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Objective: To determine if TEST-yolk buffer, consisting of TES (N-tris [hydroxymethyl]-methyl-2-aminoethanesufonic acid), Tris (Tris[hydroxymethyl]aninomethane), and chicken egg yolk, affects the presence of antisperm antibodies on the sperm surface as detected by the immunobead test., Design: A prospective study of antisperm antibodies on sperm surface before and after incubation in TEST-yolk buffer. Direct immunobead test and indirect immunobead test were done the day of collection of the semen sample to detect the presence of human immunoglobulin class G (IgG) and immunoglobulin class A (IgA); immunobead tests were repeated on the same sperm samples after 24 hours of storage in TEST buffer., Setting: Academic tertiary institution., Participants: Patients undergoing evaluation for infertility., Results: There was no significant difference in the outcome of the direct immunobead test after extending semen samples with TEST-yolk buffer for 24 hours at 4 degrees C. Eleven samples that were initially negative for IgG and 13 samples that were negative for IgA remained negative after 24-hour storage in TEST-yolk buffer. Eleven samples that were positive for IgG and nine samples that were positive for IgA by the direct immunobead test the first day remained positive the next day. Five extended sperm samples used in the indirect immunobead test with IgG positive serum gave positive results and four of five used with IgA positive serum gave positive results., Conclusions: These findings suggest that TEST-yolk buffer can be used to extend semen without affecting the presence of antibodies on the sperm surface as indicated by the direct immunobead test. The higher variability of the indirect immunobead tests indicates there may be some alteration of sperm antigens after storing in TEST-yolk buffer. These findings indicate that TEST-yolk buffer can be used to store semen for batched processing of samples or as a transport medium for delivery to a central laboratory for antibody testing.

Objective: To study the effect of human follicular fluid (FF) and the specific contribution of its epidermal growth factor (EGF) component on the in vitro maturation of cumulus-enclosed mouse oocytes., Design: A previously described mouse oocyte model system was used to study the effect of FF on oocyte maturation before and after extraction of EGF by immunoprecipitation. Follicular fluid specimens enclosing both mature and immature human oocytes were tested., Main Outcome Measures: The endpoints assessed were the percentage of oocytes undergoing germinal vesicle breakdown (GVBD) and polar body one formation at different intervals over a 24-hour period and the final degree of cumulus expansion achieved., Results: A concentration-related stimulatory effect of mature FF was noted when compared with the spontaneous increase of GVBD and polar body one formation observed for the EGF-free control medium. Overall, the effect of immature FF was inhibitory. After extraction of EGF from FF by immunoprecipitation from both immature and mature FF, the rates of GVBD and polar body one formation were decreased in both groups. The addition of 5 ng/mL of EGF to the extracted groups reversed this effect on polar body one formation. Cumulus expansion was maximal for oocytes incubated with mature FF and minimal for those incubated with EGF-free media., Conclusions: The positive effect of mature human FF on mouse oocyte maturation and cumulus expansion is to a large extent because of the presence of EGF.

Objective: Immature mammalian oocytes cultured in vitro undergo inadequate cytoplasmic maturation and hence have a limited potential for fertilization. Our primary objective was to determine if the addition of epidermal growth factor (EGF) to the in vitro culture system would have a positive effect on oocyte cytoplasmic maturation., Design: We studied the effect of different EGF concentrations on both denuded and cumulus-enclosed mouse oocytes cultured in vitro., Main Outcome Measures: The percentage of oocytes undergoing germinal vesicle breakdown (GVBD) and polar body one formation over time as a function of EGF concentration was determined., Results: A dose-related positive effect of EGF on both GVBD and polar body one formation over time was observed for mouse oocytes. As well, a similar effect of EGF was seen on immature human oocytes that had not been stimulated with exogenous gonadotropins., Conclusions: The use of EGF may allow for the performance of successful in vitro fertilization procedures using immature human oocytes retrieved during unstimulated cycles.

Nicotine at concentrations found in the cervical mucus of female smokers appeared to enhance in vitro human sperm penetrability through ovulatory bovine cervical mucus. Sperm motility parameters were not affected by the addition of nicotine to semen samples incubated with BWW medium. Overall, these results suggest that a direct inhibitory effect of nicotine on sperm penetrability through cervical mucus is not responsible for the apparent increase in cervical factor infertility present in smoking women.

Fetal fluid was studied in mice of the Q, C57BL/McL, C3H/Bi/McL and JU/Fa strains. The amount increases to day 16 of gestation, and then decreases. Amniotic fluid and fetal plasma contain three transferrins, five alpha-fetoproteins and albumin. Protein concentration in both increases during gestation; glucose decreases in amniotic fluid although it rises in fetal plasms; urea levels remain constant. The regulation of volume and biochemical composition of amniotic fluid, and its relation to the yolk-sac membrane, are discussed.

The relation of extra-embryonic fluid weight to litter number and foetal and placental weight was studied in mice on the 18th day of pregnancy, in litters both of experimentally reduced and of normal number. Partial regression analysis showed that litter number and foetal weight both exerted a negative effect on fluid weight; placental weight had no significant effect. Increased foetal weight reduced weight locally; on the other hand the effect of litter number was exerted systemically, throughout both horns of the uterus.

Cumulus enclosed primary oocytes from 2 to 4-mm bovine follicles were matured in vitro in Minimum Essential Medium containing follicle-stimulating hormone (0, .1, 1, 10, 50, or 100 micrograms/ml) or human chorionic gonadotropin (0, .1, 1, or 10 IU/ml) for 48 h at 37 degrees C under paraffin oil. Cumulus mass expansion comparable to that seen in vivo occurred in 18% of the control oocytes, 39% of those cultured in human chorionic gonadotropin, and 56% of those cultured in follicle-stimulating hormone. The optimum follicle-stimulating hormone concentration for cumulus expansion was 1 microgram/ml, and this was then used to mature oocytes individually or in groups of 5 for in vitro fertilization. Ejaculated bovine semen, extended 1:10 with yolk-TES-Tris extender and stored 24 to 48 h at 4 degrees C, was warmed, washed once with Minimum Essential Medium, and 500,000 motile sperm/ml were used to inseminate the matured oocyte-cumulus cell complexes. Criteria for fertilization was cleavage to the two-cell stage 48 h after insemination. Oocytes, inseminated individually, cleaved with a frequency of 5%, whereas 15% of those inseminated in groups of 5 cleaved, perhaps as the result of cumulus factors enhancing capacitation. The cleavage rate for the parthenogenetic control with killed spermatozoa was 0%. Therefore, primary oocytes matured in vitro to secondary oocytes were successfully fertilized in vitro and cleaved to at least the two-cell stage in the Minimum Essential Medium. Individual differences between bulls in ability to fertilize in vitro were noted.