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mR1ECM (in vitro culture of rat fertilized eggs)


Superovulation induction and mating

Mature female Wistar rats (8-12 weeks old) received intraperitoneal administration of PMSG (150 IU/kg/rat) and hCG (75 IU/kg/rat) at 48-hour intervals to induce superovulation. After hCG administration, they were mated with male rats. On the next day, vaginal plugs were confirmed to ensure mating. Mated female rats were used in the experiment. Pronuclear-stage fertilized eggs can be collected on the day of vaginal plug confirmation, and 2-cell-stage embryos on the next day.

Recommended schedule

PMSG:@11:00 -13:00
hCG F 11:00 -13:00 (48 hours after injecting PMSG)

Collection time
Pronuclear-stage fertilized eggs :15:00-16:00 (on the day of vaginal plug confirmation)
2-cell-stage embryos                :15:00-16:00 (on the next day of vaginal plug confirmation)

Preparation of drops

Place three drops of mR1ECM (100 μL each) on a dish. Cover the dish with liquid paraffin and allow it to stand for at least 30 minutes in a 5% CO2 incubator before use for gas equilibrium.

Collection of fertilized eggs (fallopian tube perfusion)

Disinfect all dissectors with alcohol.
Heat culture medium for perfusion (M2 or PB1) to 37oC.

(1) Euthanize a mature female rat. Remove the uterus, ovary, and some fat using scissors-tweezers. Excise only the fallopian tube on filter paper, and remove the blood.

(2) Perfuse the fimbriae of uterine tube with culture medium using a glass capillary or perfusion needle to collect fertilized eggs.
* If you collect Pronuclear-stage fertilized eggs, please add hyaluronidase (final concentration: 0.1%) to the culture medium for perfusion (M2 or PB1) because early embryo is wrapped in cumulus cell. 

(3) Put the fertilized eggs into the mR1ECM drops.
The resulting two-cell stage embryos can be cultured in vitro to the blastocyst stage. For culturing, use mR1ECM (400 μL drop).

    mR1ECM Product Page 

To be used for research only. DO NOT use for human gene therapy or clinical diagnosis.

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