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RESEARCH DIVISION Laboratory Manual

 


 

Vasectomy of Male Mice

Mice are vasectomized to be mated to females used for embryo transfer. The female becomes pseudopregnant, her body geared for a pregnancy because she has been mated.

Vasectomy is performed on mice at 6-8 weeks of age. We use the hybrid (C57BL/6J x CBA/CaH)F1 as these males have a higher libido than most other strains.

Before starting:

  Sterilize-
- -3 pairs of curved forceps
- -2 pair of iris scissors
- -1 pair suture holders
- -1 pair Michel clip applicators
- -Michel clips
- -Suture
  Make up Rompun/Ketavet anaesthetic
- Add 0.5 ml 2% Rompun (20mg/ml Xylazine) and 0.5ml Ketavet (100mg/ml Ketavet) to 9.0 ml of PBS (Phosphate Buffered Saline)
- DOSE (0.02 ml/g bodyweight, given i/p ):
- <25g bodyweight give 0.4ml
- >25g bodyweight give 0.45ml

We do not use mice which weigh less than 20grams.

  Procedure

The mouse is weighed and anaesthetized accordingly. Lie the mouse on its back and shave lower abdomen with clippers. Swab clipped region with 70% EtOH. Using a pair of curved forceps and iris scissors,(the outside instruments) cut across the lower abdomen, making the incision about one cm wide. Using separate forceps and scissors (the inside instruments) to those that the skin was cut with, incise the body wall, reach in and locate the bladder. Both tubes of the vas defrens will be visible lying on either side. Grasp the left vas deferens gently with the other (third) pair of forceps and lift a section clear of the incision. Using the suture, make two firm knots in the vas deferens, about 4-5 mm apart, tying both knots firmly. Cut out a section of vas deferens from in between the knots and place on a tissue to be sure that one side has been done. Gently ease the two cut ends of the vas deferens back inside the abdominal cavity and repeat procedure on the right hand side. When both sides have been done, sew up the incision in the body wall with separate stitches - two or three should suffice. The skin is closed up using two Michel clips to hold the wound closed. The mouse must be wrapped in a tissue to keep it warm (loss of body heat is common in abdominal surgery) and placed in a box by itself to recover. Animals which are placed under anaesthetic should always be supervised and monitored until fully awake.

Following the operation, the mice are allowed to recover for two weeks before being test bred to confirm their sterility.

  Test Breeding

One or two female mice are placed with the vasectomized male and are checked for plugs the following morning. Females which plug are flushed (see page 89) 24 hours later with PBS. The eggs should be at one cell stage or unfertilized. If they are at two cell stage then the vasectomy was not performed correctly and the male should be culled.

Disclaimer: The following set of protocols were contributed by various members of our lab (past and present): Christine Andrews, Fiona Christensen, Neil Della, Ross Dickins, Debbie Donald, Andrew Holloway, Gary Hime, Colin House, Yinling Hu, Rachael Parkinson, Nadia Traficante, Hannah Robertson, Ping Fu and Dennis Wang. Special thanks to Vicki Hammond, Frank Kontgen and Maria Murphy, who contributed many of the ES cell protocols. Sections dealing with Photomicroscopy, Polyclonal and Monoclonal Antibody Production were provided by members of Gerry Rubin's Laboratory (Berkeley). Any comments in the methods (technical errors etc.) E-mail: d.bowtell@pmci.unimelb.edu.au
David Bowtell PMCI October 1998