentages of spermatozoa with functional mitochondria and intact membranes along with the proportio n of dead cells were identified and quantified by flow cytometry. Quad rant analyses of these data were used to determine the relative red an d green fluorescent intensities. The initial sperm motility was correl ated to the motility observed for the 24-hour stored and the CP sample s. The sperm velocity of both the initial and the 24-hour extended sam ples was correlated to the velocity of CP samples. As for the comparis on of the two automated methods for assessing seminal quality, the onl y sperm motion parameter that was correlated with a sperm population i dentified by flow cytometry (quadrant 4) was the curvilinear velocity of the sperm after 24 hours storage (r= 0.69) and after cryopreservati on (r = 0.74). The present findings indicate that additional research is needed to determine if prefreeze analyses of donor sperm could be u seful in predicting the post-thaw integrity of CP samples and, thereby , be useful in screening potential semen donors.