I received a shipment for which INRA 96 was used as the extender. We were using the shipment for just two mares but apparently the sending facility was concerned about not giving us enough, so they sent 3-50 ml doses, processed as follows (according to the information on the included sheet):
Volume Raw Semen per insemination dose: 20 ml Colume of Extender per insemination dose: 30 ml Total volume of extended semen dose: 50 ml Progressive motility: 60% Total #PMS per insemination dose: 2.74 billion PMS/ml of extended semen: 54.9 million
The concentration of the raw semen was not listed but can be calculated backward from this information, I believe, as 228 million/ml and 137 million/ml PMS
The low extension rate and the overly-high concentration of the extended semen set off an alarm, but I was wondering how detrimental this was as I understand the INRA extender has some tolerances beyond "normal" extenders. Was this processing acceptable with that extender?
Your mathematical calculations would appear to be correct.
INRA '96does have some slightly different properties from other extenders in that it can be used at as low a dilution ratio as 1:1 and still have good success.
As the old saying runs "the proof of the pudding is in the eating"... what did the semen look like under the microscope once warmed? If you had an acceptable number of progressively motile sperm (>100 million) per insemination dose, then you are still probably just fine, no matter what the dilution ratio was!
Thanks for the reply, Jos. Well, I am hoping "the eating"--the actual preg checks--prove my concern is unwarranted.
I didn't look at the semen until it was 42 hours old, though the mares were bred when it was 26 hours old (the sample I looked at was kept in the Equitainer). It looked pretty bad by 42 hours, maybe 5% motile, probably more of those just twitching than actually PM. The slide was covered so densely it made me think of a battlefield, the few survivors feebly trying to move around their fallen comrades.
I looked at a sample that was appropriately warmed on a warm slide, then covered with a warm cover slip, and at one that went from the container to a cold slide that I watched over a period of time while it sat on the scope. Oddly, both "sets" looked about the same.
I was hoping that either I had some kind of seizure that prevented me from correctly reading the slide, or that the stuff was a lot better at 26 hours, or that the guys that were alive and kicking were "Superman." I guess, since there were so many cells in the dose, it could have dropped down to 2.2% and we would have had 100 million...
If the girls are in foal, no worries. But if not, I think I would ask the stuff to be extended differently next time. Would you expect that the conditions of extending would have rendered it in such poor shape, or would you suspect some other issues are affecting the quality? I mean, it was just 5 million higher than the upper end of the acceptable range, right?
Did you actually warm the semen on a slide warmer before evaluation? Simply putting it on a warm slide will be unlikely to get the job done.
A significant drop-off in quality between 26 and 42 hours would not be in the least unusual.
There's no real way to pinpoint a problem, as one doesn't even know if there was a problem (not having evaluated prior to 42 hours, nor having other extenders to compare the results to). The final concentration was higher than I would personally like to ship at, but with some stallions that is not a problem. With others it is...
I'd wait and see what your mares think before stressing more if I were you...
I really got a laugh out your post! It lightened my mood. So, did the girls like the Superman? Are they or aren't they?
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