The effects of method of handling blood and saliva on progesterone concentration in the dairy cow

The radioimmunoassay for progesterone was used to determine the effects of holding blood for increasing lengths of time (0, 6, 12 hours) prior to serum separation on the concentration of progesterone. The radioimmunoassays for progesterone in whole blood and whole saliva were also evaluated as possible applications for progesterone testing under field conditions. Five consecutive l0 ml blood samples and two 10 ml saliva samples were collected from each of the ten Holstein cows in this study and were subjected to various handling treatments.

Significant differences in progesterone concentrations existed between serum and whole blood samples and between serum and saliva samples (P<.00l). Blood samples held for 6 or 12 hours at 4°C prior to centrifugation and separation of serum from cells resulted in lower serum progesterone concentrations than blood centrifuged immediately upon return to the laboratory (P<.05). Significant loss of progesterone was also determined in serum from blood held for 12 hours at 4°C when compared to serum from blood held for 6 hours (P<.05). The use of serum separation tubes (SST; Becton-Dickinson, Rutherford, N.J.) gave comparable results to serum collected from blood samples centrifuged immediately upon return to the laboratory.

The determination of progesterone in whole saliva samples showed a positive correlation (r=.76) (P<.00l) to the concentration of progesterone in serum samples. The concentration of progesterone was 23 + 5.3 percent (range 19-49 percent) of the progesterone levels determined in serum. No loss of progesterone was observed in saliva samples held at room temperature for 24 hours prior to storage at -20°.

iv The average recoveries of unlabeled progesterone added to serum and saliva samples were 88 + 7 percent and 64.7 + 9 percent, respectively. When increasing volumes of serum and saliva were analyzed, measured progesterone concentrations differed from expected concentrations by 15.9 + 3 percent and 30.0 + 11 percent. The interassay variation for serum samples in this study was 6.3 percent, while intraassay variation ranged from 5.8 to 2l.3 percent. The interassay variation for saliva samples was 32.7 percent, while intraassay variation ranged from 16 to 36 percent.

An intravenous dces of 250 mg progesterone resulted in a peak of 3400 ng/ml and 800 ng/ml in the plasma and saliva of a lactating non-pregnant dairy cow at five minutes post infusion. The progesterone concentrations in 15 matched plasma and saliva samples taken over a 24-hour period were highly correlated (r=.96) (p<.00l).