Visual ELISA for detection of pregnancy-associated glycoproteins (PAGs) in ewe serum

The aim of the present study was to evaluate the efficiency of visual ELISA-PAG for early pregnancy diagnosis based on the presence of PAG (pregnancy-associated glycoprotein) using sheep blood serum. Experiment 1: 140 ewes were divided into three groups with different stages of pregnancy. In the first group, 41 pregnant ewes were sampled at 26, 28, 30, and 32 days of pregnancy; in the second group, 65 ewes (49 pregnant with 30 days and 16 non-pregnant) were sampled; in the third group, 34 non-pregnant ewes were sampled. Experiment 2: 10 pregnant ewes were sampled weekly from day 35 of gestation until day 70 post-partum to verify the total period in which PAG can be detected in the blood serum by the test. Transrectal ultrasound was used as a gold standard. The detection or non-detection of PAG was analyzed by the logistic model PROC GENMOD of SAS; differences were detected by the chi-squared test. In group 1, there were no differences between the results from 28, 30, and 32 days of gestation, but samples from 30 days were easier to interpret in comparison to 28 days, with a sensitivity of 97.56%. In the second group, using 65 sheep, visual ELISA-PAG showed 100% sensitivity and 93.75% specificity, which indicates the diagnosis of an animal as a false positive. In the third group, 97.06% of the sheep were confirmed as negative and 2.94% as positive, again indicating the presence of a false positive. In 100% of the sheep, the PAG remained in the blood circulation throughout the antepartum period until birth and seven days post-partum, declining thereafter. Based on our results, the visual ELISA-PAG is an effective method for the early diagnosis of pregnancy in sheep and can be performed from day 30 of gestation.