The aim of this study is to compare different laboratory tests, performed in order to evaluate the effect of some laboratory challenges on in vitro quality of frozen-thawed semen, and to compare the laboratory results with the in vivo semen fertility of a Timed-AI program. Semen doses of five different batches from a Brangus bull will be challenged and evaluated. The same semen batches that will be evaluated in vitro were previously utilized in a Timed-AI program in a commercial beef farm, which approximately 250 Brangus cows were inseminated. For each batch, three repetitions of each experimental design will be performed. Hence, for each semen doses, the following procedure will be performed: a 0.5 mL semen sample will be thawed at 37ºC for 30 sec (control group). Then, the following laboratory tests will be performed: sperm motility and vigor, Hypo-osmotic swelling test and sperm morphology. Then, an aliquot of 150 uL of the control sample will be incubated in a water bath at 45°C for 2 h (thermal challenge Group; GDT) and another aliquot of 150 uL of control sample will be centrifuged at 500 x g in Percoll gradient 45%/90% by 15 minutes (centrifugation challenge Group; GDC). In addition, the diluted and centrifuged semen (GDC) will be further submitted to thermal challenge, being incubated in a water bath at 45°C for 2 h (centrifugation + thermal challenge Group; GCDT). At the end of each challenge (GDT, and GCDT GDC), the same laboratory tests performed for the control group will be repeated. Finally, at the end of the present work, the results of control, GDT, GDC and GCDT groups will be compared with field fertility results (conception rate ate 45 days post-insemination) in order to evaluate, which laboratory challenge was better related with field bull fertility.
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