Retrieval of Mouse Oocytes

Summary

This protocol illustrates the technique for extracting oocytes or early-stage fertilized embryos from the oviduct of mice. The ability to identify the infindibulum and insert a blunt end needle into it is essential to correctly performing the procedure.

Abstract

To date, only a few studies have reported successful manipulations of Peromyscus embryogenesis or reproductive biology. Together with the Peromyscus Genetic Stock Center (http://stkctr.biol.sc.edu), we are characterizing the salient differences needed to develop this system. A primary goal has been to optimize oocyte/early embryo retrieval.

Protocol

Embryo/Oocyte retrieval

1. At least two hours prior to retrieval of embryos/oocytes, KSOM is placed in the 37ºC CO₂ incubator and FHM must be thawed from -20°. A glass dish containing three concave shallow wells is used to house drops of KSOM in the incubator. To prevent dehydration, the KSOM and dish are placed in a lidded container that has water at the bottom. This apparatus prevents evaporation without the use of mineral oil.
   
   
2. The oviducts from an ovulating female are placed in FHM media. To ensure orientation and an intact infindibulum, a small section of the ovary is cut along with the oviduct and small section of ....

Discussion

In this article, we demonstrate the retrieval of oocytes from ovulation-induced (superovulated) Peromyscus. The use of superovulation (as opposed to natural estrous cycle) is common due to the much greater resulting numbers. However, caution must be taken in interpreting results utilizing such oocytes/embryos, as both the hormones used to induce ovulation and culturing may have effects.

When estrous has been confirmed, females are euthenized and oviducts recovered along with a small porti.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Flushing needle	Tool	Zephyrtronics	ZT5-130-L	30 G blunt
Forceps (2)	Tool	Roboz Surgical Instruments Co.	RS-5060
Peromyscus	Animal	Peromyscus Genetic Stock Center
Scissors	Tool	Roboz Surgical Instruments Co.	RS-5880
FHM HEPES Buffered Media	Reagent	Specialty Media	MR-025-d
KSOM Embryo Culture Medium (5x 10 mL)	Reagent	Specialty Media	MR-020P-5F
Mouth Pipette	Tool			Pulled from glass capillaries and attached to rubber tubing
Oocyte holding dish	Tool			Glass container with concave divets