Use Progesterone Testing to Decide When to Breed

Puppy drinking mother's milk

Precise timing of dog breeding through progesterone testing enhances litter production efficiency. Understanding the optimal time to breed a female dog in heat is fundamental to effective breeding management.

“Progesterone increases in a predictive manner, so we can use it to determine the important landmarks for breeding, such as the LH surge, ovulation and the fertile window,” says Gail McRae, DVM, who is completing a theriogenology residency at The Ohio State University College of Veterinary Medicine. She presented on progesterone testing at the AKC Breeder Symposium last fall in Columbus, Ohio.

Progesterone, a reproductive hormone, rises in the female's bloodstream before ovulation. This increase in progesterone is accompanied by the release of luteinizing hormone (LH) from the pituitary gland, which stimulates the release of eggs from the follicles, leading to ovulation and further progesterone elevation. The eggs typically mature within 48 to 72 hours, preparing them for fertilization by sperm.

“Progesterone is the only way we truly know when a female is ready to be bred. It also helps narrow down the whelping date to 63 days plus or minus one day from ovulation or 65 days plus or minus one day from the LH surge,” explains Dr. McRae who is also completing a master’s degree in which she is studying infertility in females.

Many breeders choose to collaborate with reproduction specialists who are board-certified in theriogenology or veterinarians with expertise in reproduction. This ensures the implementation of best practices in dog breeding. In cases where frozen semen is used, veterinary reproduction specialists play a crucial role in determining the optimal breeding time to achieve the best outcomes.

“The semen type dictates the right time to breed. Fresh semen on average lasts over 48 hours, chilled semen lasts about 48 hours, and frozen semen lasts 12 to 24 hours. If we are doing a frozen semen breeding, we usually wait until day three or four post-ovulation to breed a female to ensure the eggs are mature. We want the progesterone level to be around 15 nanograms per milliliter in the blood,” Dr. McRae says. 

“If we are using fresh or chilled semen, then we breed during post-ovulation on days one and three or on days two and four. We also can breed one time on days two to three post-ovulation for best results. This is because the sperm is viable for five to seven days and because it takes the eggs around two days to mature prior to fertilization. The rate of maturation of the eggs differs based on the amount of progesterone present.”

At Ohio State, the theriogenology team utilizes the Immulite machine to measure progesterone levels in female dogs. This two-hour progesterone test relies on chemiluminescence, a method that detects specific enzymes to provide an accurate quantitative measurement of progesterone.

“We recommend that breeders work with their veterinarian and have their blood samples sent out to a diagnostic testing laboratory to be tested on an Immulite machine to ensure accurate, comparable results,” Dr. McRae says. “Although there are many different progesterone-testing machines, some may only be reliable in the lower progesterone numbers, not when progesterone reaches the higher numbers that are so important when we are doing frozen semen or artificial insemination breedings.”

Breeders often opt for progesterone testing every two to three days, starting around five to seven days into the heat cycle. The cost of progesterone testing may vary depending on the veterinary clinic and location, but it typically ranges from $100 to $120 per test. It is common to conduct four to seven progesterone tests throughout the breeding cycle to accurately determine the optimal time for breeding.

Vaginal cytology is a technique veterinarians use to assess the stage of a female dog's heat cycle. By examining the degree of cornification in the epithelial cells of the vaginal lining, veterinarians can determine whether the female is in or out of season. Cornification refers to the process where cells lose their nuclei and flatten out. The levels of estrogen hormone, which rise during the heat cycle, contribute to the cornification of the vaginal epithelium.

“Vaginal cytology is an extremely useful tool for a veterinarian to use,” Dr. McRae says. “The changes we see during estrus are from the rise in estrogen. When we see lots of cornified cells, over 95 percent, the female is in estrus. If we think she is near the end of the fertile window, we can use vaginal cytology to see if she is still in heat. If we see 40 to 60 percent cornified cells with some white blood cells, this could indicate that we have missed the fertile window for breeding, though this also could indicate she is in proestrus and is too early to be bred.”

“A progesterone test taken at the same time as vaginal cytology will confirm whether she is going out of heat, or in diestrus, and thus too late to breed, or still very early, or in proestrus. If progesterone is low, this correlates with proestrus, and if progesterone is high, this correlates with diestrus.”

The value of using progesterone testing to plan and time a breeding is immeasurable. 

“If an owner is doing natural breedings and not having success, progesterone testing can be done to help nail down the timing better,” Dr. McRae says. 

“We recommend doing progesterone testing on females that may need a cesarean section, as this provides a three-day window for the due date if we can know when she had her LH surge and ovulated. When an important breeding using frozen semen is planned, we highly recommend using progesterone testing to ensure the best opportunity for success.”

“Progesterone testing is another tool in our toolbox that takes the guesswork out of breeding.”

Tools To Determine Ovulation

Veterinarians who specialize in reproduction often use a toolbox approach that include the following diagnostics to determine the appropriate time to breed.

  • Hormonal testing, including progesterone and luteinizing hormone (LH) assays, helps determine the presence and/or the levels of reproductive hormones in the blood. A surge in LH triggers the release of eggs, leading to an increase in progesterone levels just before ovulation in the female.
  • Vaginal cytology is used to determine the reproductive stage of a female dog, whether she is in estrus (in season) or diestrus (out of season). This is done by examining the degree of cornification in the epithelial cells, where the cells lose their nuclei and flatten out. Veterinarians frequently perform vaginal cytology on days when breedings are scheduled to confirm that the female is still within her fertile window.
  • Vaginoscopy, conducted with an endoscope, is a valuable tool during transcervical artificial insemination. It allows for a visual examination of the vaginal epithelium, aiding in determining the peak fertility period. The vaginal epithelium exhibits a crenulated or wavy outline during the optimal fertility phase. 

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