The TARGET kit contains testing cups and four different reagents that are used to determine the progesterone level from a blood sample. Although the testing protocol uses seven different steps and involves mixing several of the reagents, it provides results in about 15 minutes. This test requires blood serum, so the sample must be spun or allowed to clot for about an hour before performing the test.
Because the reagents are mixed at the time of testing, this kit has a longer shelf life than many others (about 15 months) and does not require that any of the reagents be stored frozen.
DETERMINATION OF THE FERTILE PERIOD
The visible signs of heat and breeding time (such asvulval swelling, end of vaginal discharge, or flagging) are only an approximate indication of the time of ovulation. These signs can vary by more than a week. Ovulation is triggered by the Luteinizing Hormone or LH which peaks two days before ovulation. Progesterone levels begin to rise at the same time as the LH surge. Ovulation occurs two days after the initial rise in progesterone. Predicting ovulation is doneaccurately by detecting the initial rise in progesterone. After ovulation occurs, the ovacytes take 2-3 days to ripen, making the most fertile period 5-6 days after the initial rise in progesterone. The relationship of the hormones, progesterone andLH, to ovulation and to the fertile period is shown in the figure. With the TARGET Canine Ovulation Timing kit, it is possible to detect the initial rise in progesterone. As seen in the figure, the progesterone level before ovulation is low (between 0 and 1.0 ng/ml). This corresponds to a bright blue (C1) test result. When the initial rise in progesterone occurs, the test result will be a light blue (C2).
GUIDELINE FOR WHEN TO TEST
1. In order to determine the baseline progesterone level, one serum sample can be run during the first five days of the proestrus cycle (characterized by the presence of blood and vulval swelling). The baseline progesterone level varies between 0-1 ng/ml in individual dogs and gives a bright blue color result similar to the C1 color on the color chart. Note the color shade.
2. To determine the initial rise in progesterone, begin testing 7-8 days after observing the first day of vaginal bleeding and vulval swelling (the beginning of proestrus). If the first result is the baseline color, bright blue (C1), then the progesterone level is still very low.
3. Retest every two days until the test results are a light blue that is similar to the C2 color on the color chart. This first fading of the color from C1 to C2 represents the initial rise in the progesterone level; ovulation will occur 1-2 days later.
4. Retest every day until the test results are faint blue similar to the C3 color on the color chart. This means ovulation has occurred and the egg is beginning to ripen and the fertile period begins 2-3 days later (See Figure). After obtaining a C3 color, natural matings and inseminations with fresh semen should take
TEST PREPARATION (For Serum Or Plasma)
Allow all kit components and samples to come to room temperature. Collect the blood sample into an EDTA or heparin coated tube (purple top tube) or in a dry tube (red top tube). Immediately after sample collection, invert the tube several times to mix. Spin the blood down with a centrifuge or allow the blood to clot by staying at room temperature for 1/2 to 1 hr and pour the serum into a clean glass tube. If the sample is not run immediately, store the sample in the refrigerator. For long term storage the sample should be frozen. Label the sample with the name, sample day and cycle day. Do not use whole blood. Do not run test with cold kits.
Store the kit in the refrigerator when not in use. Do not freeze.
Always reseal the plastic bag after removing a test cup.
Timing of the enzyme during Step 2 is IMPORTANT: one (1) minute.
Do not exchange test cups or reagents between different kits.
If you want to do a control, do steps 2-7, the result should be C1.
The TARGET Canine Progesterone kit can also be used to accurately determine the right time for a Cesarean section. About 24 hours before normal
INTERPRETATION OF THE RESULTS
The recommendations in the following table are only a guideline and all other available information concerning the dogs cycle should be considered. The color result of the test corresponds to the level of progesterone present. Use the enclosed color chart as a guide for evaluating and recording the result.
The C1 color corresponds to baseline serum progesterone levels.
The C2 color corresponds to the initial rise in progesterone levels. This is Day 0, the day of the LH peak, which is usually 1-2