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Purification of pregnancy-associated glycoproteins from late-pregnancy Bubalus bubalis placentas and development of a radioimmunoassay for pregnancy diagnosis in water buffalo females

Olimpia Barbato1*, Noelita Melo de Sousa2, Vittoria Lucia Barile3, Claudio Canali1 and Jean-François Beckers2

  • * Corresponding author: Olimpia Barbato

  • † Equal contributors

Author Affiliations

1 Department of Biopathological Veterinary Science, Faculty of Veterinary Medicine, University of Perugia, Perugia, 06126, Italy

2 Laboratory of Animal Endocrinology and Reproduction, Faculty of Veterinary Medicine, University of Liege, Liege, 4000, Belgium

3 Animal Production Research Centre, Italian Agricultural Research Council, Monterotondo, 00016, Italy

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BMC Veterinary Research 2013, 9:89  doi:10.1186/1746-6148-9-89

Published: 1 May 2013



Pregnancy-associated glycoproteins (PAGs) were first described as placental antigens present in the blood serum of the mother soon after implantation. Here, we describe the purification of several pregnancy-associated glycoproteins from water buffalo placenta (wbPAGs). A specific radioimmunoassay (RIA) was developed for early pregnancy diagnosis in buffalo species.


Amino-terminal microsequencing of immunoreactive placental proteins allowed the identification of eleven wbPAGs sequences [Swiss-Prot accession numbers: P86369 to P86379]. Three polyclonal antisera (AS#858, AS#859 and AS#860) were raised in rabbits against distinct wbPAG fractions. A new RIA (RIA-860) was developed and used to distinguish between pregnant (n = 33) and non-pregnant (n = 26) water buffalo females.


Our results confirmed the multiplicity of PAG expression in buffalo placenta. In addition, the RIA-860 system was shown to be sensitive, linear, reproducible, accurate and specific in measuring PAG concentrations in buffalo plasma samples from Day 37 of gestation onwards.

Water buffalo; Placenta; Pregnancy-associated glycoprotein; Purification; Radioimmunoassay; Polyclonal antiserum; Pregnancy diagnosis; N-terminal amino acid sequence