Sperm Preparation

1. Swimup
2. Percoll

Percoll Separation Procedure

Next you will fractionate the Percoll column so that you can observe how sperm are separated.

• Attach a syringe to a pasteur pipet.
• Plunge the pipet to the bottom of the Percoll tube. From the bottom fraction just remove the pellet of sperm by aspirating with the syringe. This might only be 100 to 200 µl so be careful. Place the first aliquote in a small 1.5 ml centrifuge tube.
• Repeat the previous step but remove 0.5 ml aliquotes from the bottom, placing each in a separate 12 X 55 mm test tube. Once you reach the extender, you can pool the extender fractions in one tube.
• Evaluate the motility of each fraction and fill in the accompanying table at the bottom. Do this as quickly as possible as you have 5 - 10 minutes at most. 

While motility is being evaluated, other members of the lab group will need to go to the next page and determine the concentration of sperm in the sperm pellet (bottom fraction).