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jenny,
20 February 2012
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Dear All,
I need your advice on how to wash thawed sperm with very low count. Before freezing the count was less than 100 000/ml, freezing was done using sperm freezing medium with a ration 1:0.7 or 1:1 ratio after concentrating the sperm (doing simple wash to increase the count)
After thawing i cant do density gradient because of the low count. and if i do simple wash, how will i be sure that the glycerol or toxic ingredients of the freezing media was removed before doing icsi?? |
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Low sperm count wash after cryopreservation
Akanksha Mishra said on 05 March 2012
"Hi Jenny, you have to do simple wash in case of low sperm count. You have to ensure that you have to add the washing media over a period of time. Add two-three drops of media, shake the tube, wait for a minute, then repeat. this ensures the removal of the cryoprotectant from the sperm cells as you gradualy decrease the molar concentration of the medium."
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do simple wash
achmad muhyiddin said on 22 February 2012
"for the sperm with very low count, you only can do simple wash methode with three times centrifugation. you can use 2-5ml sperm preparation medium for rinse with 1000rpm for 5-10 minutes every centrifugation. that will reduce toxic ingredients of the freezing media."
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sperm washing
Andrew Glazier said on 20 February 2012
"jenny,
your best option is to spin down as is and then very carefully remove the supernatent. Resuspend the pellet in as small a volume as possible (certainly less than 0.5ml)and then plate sperm suspension in microdrops in the ICSI plate. The spin plus resuspension and subsequent plating out will be sufficient to dilute out the cryoprotectant. Good luck!
For future reference, you may have been better placed to concentrate the intial sample to have a better recovery, allowing a wash step. Performed correctly, you should only loose approx. 10% of the sperm per wash/gradient, leaving sufficient for ICSI"
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