Recombinant bovine pregnancy-associated glycoprotein 1 (PAG1) production involves several critical steps, starting with the isolation of the target gene that encodes the 54-380aa of the bovine PAG1. The gene is fused with an N-terminal 6xHis-tag gene and then cloned into an expression vector, which is introduced into E. coli cells using transformation. The E. coli cells express the recombinant PAG1 protein, which is collected from the cell lysate. Purification of the protein is typically achieved using affinity chromatography. The final stage involves measuring the protein's purity through SDS-PAGE. Its purity is over 90%.
Bovine PAG1 is a glycoprotein produced by trophoblast cells in the placenta and is considered an important marker for pregnancy in cows [1][2]. It can be detected in the serum and milk of pregnant cows. Various studies have focused on developing assays, such as radioimmunoassays and ELISAs, to measure PAG1 levels for accurate pregnancy diagnosis in cattle [3][4][5]. Research has shown that PAG1 levels in plasma can vary throughout gestation and are influenced by factors like fetal number [6]. Additionally, PAG1 has been associated with predicting abortion in dairy cows infected with Neospora caninum [7].
The detection and quantification of PAG1 play a crucial role in reproductive management in cattle. Studies have compared methods such as ultrasonography, ELISA, and radioimmunoassays for pregnancy diagnosis, emphasizing the importance of PAG1 testing in dairy cows [5][8]. Furthermore, research has explored the potential of using PAG1 levels to monitor induced abortions and hormonal profiles in heifers [9].
References:
[1] Z. Perényi, O. Szenci, J. Sulon, P. Drion, & J. Beckers, Comparison of the ability of three radioimmunoassay to detect pregnancy‐associated glycoproteins in bovine plasma, Reproduction in Domestic Animals, vol. 37, no. 2, p. 100-104, 2002. https://doi.org/10.1046/j.1439-0531.2002.00341.x
[2] K. Klisch, N. Sousa, J. Beckers, R. Leiser, & A. Pich, Pregnancy associated glycoprotein‐1, ‐6, ‐7, and ‐17 are major products of bovine binucleate trophoblast giant cells at midpregnancy, Molecular Reproduction and Development, vol. 71, no. 4, p. 453-460, 2005. https://doi.org/10.1002/mrd.20296
[3] A. Zoli, L. Guilbault, P. Delahaut, W. Ortiz, & J. Beckers, Radioimmunoassay of a bovine pregnancy-associated glycoprotein in serum: its application for pregnancy diagnosis1, Biology of Reproduction, vol. 46, no. 1, p. 83-92, 1992. https://doi.org/10.1095/biolreprod46.1.83
[4] M. Friedrich and W. Holtz, Establishment of an elisa for measuring bovine pregnancy‐associated glycoprotein in serum or milk and its application for early pregnancy detection, Reproduction in Domestic Animals, vol. 45, no. 1, p. 142-146, 2010. https://doi.org/10.1111/j.1439-0531.2008.01287.x
[5] O. Szenci, J. Beckers, P. Humblot, J. Sulon, G. Sasser, M. Taverneet al., Comparison of ultrasonography, bovine pregnancy-specific protein b, and bovine pregnancy-associated glycoprotein 1 tests for pregnancy detection in dairy cows, Theriogenology, vol. 50, no. 1, p. 77-88, 1998. https://doi.org/10.1016/s0093-691x(98)00115-0
[6] O. Patel, J. Sulon, J. Beckers, T. Takahashi, M. Hirako, N. Sasakiet al., Plasma bovine pregnancy-associated glycoprotein concentrations throughout gestation in relationship to fetal number in the cow, Acta Endocrinologica, p. 423-428, 1997. https://doi.org/10.1530/eje.0.1370423
[7] I. García-Ispierto, S. Almerı́a, B. Serrano, N. Sousa, J. Beckers, & F. López‐Gatius, Plasma concentrations of pregnancy‐associated glycoproteins measured using anti‐bovine pag‐2 antibodies on day 120 of gestation predict abortion in dairy cows naturally infected with neospora caninum, Reproduction in Domestic Animals, vol. 48, no. 4, p. 613-618, 2012. https://doi.org/10.1111/rda.12134
[8] O. Patel, K. Kizaki, T. Takahashi, K. Imai, & K. Hashizume, Quantitative analysis throughout pregnancy of placentomal and interplacentomal expression of pregnancy‐associated glycoproteins‐1 and ‐9 in the cow, Molecular Reproduction and Development, vol. 67, no. 3, p. 257-263, 2004. https://doi.org/10.1002/mrd.20017
[9] F. Lobago, H. Gustafsson, M. Bekana, J. Beckers, & H. Kindahl, Clinical features and hormonal profiles of cloprostenol-induced early abortions in heifers monitored by ultrasonography, Acta Veterinaria Scandinavica, vol. 48, no. 1, 2006. https://doi.org/10.1186/1751-0147-48-23
[10] T. Lestari, Study of the anti – pregnancy - associated glycoprotein (anti – pag) resulted from antigen pag immunization, as prospective early pregnancy detector in animals, Cercetari Agronomice in Moldova, vol. 49, no. 3, p. 111-118, 2016. https://doi.org/10.1515/cerce-2016-0030
