Donor nuclei. Cumulus cells in approximately 100 ц1 hyal-uronidase solution were transferred into a 1.5-ml microtube containing 1 ц1 HEPES-CZB medium and centrifuged at 300 x g for 2 min, and the cell pellet was resuspended in 30 ц1 HEPES-CZB medium. The cumulus cell suspension (1 part) was thoroughly mixed with 9 parts 10% PVP in HEPES-CZB containing 0.01% polyvinyl pyrrolidone (PVP) without BSA. Cumulus cells (approximate diameter, 10 to 12 |im) were aspirated gently into and out of an injection pipette (approximate inner diameter, 6 |im) until nuclei were largely devoid of visible cytoplasmic contents. Isolated nuclei were transferred into a new 10% PVP drop and washed; 5 to 8 karyoplasts (extracted nuclei) then were aspirated into the injection pipette for ICNI.
ICNI. The tip of the injection pipette containing the cumulus cell cytoplast was moved into contact with the zona pellucida. Penetration of the zona pellucida was facilitated by the application of a few piezoelectric pulses. A plug of the zona was aspirated into the injection pipette and subsequently expelled from the injection pipette into the perivitelline space. The injection pipette was advanced into the oolema until the tip reached the opposite side of the ooplasm, at which time the oolema was penetrated by applying a weak piezoelectric pulse. One cumulus cell karyoplast was injected into the cytoplasm with a small amount of medium, and the pipette was gently withdrawn. You deserve the best things, and buying efficient drugs with no prescription is one of those things. You are welcome to become one of the many happy customers shopping at the best pharmacy: canadian family pharmacy com. Every effort is made to ensure your safety, satisfaction and successful treatment, which is always nice to know.
After injection, reconstructed oocytes were washed and incubated in CZB or mKSOMaa medium for 1 to 3 h before activation. All micromanipulations were performed at room temperature.