Porcine Cumulus Oocyte Complex Isolation: A Technique to Isolate Cumulus Oocyte Complex from Porcine Ovarian Follicle for In Vitro Fertilization

Abstract

Source: Gorczyca, G. et al., Proteolytically Degraded Alginate Hydrogels and Hydrophobic Microbioreactors for Porcine Oocyte Encapsulation. J. Vis. Exp. (2020).

This video describes the technique to isolate the cumulus-oocyte complex or COC from porcine ovaries. The isolated intact COC can be used to carry out the in vitro fertilization - the assisted reproductive technique for developing fertility treatments.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Isolation of porcine cumulus-oocyte complexes

1. To collect material for the isolation of ovarian follicles, excise porcine ovaries from prepubertal gilts (approximately 6-7 months of age, weighing 70 to 80 kg) at a local slaughterhouse. Choose approximately 20 pig ovaries from 10 animals for cumulus-oocyte complexes (COCs) isolation in each experiment.
   NOTE: Assuming each ovary yields 3–5 follicles, the total number of follicles varies from 60 to 100.
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Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
General
Antibiotic Antimycotic (100x) 100ml	Thermo Fisher	15240062	2.5% final concentration for Handling Medium. 1% in PBS (step 1.2)
DMEM/F12 (500ml)	Sigma-Aldrich	D8062	Handling Medium
FCS (100 ml)	Thermo Fisher	16140063	10% final concentration for Handling Medium. (step: 1.5)
PBS (1x, pH 7.4) 500ml	Thermo Fisher	10010023
Maturation Medium
hCG (1 VIAL of 10 000 U)	Sigma-Aldrich	CG10
PMSG	BioVendor	RP1782721000
Specific instruments, tools
30 mm Pteri dish	TPP	93040
60 mm IVF Petri dish	Falcon	353653
Incubator	Panasonic	MCO-170AIC-PE	Other brands can be used
Sterile petri dish (10 cm)	NEST Biotechnology	704002
Thermos	Quechua	5602589	Other brands can be used