alexa Evaluation of acrosome reaction and viability of human sperm with two fluorescent dyes.
Molecular Biology

Molecular Biology

Journal of Cytology & Histology

Author(s): Ozaki T, Takahashi K, Kanasaki H, Miyazaki K

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Abstract OBJECTIVE: To evaluate the usefulness of the Hoechst 33258/FITC-Pisum sativum agglutinin (FITC-PSA) staining for simultaneous assessment of viability and acrosome reaction rate (\%AR) of human sperm. MATERIAL AND METHODS: Fresh sperm was collected 13 fertile donors provided fresh semen. We used motile sperm selected by the "swim-up" procedure using modified HTF. Hoechst 33258 was added and co-incubated with sperm for 10 min. Samples were washed free of unbound stain and the sperm were mounted as smears on glass slides. After drying, sperms were incubated with FITC-PSA for 30 min. Sperm were examined by fluorescence microscopy. Also, FITC-Concanavalin A (FITC-ConA) staining and vital staining with yellowish eosin were performed simultaneously. The correlation of viability and \%AR were analyzed. RESULTS: Four different staining patterns were observed and clearly distinguished as follows: a) Viable acrosome-reacted sperm, b) Viable acrosome-intact sperm, c) dead acrosome-reacted sperm, d) dead acrosome-intact sperm. There was significant correlation between the results obtained by Hoechst 33258 and vital staining methods in viability of human sperm (r = 0.927, P < 0.001). There was significant correlation between the two methods (FITC-PSA and FITC-ConA) in \%AR of human sperm (r = 0.92, P < 0.001). CONCLUSION: Viability and acrosome status of a human sperm can be easily assessed simultaneously by Hoechst 33258/FITC-PSA staining method. This combination method is considered useful to evaluate sperm function.
This article was published in Arch Gynecol Obstet and referenced in Journal of Cytology & Histology

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