Semen Assessments
The acrosome is a membrane-bound organelle located over the anterior half of the sperm head. The acrosome contains degradative enzymes, such as hyaluronidase and acrosin, functioning in breaking down the zona pellucida of the oocyte, thereby facilitating fertilisation. Acrosome integrity is associated with increased fertilising potential and fertility [1, 2]. Damage to the sperm acrosome, or premature acrosome reaction renders the spermatozoon incapable of fertilisation.
Acrosome integrity is assessed using flow cytometry, which allows us to objectively assess 10,000 sperm per sample, as opposed to only a few hundred via microscopy. Spermatozoa are incubated with a vitality stain (Live/Dead™; LD), before fixing in paraformaldehyde. Cells are then briefly permeabilised to allow the fluorescent probe, FITC-PNA (fluorescein isothiocyanate-conjugated peanut agglutinin), entry into the cell. FITC-PNA binds specifically to the outer acrosome membrane [3]. Where the acrosome is intact, spermatozoa will fluoresce green. The vitality counterstain (red) enables us to distinguish between ‘true’ acrosome loss (live cells with no FITC-PNA fluorescence – occurring either through premature acrosome reaction or through damage) and necrotic acrosome loss which has occurred following a loss of vitality (dead cells with no FITC-PNA fluorescence). The populations reported are:
1. Kumaresan, A., Johannisson, A., Al-Essawe, E.M., and Morrell, J.M. (2017) Sperm viability, reactive oxygen species, and DNA fragmentation index combined can discriminate between above- and below-average fertility bulls. Journal of Dairy Science. 100(7): p. 5824-5836.
2. Harayama, H., Nishijima, K., Murase, T., Sakase, M., and Fukushima, M. (2010) Relationship of protein tyrosine phosphorylation state with tolerance to frozen storage and the potential to undergo cyclic AMP-dependent hyperactivation in the spermatozoa of Japanese Black bulls. Molecular Reproduction and Development. 77(10): p. 910-921.
3. Mortimer, D., Curtis, E.F., and Miller, R.G. (1987) Specific labelling by peanut agglutinin of the outer acrosomal membrane of the human spermatozoon. J Reprod Fertil. 81(1): p. 127-35.