One-step collagenase method for zona pellucida removal in unfertilized eggs: easy and gentle method for large-scale preparation

Purpose  

Zona pellucida (ZP)-free eggs are often used for studies such as evaluating the interaction of sperm-oolemma. To acquire ZP-free eggs, the most commonly used methods employ acidified Tyrode’s solution, enzymatic digestion with a trypsin-like enzyme, or mechanical methods using micropipettes. However, acidified Tyrode’s solution and trypsin-like enzymes often damage the oolemma, especially when many eggs are treated at once for mass sample analyses. The mechanical method requires skill, and it is time-consuming to prepare many ZP-free eggs. Therefore, in this study, to establish an easy, reliable method for preparing ZP-free eggs, we examined the ZP digestion method originally reported by Zuccotti et al. (J Reprod Fertil 93:515–520, 1991) that uses collagenase.