Evaluation of buffalo semen by Trypan blue/Giemsa staining and related fertilityin vitro

The aim of this work was to verify the feasibility of an easy, quick double staining technique for evaluation of frozen-thawed semen to predict the fertilizing capability in vitro of buffalo bulls. In Experiment 1, frozen-thawed semen from 6 bulls was stained with double Trypan blue/ Giemsa and the incidence of acrosome-intact live (AIL), acrosome-intact dead (AID), acrosome-lost live (ALL) and acrosome-lost dead (ALD) sperm was recorded. In Experiment 2, sperm from the same bulls were used to fertilize in vitro matured oocytes. The data obtained confirm that there is a strong “bull effect” in buffalo species, with differences in the percentage of AIL sperm at thawing, in cleavage and blastocyst rates among bulls. Interestingly, it was found that this staining technique can be used for a preliminary screening to select semen to use for IVF, as shown by the correlation existent between the percentages of acrosome-intact viable sperm cells at thawing and the blastocyst yields for 4/6 bulls.