Fluorescent labelling of boar spermatozoa for quantitative studies on competitive sperm–oviduct binding
Heiko H. W. Henning
A B , Julia Batz-Schott A , Benita Grünther A , Xuyen Le Thi A and Dagmar Waberski A C
+ Author Affiliations
- Author Affiliations
A Unit for Reproductive Medicine, Clinic for Pigs and Small Ruminants, University of Veterinary Medicine, Bünteweg 15, 30559 Hannover, Germany.
B Department of Equine Sciences, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 112, 3584 CM Utrecht, Netherlands.
C Corresponding author. Email: dagmar.waberski@tiho-hannover.de
Reproduction, Fertility and Development 31(9) 1520-1532 https://doi.org/10.1071/RD19081
Submitted: 3 July 2018 Accepted: 8 April 2019 Published: 10 May 2019
Abstract
In vitro sperm–oviduct binding assays enable assessment of the capacity of spermatozoa to form a ‘reservoir’ in the oviduct. Competitive approaches, such as experimental set-ups that test multiple males or semen samples simultaneously on the same tissue explants, are desirable because they reduce the likelihood of bias when using material from different females. Therefore, we established a fluorescent labelling technique that allows tagging and storage of spermatozoa before competitive studies of sperm–oviduct binding in vitro. Fluorescent markers were tested for reliability and compatibility with parameters of boar spermatozoa viability. The addition of seminal plasma after density gradient centrifugation was essential to counteract centrifugation stress during the labelling procedure. It was demonstrated that sperm tagged with MitoTracker Green FM or MitoTracker Red FM can be successfully used in competitive sperm–oviduct binding studies. The assay was sensitive enough to indicate subtle effects of semen storage temperature on the ability of the spermatozoa to contribute to the female sperm reservoir.
Additional keywords: sperm competition, sperm reservoir.
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