Compositions and methods for early pregnancy diagnosis
View Patent ↗Pregnancy-associated glycoproteins (PAGs) are structurally related to the pepsins, thought to be restricted to the hoofed (ungulate) mammals and characterized by being expressed specifically in the outer epithelial cell layer (chorion/trophectoderm) of the placenta. By cloning expressed genes from ovine and bovine placental cDNA libraries, the inventors estimate that cattle, sheep, and most probably all ruminant Artiodactyla, possess possibly 100 or more PAG genes, many of which are placentally expressed. The PAGs are highly diverse in sequence, with regions of hypervariability confined largely to surface-exposed loops. Selected PAG that are products of the invasive binucleate cells, expressed highly in early pregnancy at the time of trophoblast invasion and expressed weakly, if at all, in late gestation are useful in the early diagnosis of pregnancy. In a preferred embodiment, the invention relates to immunoassays for detecting these PAGs.
1. A method for detecting pregnancy in a bovine animal comprising:
(a) obtaining a sample from said animal; and
(b) detecting at least one pregnancy associated antigen (PAG) wherein said PAG is selected from the group consisting of BoPAG5, BoPAG9, BoPAG15, BoPAG18, and BoPAG19; wherein the PAG is present in early pregnancy and absent at about two months post-partum;
whereby the presence of the PAG indicates that said animal is pregnant.
2. The method of claim 1 , wherein said sample is saliva, serum, blood, milk or urine.
3. The method of claim 1 , wherein said detecting comprises immunologic detection.
4. The method of claim 3 , wherein said immunologic detection comprises detection with polyclonal antisera or with a monoclonal antibody preparation.
5. The method of claim 3 , wherein said immunologic detection comprises ELISA, RIA or Western blot.
6. The method of claim 1 , wherein the PAG is BoPAG5.
7. The method of claim 1 , further comprising detecting a second or third PAG in said sample.
8. The method of claim 5 , wherein said ELISA is a sandwich ELISA comprising binding of a PAG to a first antibody preparation fixed to a substrate and a second antibody preparation labeled with an enzyme.
9. The method of claim 8 , wherein said enzyme is alkaline phosphatase or horseradish peroxidase.
10. The method of claim 8 , wherein said first antibody preparation is monoclonal.
11. The method of claim 1 , wherein the PAG is BoPAG9.
12. The method of claim 1 , wherein the PAG is BoPAG15.
13. The method of claim 1 , wherein the PAG is BoPAG18.
14. The method of claim 1 , wherein the PAG is BoPAG19.