A Simple Approach to Progesterone Testing for Mating and Parturition
British Small Animal Veterinary Congress 2008
Angelika von Heimendahl, DVM, MScAg, BVM(Berlin), DECAR, MRCVS
Veterinary Reproduction Service, Clarendon Street Veterinary Surgery

Progesterone is the most important hormone in female dog reproduction. It is easy to test for and can be used to look at ovulation, parturition and other cyclical abnormalities. In order to interpret results one needs to be aware of the unique pattern of progesterone secretion in the bitch.

Progesterone Cycle in the Bitch

Progesterone is a steroid hormone produced by the corpora lutea and, uniquely in the bitch, by maturing follicles. Granulosa cells grow within the unruptured follicle and plasma progesterone levels follow a characteristic rise before, during and after ovulation. Maximum concentrations are reached between 20 and 30 days after the end of oestrus, whether the bitch is pregnant or not. From then on a gradual decline takes place with basal levels around 60-70 days after ovulation. Non-pregnant animals tend to have a slightly longer luteal phase than pregnant animals. The drop in progesterone will induce a rise in prolactin, which will stimulate milk production. In a pack animal situation most of the females will come into milk regardless of whether or not they have produced offspring themselves and will be able to help feeding.

How To Test For Progesterone

There are basically two ways of measuring progesterone: in-house or at a laboratory.

The in-house tests come in packs of five or ten and have to be kept refrigerated. They are semiquantitative enzyme-linked immunosorbent assay (ELISA) tests based on a colour change. The author prefers the tests that run two progesterone standards alongside the sample rather than a colour chart. The standards are set at raised levels of 3 ng/ml and post-ovulation at 10 ng/ml. The tests are fairly easy to run, although accuracy in handling and timing is required, and will give results within 30 minutes. The shelf-life of the tests can make them expensive to keep, but the author finds that freezing the conjugates and thawing them in warm water before use can extend the life. The standards themselves are stable for a long time.

The laboratory tests are either radio immunoassays or chemiluminescence assays and, just to make things more complicated, are reported either in ng/ml or nmol/l (1 ng = 3 nmol). Laboratory tests are more accurate and, because of the precise value of progesterone levels, predictions can be made more accurately. As a rule of thumb once the progesterone level starts to rise, it doubles every 2 days.

Progesterone Testing For Mating

To determine the days of mating two factors have to be considered: ovulation and maturation. Bitches will ovulate at around 6 ng/ml (18 nmol/l). After ovulation the oocytes have to mature for a further 2 days before they can be fertilised. Once mature they will be viable for another 2 days. Spermatozoa will take 6-12 hours to capacitate and reach the oviduct.

Possible testing regime:

 Plasma progesterone levels of <1 ng/ml (3 nmol/l): retest in 4 days

 Plasma progesterone levels of <2 ng/ml (6 nmol/l): retest in 3 days

 Plasma progesterone levels of >2 ng/ml (6 nmol/l): retest in 2 days

 Plasma progesterone levels of >25 ng/mol (75 nmol/l) usually indicate the end of the fertile period

Possible mating regime:

 Fertilisation period: ovulation +1 day until +4 days

 Two matings: ovulation +1 day and +3 days or ovulation +2 days and +4 days

 One mating: ovulation +2 days or ovulation +3 days

When using a laboratory the post office 'next day delivery' service is very useful. One should also consider weekends and bank holidays. Given the predictable rise of progesterone and the time available after ovulation this system normally works well.

Progesterone Testing For Parturition

The relatively short gestation length of the bitch makes it important to be able to determine the physiological end of the pregnancy, especially when intervention is necessary. Any puppies born before 57 days after fertilisation have poor survival rates. Although the actual duration of a pregnancy from fertilisation in the bitch is around 61 days ± 48 hours the information available to the veterinary surgeon can be very uncertain. Multiple matings and long sperm survival in the reproductive tract (up to 7 days) can make it very difficult to know how many days the bitch is pregnant. The author also finds it useful to count the days from mating in her own calendar as owners sometimes make mistakes.

Letting owners measure body temperature three times a day can be useful especially in smaller and medium-sized dogs. The temperature will fluctuate slightly in the last week but drop 2°C 12-24 hours before parturition. The drop is not always so marked in larger dogs. The change in body temperature is caused by the drop in plasma progesterone.

Plasma progesterone levels decline to <1 ng/ml 24 hours before parturition. As it is important to have the results immediately, the ELISA kits for ovulation testing can be used. The sample will be stronger in colour (lower progesterone) than the low standard. In cases of suspected primary inertia the test can give results within 30 minutes. If the progesterone is still high the bitch is not ready, if it is low the puppies are ready and a Caesarean section can be attempted. In case of an elective Caesarean (e.g., single puppy pregnancy, Bulldog) the test can be repeated every day towards the end of gestation until the drop in progesterone is visible.

Progesterone Testing For Some Other Conditions

Progesterone testing can be useful in a range of other reproductive conditions:

 Split heats are quite common in young bitches, but also occur in around 5% of older animals. The first season is quite short and non-ovulatory, followed by a shorter than normal interoestrous period with a normal season. Progesterone rises only slightly and drops off again as ovulation hasn't taken place. This can be tested any time in the next 40 days following the season.

 Delayed puberty is suspected if a bitch has not shown any seasons by the time she is 2 years of age. To exclude a silent oestrus having taken place in the previous 2 months a progesterone level would be the first test before further investigations are attempted.

 Silent oestrus can be a big problem as some animals show no or hardly any signs of swelling or discharge during their season. In order to breed from theses females testing has to start on the first day of any signs to make sure the fertile period is not missed.

 Hypoluteoidism is a condition that seems to affect some breeds (e.g., German Shepherd Dog, Old English Sheepdog) causing the reabsorption or abortion of puppies before the end of gestation. Regular progesterone testing from 30 days of gestation and supplementation can be useful.


Progesterone testing is a very useful, inexpensive and readily available tool in female dog reproduction. It can be used at different stages of the reproductive cycle and give information about the right time of mating and parturition as well as some abnormalities.

Speaker Information
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Angelika von Heimendahl, DVM, MScAg, BVM(Berlin), DECAR, MRCVS
Veterinary Reproduction Service
Clarendon Street Veterinary Surgery
Cambridge, UK

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