IVF Mail - Overnight IVF insemination concentration

Overnight IVF insemination concentration

By: Wendy,
05 May 2008


Question:
	I would like to ask "What is the maximum concentration of sperms added in the overnight IVF culture?" 0.1 million motile sperms or above?

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Michael L. Reed said on 19 May 2008

"A good question, and the answer in terms of sperm concentration per egg per ml really depends on the system you are using for insemination - well dishes vs test tubes vs micro or macrodrops. You will see a lot of references to sperm per egg per ml, designed originally for use with well dishes or test tubes. For example, 50,000 motile sperm per egg per ml was/is common with well dishes, each dish having one ml medium either under oil or with lots of medium around the well in a moat, and a single egg in the one ml volume. Similar for test tubes, where an egg is placed with one ml medium. The dynamics of the well, or tube are different as well, as wells may have a greater diameter than the test tube, changing the local concentration around the egg, and hence you may not need quite as many sperm per egg per ml in a narrow test tube. In micro or macrodrops, the concentration/egg/ml may be the same, but you will use a lot less sperm. For example, I target a final drop size of 200 microliters after the addition of sperm, so to get to 50,000 sperm/egg/ml, I'm adding approximately 10,000 sperm to the drop. I'll increase the number of sperm per drop if I add an extra egg (up to three eggs per drop), and the maximum number of sperm per drop that i use is approximatley 20,000 to 25,000 sperm. You can use fewer sperm in micro or macrodrops due to the fact that the drops trap the sperm in closer approximation to the egg(s). I believe that I've seen a concentration as low as 2,000 sperm per egg in a microdrop setting, but I would not personnaly want to go that low with sperm numbers.

If you are using micro or macrodrops, you should be even more careful of the pH management, as a lot of sperm in a confined volume will change the pH, as metabolic acids are released by the motile sperm, you'll see a shift towards a more acidic environment. This is very easy to test with medium that has phenol in it. You can inseminate drops, wells, tubes, whatever with different numbers of motile sperm, and watch what happens overnight.

Mike"

Simon Aiken said on 11 May 2008

"The amount to inseminate with varies, but its generally between 50,000-100,000 sperm per well (or ml). I have seen clinics that will go up to 150,000, but only if they're concerned about the "fair" motility of the sperm."