Physiology: Physiological Induction of the Acrosome Reaction in Human Sperm: Validation of a Microassay Using Minimal Volumes of Solubilized, Homologous Zona Pellucida

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Abstract
Purpose: The objective was to develop a method that couldaccommodate microvolumes of solubilized human (ZP) andsperm for assessing the induction of the acrosome reaction. Methods: A microassay using 1 μl of 2.5, 1.25, 0.6, 0.3,and 0.125 ZP/μl incubated with 1 μl of a highly motilesperm suspension for 60 min. As a control and parallelto the microassay a standard acrosome reaction techniquewas performed. Results: No significant differences were observed betweenthe percentage acrosome-reacted sperm reported by the twoassays under basal conditions (spontaneous) or after inductionwith a Ca2+ ionophore or solubilized ZP. At a ZPconcentration of 0.6 ZP/μl, the percentages ofacrosome-reacted spermatozoa in both techniques were significantlyhigher compared to the spontaneous acrosome reactionresults, namely 18% and 17%, compared to 10% and 10%,respectively. Approximately a 30% level of acrosomal exocytosis was induced with 2.5 ZP/μl in both methods. Conclusions: This newly devised microtechnique is easy andrapid to perform, is repeatable, and facilitates the use ofminimal volumes of solubilized human ZP (even a single ZP)for assessment of the inducibility of the acrosome reaction ofa homologous sperm population.

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