Canine Artificial Insemination Extender
A universal practice in artificial insemination (AI) procedures, a semen extender is added to an animal’s semen to preserve its fertilizing capabilities. Initiated within animal husbandry, AI has spread from cows and pigs to a wide variety of animal species, including endangered animals in zoos for wildlife conservation and domestic dog breeding.
Artificial insemination has been embraced by dog breeders maintaining pedigrees who are restricted from traditional mating for various reasons. There has been much study regarding canine semen extenders, with formulas modified from successful bovine semen extenders. A typical formula is one shown below:
| Ingredient | Purpose |
| Double distilled water | Solvent |
| Tris(hydroxymethyl) aminomethane | pH buffer |
| Citric acid | pH buffer |
| Fructose | Energy source |
| Penicillin | Antibiotic |
| Glycerol | Cryoprotectant (freezing semen only) |
| Egg yolk | Protection from cold shock |
The Tris, citric acid, and fructose are stirred in water until dissolved, then sterilized. For fresh chilled semen to be stored at 4°C, an antibiotic and fresh egg yolk (20% v/v) are added and mixed well, followed by mixing into the semen to make a known concentration. Versions of this formula successfully maintain semen’s viability for 2-48 hours after collection, allowing for cold-shipments nationally or globally to dog breeders.
The yolk’s fat globules must be broken down into small particles, achieved by vigorous mixing.
Preparation of egg yolk is important for preservation of the semen, and must be as free of albumin and the yolk membrane as possible. As well, the yolk’s fat globules must be broken down into small particles, achieved by vigorous mixing. The prepared yolk is added to pre-warmed Tris buffer at 30°C, then stirred to homogeneity while being warmed to 35°C. The completed extender is now ready to be gently mixed into the collected semen.
For frozen long term storage, antibiotics and the glycerol are mixed with the Tris buffer first, with egg yolk added just prior to use. After adding extender to the semen similar as for cold storage, the extended semen is cooled, transferred to cryovials and frozen using liquid nitrogen.
Because of the small volumes being used and the variety of mixing tasks involved, Caframo’s Petite stirrer 250 is ideal for preparing canine AI extender media. The 50‑2500 rpm speed range encompasses both the vigorous mixing needed to prepare the egg yolk, and the gentle homogenization of semen as it is diluted in the extender. Consistency and reproducibility in the process is achieved with the digital display and push button settings. The impellers provide both axial and radial mixing options, and the small diameter allows direct mixing in vessels without needing to transfer. The 250 package includes everything you need to begin immediate mixing, is easily portable, and occupies a small footprint on the lab bench. Contact us to discuss how the Petite can meet your small volume mixing needs.
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Bateman HL (2001) Effects of Semen Extender Composition and Cooling Methods on Canine Sperm Function and Cryo-Survival. (Master’s Thesis). University of Guelph, Guelph Ontario. Retrieved from http://www.collectionscanada.gc.ca/obj/s4/f2/dsk3/ftp04/MQ61869.pdf
Dresser, BL (1988) Cryobiology, Embryo Transfer, and Artificial Insemination in Ex Situ Animal Conservation Programs. In Wilson EO and Peter FM (Eds.) Biodiversity (296-308). Washington, D.C: National Academy of Sciences.
Farstad, WK (2010) Artificial Insemination in Dogs. In England G and von Heimendahl A (Eds.), BSAVA Manual of Canine and Feline Reproduction and Neonatology 2nd ed. (80-88). Gloucester, UK: British Small Animal Veterinary Association.
Hori T, Yoshikuni R, Kobayashi M, and Kawakami E. (2014) Effects of Storage Temperature and Semen Extender on Stored Canine Semen. Journal of Veterinary Medical Science 76: 259-263. doi: 10.1292/jvms.13-0303
Howard JG, Lynch C, Santymire RM, Marinari PE, and Wildt DE. (2015) Recovery of gene diversity using long-term cryopreserved spermatozoa and artificial insemination in the endangered black-footed ferret. Animal Conservation 19: 102–111. doi:10.1111/acv.12229
Queiro A (2015, Feb 3) Why are dog breeders turning to AI? Retrieved from http://www.bbc.com/news/uk-scotland-31091695.
Umut, CA (2015, Aug 30). Are there any ways to filter egg yolk for semen extender?. Message posted to https://www.researchgate.net/post/Are_there_any_ways_to_filter_egg_yolk_for_semen_extender.
wiseGEEK (n.d.) What is a Semen Extender? Retrieved Oct 2016, from http://www.wisegeek.com/what-is-a-semen-extender.htm
