FERTILIZATION POTENTIAL AND QUALITATIVE CHARACTERISTICS OF HUMAN SPERMATOZOA AFTER SHORT-TERM CRYOSTORAGE AT 5-DEGREES-C IN 2 DIFFERENT TEST-YOLK BUFFER PREPARATIONS

Use of the media TEST-yolk buffer (TYB) in semenology today enables th e short-term incubation and cryostorage of spermatozoa and its subsequ ent use in the various assisted reproductive technologies (ART). Prepa ration of TYB media involves the addition of egg yolk (20% v/v) to a p hysiological solution of the zwitterion buffers TEX and Tris. The TYB is usually thermoprecipitated to remove the majority of the egg yolk g lobules and other macromolecules;from the medium. However, removal of these egg yolk constituents could possibly eliminate or reduce essenti al factors that could enhance the sperm viability and fertilization po tential after short-term dilution and storage. Improvements in the qua lity of the TYB could add greater benefits to those techniques employe d in the various forms of ART. The objectives of the investigation wer e 1) to study the sperm qualitative characteristics following short-te rm cryostorage at 5 degrees C in either thermoprecipitated (T-TYB) or non-thermoprecipitated (NT-TYB), and 2) to compare the fertilizing pot ential of spermatozoa stored for 24 hours at 5 degrees C in the two TY B preparations. In Experiment 1, semen specimens from 15 patients mere collected, assessed and split into two aliquots. Sperm specimens were processed by diluting 1:1 (v/v) with the T-TYB or NT-TYB, followed by centrifugation and reconstitution of the specimen to its initial volu me with the corresponding TYB medium. Sperm specimens were cryostored for 1, 2, 24, 48 and 72 hours. Samples were taken at each interval and placed in a 37 degrees C water bath and allowed to warm for 15 minute s after each cryostorage interval. Semen specimens mere assessed for p ercentage and grade of motility. The results of this study indicated t hat, although the NT-TYB yielded better results than the T-TYB, overal l those differences were not statistically significant. In Experiment 2, the fertilization potential of spermatozoa recovered after 24 hours of cryostorage in the two TYB preparations and further prepared via f iltration, was assessed by the sperm penetration assay (SPA) using zon a-free hamster oocytes. The average penetration rate (PR) and penetrat ion index (PI) were significantly better for the NT-TYB than for the T -TYB. The PR was 54% vs. 25%, and the PI 0.78 and 0.27 for spermatozoa incubated in the NT-TYB vs. T-TYB. The range of penetration was also much lower for the T-TYB (6 to 100%) preparation when compared to the NT-TYB (22 to 100%). The highest penetrator showed 100% for both prepa rations. However, the lowest penetrator showed 6% for the T-TYB and 22 % for the NT-TYB. The data obtained in this study suggest that both TY B preparations can be employed in short-term cryostorage (5 degrees C) of human spermatozoa and can adequately maintain the qualitative char acteristics of those spermatozoa. The data also showed that the NT-TYB preparation yielded sperm samples of higher fertilization potential, thus possibly establishing the superior usefulness of the NT-TYB in an ART program. (C) 1998 Tohoku University Medical Press.