INSTRUCTIONS FOR STORAGE, THAWING, and INSEMINATION
of FROZEN EQUINE SEMEN

PLEASE READ THESE INSTRUCTIONS FIRST. VERIFY THAT THE STRAWS YOU HAVE RECEIVED ARE THOSE YOU ORDERED, PRIOR TO THAWING, and CORRESPOND WITH THE POST-THAW INFORMATION ON THE ATTACHED CARD, AFTER THAWING!!

If sperm motility upon thawing does not approximate that stated for this batch on the accompanying card, or if you have any other questions, please call immediately, 24 hours per day. Our office number is 530-737-3637. DO NOT THAW MORE THAN ONE STRAW PRIOR TO CALLING US IF SOMETHING DOES NOT SEEM RIGHT!! Our consultation is free of charge, and the live-foal guarantee provision of the breeding contract is null and void if these instructions are not followed!

UPON ARRIVAL:

If you have a separate storage tank, inspect and transfer the straws from the shipper to your storage tank as soon as possible. Frozen cotton is packed inside the shipper canister to prevent the straws from rattling and possibly cracking during shipment. This should be withdrawn using cold tweezers BEFORE attempting to remove the straws. We suggest that a styrofoam tray of liquid nitrogen be used to precool your straw tweezers, and that withdrawn straws be placed in this tray for verification of identity and integrity (check for cracks) prior to transfer to your storage tank. Larger 5ml straws should have the air bubble in the MIDDLE of the straw. If the air bubble is at the straw END, then the straw has been fully thawed since it left our hands, and both the carrier and ourselves must be notified IMMEDIATELY.

If you have no separate storage tank, the shipper itself will work well for storage, provided that you top it off with additional nitrogen. Please be aware that your client is charged an additional $10. per day rental fee for tanks not returned the next business day using our enclosed airbill.

TANK RETURN:

Please follow the return instructions sent with the tank. You only need to weigh, sign, and date the pre-prepared airbill, and contact FedEx for pickup. Any unused straws MUST BE RETURNED. In this case, please recharge the tank with liquid nitrogen, then empty all excess just prior to shipping.. Top up the tank with liqNote that you only need to weigh, sign, and date the airbill for return.

QUICKIE INSTRUCTIONS:

Temperature fluctuations from 37 degrees C, exposure to sunlight, and any delays in insemination once the semen has been thawed are highly detrimental events to be avoided at all costs!

  1. Single, periovulatory inseminations may be made, provided that the mare's follicle is being scanned at least every six hours, round the clock. For inseminations based upon palpation alone, or when follicle evaluation will be less frequent than every six hours, we recommend "waiting as close as you dare" to ovulation, and then AI every 12 hours, including the first postovulatory check. Note that some stallions only provide single doses per shipment.
  2. Straws should not be handled or thawed until all supplies are prewarmed, the mare is wrapped and washed, and the thaw-bath is up to temperature.
  3. Using PRECOOLED straw tweezers (loaned free of charge upon request), quickly transfer one straw from liquid nitrogen to the thawing bath, grasping the straw firmly and directly over the end sealing bead. Grasping the straw elsewhere will increase the internal pressure, and increase the possibility of straw cracking or sealing-bead expulsion. The thawing protocol for THIS shipment is one of the following (will be circled upon shipment):

    a.) 15 seconds at 37 degrees C

    b.) 60 seconds at 37 degrees C

    c.) 45 seconds at 50 degrees C

    d.) 7.0 seconds at 75 degrees C

    All temperatures are +/- 0.25 degrees C. Use a DIGITAL or GLASS-ROD LABORATORY THERMOMETER ONLY. Other types are not accurate enough!

  4. Wipe the thawed straw dry with a tissue before cutting one end open with a sterile razor blade or sharp pair of scissors. For 0.5ml straws, load the opened straw, open-end first, into the sterile AI gun. The blue, plastic cylinder is used to firmly seat the straw inside the gun; the metal stylet expels semen once inside the mare. For MULTIPLE 0.5ml doses, empty all straws into a pre-warmed, sterile red-top tube and aspirate the total volume into a pre-warmed AI pipette using a sterile, pre-warmed, all-plastic syringe.  Alternatively, cap the syringe tip with a sterile cap, withdraw the plunger, and empty the semen into the syringe barrel.  Replace the plunger and immediately AI through a pre-warmed AI pipette, using air to slowly and thoroughly flush semen out of the pipette, while at the same time intoducing as little air into the uterus as possible. For large straws, the contents can be emptied into a PRE-WARMED sterile, red-top tube; then aspirated into a PRE-WARMED AI pipette using a sterile, all-plastic syringe (also PRE-WARMED). Alternatively, cap the syringe with the sterile cap provided, PRE-WARM it, withdraw the plunger, and empty the semen into the syringe barrel. Replace the plunger, and inseminate through a PRE-WARMED AI pipette, being careful to avoid introduction of air into the uterus, while at the same time using air to completely flush out the pipette.
  5. The mare should be inseminated as quickly as possible after thawing the semen, depositing semen SLOWLY into the uterine body. We then recommend clenching the fist, and massaging the external cervical os and anterior vaginal vault for one full minute post-insemination, to help stimulate uterine contractions which propel the sperm towards the oviducts.
  6. Now check motility of the thawed semen using a drop that invariably remains in larger straws, or in the AI pipette. Be sure to use a slide warmer and warm slide! Any desired post-breeding infusions should be done at least 90 minutes post-insemination.

GENERAL INFORMATION:

The straw-thawing process should take place in a heated room, yet as close as possible to the waiting and pre-prepared mare. We strongly advise against travelling ANY distance through sunlight or inclement weather to perform inseminations. Make other arrangements instead. Ideally, semen will be in the mare within one minute of thawing. All handling of straws pre-thawing should be done using PRECOOLED straw tweezers and beneath the surface of liquid nitrogen. Straws should be removed from the nitrogen ONLY for thawing. IT IS NOT ADVISABLE WITH OUR STRAWS TO MIX OR ADD ANY OTHER EXTENDERS OR ANTIBIOTICS TO THE SEMEN!! "Volume" per se is NOT important, as we have had excellent pregnancy rates with single, half-ml straws. We believe the massage to stimulate uterine contractions is of far greater importance. We do not recommend breeding with frozen semen very early in the season, when follicle development is slow and erratic, or on any other estrus cycle where follicular development is not progressing steadily and rapidly. Postovulatory breeding has excellent success rates PROVIDED one is certain that insemination is succeeding ovulation by LESS THAN SIX HOURS.

The shipper tank is sent as a dry shipper which will keep straws in an unopened tank cold for just a couple of days. If storing the straws in the shipper tank, NITROGEN SHOULD BE ADDED UPON ARRIVAL, thenWEEKLY. Keep the tank in a cool, draft-free location, in its plastic overpack, and out of the reach of inquisitive visitors. The lid LIFTS STRAIGHT UP, AND HAS AN INSULATING STYROFOAM CORE ATTACHED. Never raise the canister out of the tank to withdraw straws; instead use pre-chilled tweezers to reach down into the neck to grasp them. Good luck!

QUESTIONS?
Dr. Mennick, office telephone 530-737-3637 seven days