107 The effect of storage time on progesterone concentration in refrigerated whole mare blood

Abstract

The majority of equine progesterone research is over 30 years old and data is lacking regarding the use of current laboratory analysis methods. The research objective was to evaluate the effect of storage time on progesterone concentrations in refrigerated whole mare blood. Mares were ultrasounded twice weekly to confirm follicular development, ovulation, and cycling status before enrollment into the study. Twelve cycling, Quarter Horse mares were enrolled into the study from March to July, and blood samples were collected during multiple estrous cycles for an average of 2 cycles per mare. Mares were ultrasounded daily once a 35 mm follicle was detected on an ovary until ovulation, and then every other day to monitor corpus luteum formation. Open mares were reenrolled into the study on a subsequent estrous cycle, allowing for 28 total blood collection periods. Blood collections occurred 5 d after the ovulation of a previously recorded follicle and confirmation via ultrasonography of a corpus luteum on the same ovary. On each sample collection day, 6 cc of whole blood was collected from the mare into 5 vacutainer serum blood collection tubes from a single venipuncture, for a total of 30 cc of whole blood. The 5 blood samples were stored under refrigerated conditions at 4°C for 0, 6, 12, 24, and 48 h. The 0-h treatment was centrifuged once an adequate blood clot had formed or 30 min post-collection. Following the assigned storage time, each blood sample was centrifuged for 15 min and then serum was frozen at −22°C until analyzed for P4 using solid phase radioimmunoassay kit reagents validated for equine plasma. Statistical analysis of progesterone data was analyzed as a repeated measures design using the GLM procedure of SAS. Progesterone concentrations were similar among the refrigerated storage times at 0 ( P = 0.278), 6 ( P = 0.230), 12 ( P = 0.198), 24 ( P = 0.243), and 48 ( P = 0.282) hours. In summary, refrigerated samples of whole blood from mares may be stored for up to 48 h without an effect on progesterone concentration analysis.