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Freezing sperm is an efficient, cost-effective way to conserve and distribute genetics in the agricultural industry and putting male sex cells on ice is a fundamental part of human fertility programs. But the sperm of certain varieties of mice under-achieve woefully after being frozen and thawed. What's worse: the thawed sperm of the most popular mouse strain in the scientific world, the C57BL/6 or "Black 6", are known to under-perform when it comes to fertilising mouse eggs.
Scientist Live spoke with Dr's Michael V. Wiles, Charles Ostermeier, Rob Taft of The Jackson Laboratory about their discovery.
1. What is frozen mouse sperm generally used for in laboratories? Why is it important?
There are many thousands of mouse strains in the world and more are being made daily as models to help us understand and improve the human condition. The cost of making these animals is very high; the research done with them long and painstaking. Once this resource has been made and (some) data acquired, there is a need to maintain these novel mice strains so that they can be provided to other scientists as new ideas develop allowing these strains to be used again and the previous data built on. To keep these strains as live mice "on the shelf" is not viable if they are not being actively worked on; i.e. resources are limited. Not archiving them would be a massive intellectual and financial resource loss to the worldwide scientific community. Hence the need to develop a viable, cost effective system for managing mice was major goal for us.
2. What are the problems researchers currently encounter when using frozen mouse sperm?
We should point out that both mouse sperm and early embryos could be frozen successfully before we developed this approach. However, embryo cryopreservation is relatively expensive, and provides only limited numbers of animals when strains needs to be recovered. Previous sperm cryopreservation methods worked for many inbred strains. However, as fate would have it, or more accurately perhaps genetics, the main workhouse strains of the scientific community, C57BL/6J's sperm simply failed to fertilize oocytes efficiently post thaw. It was this strain which we concentrated our efforts on.
3. How has your lab circumvented these issues?
My laboratory and Dr Rob Taft's worked on this together. We approached the problem perhaps more holistically than others, combining old ideas linked with how to maintain cell functionally after the cryopreservation process with simple keen observation. As Louis Pasteur said "Chance favours the prepared mind".
4. How easy is it for a lab to implement this new system?
The strength of our work in The Technology Evaluation and Development Department is that we work very closely with the team which will use any technology we develop. In this case, Dr. Rob Taft's Reproductive Sciences group, which is an ARTs (Assisted Reproductive Technology) service lab for TJL and the scientific community, were intimately involved. So the implementation from the research lab to full production lab was simple, fast and much appreciated by all involved (as it simply worked!).
Other labs will be able use our methods as they are published and we are also teach courses on how to do this. In fact, we are developing a kit which can be sent out to lab to freeze mouse sperm. We then offer to check the quality of the sperm and store it here.
So freezing mouse sperm is simple. Having said that however, IVF (in vitro fertilization) is required for both quality control and to recover strains and it is hard to establish in a general mouse lab.
5. How is it more efficient?
The best way the answer this is to simply say before we had this method worked out very few labs would freeze sperm from C57BL/6 (or almost any other strain). Now the majority of strains can be cryopreserved as sperm and more importantly, recovered cost-effectively.
On the numbers/efficiencies, with earlier experiments done by us and many others - you were lucky to get 5-15% of the oocytes fertilizing post thaw of C57BL/6 sperm - now we get 50-70%.
(There is still variation, in part due to the large number of variables, including for example that some genetic modifications may affect sperm fertility function in ARTs.)
6. Does this efficiency translate to lower overhead?
It lowers the cost to both freeze a strain (more than 50% cheaper) and, I think much more importantly, it gives many more options on a strains recovery when it is needed; i.e. with embryo cryopreservation you tend to freeze only a few hundred embryos (because of cost). Since only about 50% of embryos make it to live born animals, you anticipate recovering tens to maybe a few hundred animals at most. With sperm, you freeze millions of gametes. Hence, if you choose to, you can recover many more animals (in fact thousands depending upon the strains background) much more rapidly.
Also by taking a mouse strain "off the shelf" you are saving animal facilities $5k to $10k/strains/year. Further you are not wasting animals which are bred and then not used productively.
Lastly, by putting strains into cryopreservation as a general measure, when (not if) disaster sticks these costly resources are not lost and can be rapidly resurrected and the work continued.