In Vitro Fertilization using Epididymal Sperm Transported at Cold Temperature

Materials and Equipment

1. Cauda epididymis transported at cold temperature
2. FERTIUP® (Preincubation medium: PM; COSMO BIO)
3. mHTF
4. Micropipettes
5. Plastic dishes (35mm x 10mm Cat.No.430588; CORNING)
6. Fine scissors
7. Pair of watchmaker's #5 forceps
8. Filter paper
9. Humidified incubator (37℃, 5% CO2, 95% air)

Procedure

1. Put 1 drop (100μL / drop) of FERTIUP®(PM) into a dish and cover it with liquid paraffin 30 minutes before collection of epididymal sperm transported at cold temperature, and place the dish in an incubator (37℃, 5% CO2 in air).
   
   
   
   
   
2. Remove the 0.2mL tube containing the sample from the foam transport box.
   
   
   [Removing the sample]
   
   
   
3. Open the tube, pick up the cauda epididymis and wipe away any preservation solution using filter paper.
   
   
   
   
   
   
4. Wash the cauda epididymis in each of the three drops of mHTF in a washing dish. After washing, wipe away any excess mHTF using filter paper.
   

   Comment:

   To make a washing dish, put 3 drops (about 100μL / drop) of mHTF into a dish without liquid paraffin just before use.

   
   
   
   
   
   
5. Place the cauda epididymis in the sperm dish containing liquid paraffin.
   Epididymal spermatozoa transported at cold temperature can be utilized for in vitro fertilization in the same manner as fresh spermatozoa.

   Note:

   If you find it difficult to release sperm from the cauda epididymis, make one more incision in cauda epididymis to release more sperm.

   Note:

   There are three different methods of preparing CARD MEDIUM, depending on whether in vitro fertilization will be carried out using fresh, frozen-thawed or cold-temperature transported spermatozoa. Please refer to the CARD MEDIUM instruction manual.

Update history

1. Updated : 20 April, 2012