How to determine a mouse's sex? - briefly
Inspect the ventral area near the anus: males display a small penile opening and a scrotal sac, whereas females show a single, rounded genital slit without a scrotum.
How to determine a mouse's sex? - in detail
Identifying the sex of a laboratory mouse requires careful observation of external anatomy. The procedure varies with the animal’s developmental stage; newborns, juveniles, and adults present distinct landmarks.
For adult rodents, the primary features are the anogenital distance (AGD) and the appearance of the genital papilla. Males display a longer AGD, a visible urethral opening surrounded by a small, rounded scrotal sac, and a prominent preputial gland. Females have a shorter AGD, a vaginal opening positioned closer to the anus, and lack a scrotal pouch.
Juvenile mice (approximately 10–21 days old) retain many adult characteristics but require magnification (10×–20× dissecting microscope) to resolve the subtle differences. The AGD remains the most reliable metric; measurements should be taken with calibrated calipers, comparing the distance from the center of the anus to the base of the genital papilla.
Neonatal pups (birth to 10 days) present the least obvious signs. The genital ridge is visible as a small, raised area; in males it is slightly elongated, while in females it appears more rounded. Palpation of the testicular tissue through the abdominal wall may be possible in slightly older neonates but should be performed gently to avoid injury.
When visual inspection is inconclusive, supplementary methods include:
- Dissection: Carefully open the peritoneal cavity to expose the gonads. Testes are oval, firm, and located near the kidneys; ovaries are smooth, pale, and contain follicular structures.
- Molecular sexing: Extract DNA from a small tissue sample (e.g., ear notch) and perform PCR targeting the Sry gene (male-specific) and an autosomal control. Presence of Sry confirms a male genotype.
- Chromosomal analysis: Karyotyping of bone marrow cells provides definitive sex determination but is labor‑intensive and reserved for research requiring precise genetic confirmation.
Best practices:
- Conduct examinations in a well‑lit environment with appropriate magnification.
- Record AGD measurements for each animal to establish baseline data.
- Use non‑invasive visual methods first; reserve dissection or molecular techniques for ambiguous cases.
- Maintain strict aseptic technique when collecting tissue for DNA analysis to prevent contamination.
Accurate sex identification underpins experimental reproducibility, breeding program management, and ethical compliance in rodent research.