I will be very grateful if somebody can help me. This is the situation: I can get fertilization and cell division, but the embryos presents fragmentatation. I obtain grade II or III embryo quality and occasionaly grade I. I would like to know which can be the posible reason and solution for that.
Some details: -
I am using medicult médium :
*flushing for aspiration and transfer
*IVF for fertilization and cleavage -Culturing on well dishes for fertilization with 100.000 esperm and tranfering to 50 ul drops over oil after pronucleus identification (day 1)
-Then (day 2) we evaluate the cleavage and grade the embryos. -Finally we transfer on day 3
-The incubator is fitted at 5% CO2 and 37 C
- I am using for recovery, pre-washed and sterile pasteur pipettes,for embryo handling on day 1 and 2 stripper tips 200 nm and 150 nm
- we don´t have termoplate for microscope, but we work as fast as possible.
- we use dry bath at 39 C on aspiration set and near the microscope
- We aspirate with 20 ml syringe Plese ask me for more details Best Regard |
alok teotia said on 29 May 2008
"possibly there is some toxicity in the envionment in which the oocyte/embryos are grown. it could come via solids (culture ware), liquids (media) and gas, check them all
all the best"
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Mohammed Ibrahim said on 12 May 2008
"All what is mentioned here is great.
No one can really tell you the reall caose of the situation . it is only you who can know. So, you have much work to do.
I assure that QC has great effect here. pH variations can have deleterious effect.
Since you have good fert rate and you are transferring the zygotes on day one to new culture dishes, try to be quick in this step, Temp varaitions also have negative effect on the spindle.
What is the Biochemical PR? Abortion R?
Check step wise and exclude one factor each time.
Oocyte quality might also have a role here. type of drugs, urinary or recom?
You have a lot to work on. Good luck.
in my opinion, the usuall rate of G1 embryos is 35-40 % and the same for G2."
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Orestes Prudencio said on 10 May 2008
"PH is the magic kye on embryo quality.
strive to reache a PH between 7.22 and 7.25 and your problem will be solved, as much there is no medium contamination.
Hope this help"
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Lakshmi Sharma said on 06 May 2008
"Try culturing your embryos in microdrps with 3-5 embryos per drop under oil. Also to eliminate variation in pH and control VOCs culture a dessicator filled with a medical grade trigas mixture of 6% co2,5%o2 and balence nitrogen. pH chnges and increased VOC levels are usually responsible for fragmentation in the lab. I would next examine the stimulation protocols and do a comparitive study to see if fragmentions is occuring across the board or is seen in cases with high e2 levels before HCG.Hope you find some answers.
Lakshmi"
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Kimball O Pomeroy said on 05 May 2008
"Large amounts of fragmentation are usually due to contaminants, especially endotoxin. I would check to make sure the medium never gets above 37 C. I would never set a dry block to 39 to get it to rad near 37 as at the very bottom where the egg sits, it might be 40 C. Next, I would make sure that you have done appropriate QC bioassays for all contact materials. I would suspect syringes, dishes, plates, etc."
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Dr.Ravindranath said on 05 May 2008
"You have to check your sperm preparation. Do a second swim up and get the clean motile sperms. It seems u r doing only IVF and not ICSI. If that is the case, also check the water in the humidity chamber of ur CO2 incubator. You must change the water every 10 days and the level must be maintained or else fragmentation comes. You should also know the pathology of the patient and if she is an untreated PCOS or Endometriosis, u r bound to get fertilization but embryos with fragmentation. Why don't you divide the oocytes, half by IVF and half by ICSI. You yourself can find the reason."
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DPankar Banerji said on 04 May 2008
"Please check the pH of Media after equilibaration ,inside the incubator. We use Eutech pH meter.
Bye"
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