DEVELOPMENT OF BuPAG ANTIBODIES BASED IMMUNODIAGNOSTIC ASSAYS FOR EARLY DETECTION OF PREGNANCY IN BOVINES
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Date
2022
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ICAR-NDRI, KARNAL
Abstract
A good reproductive management in livestock depends on an early and precise diagnosis of
reproductive dysfunctions or aberrations. Good reproductive efficiency is necessary for
dairy animals to produce more milk throughout their lives. The most significant dairy animal
on the Indian subcontinent, the buffalo (Bubalus bubalis), is known for issues with a long
calving gap, delayed adolescence, and a high prevalence of anoestrus. The problem is
further made complicated by the lack of accurate early pregnancy diagnosis techniques at
field level. In bovine species, a number of pregnancy diagnosis techniques are used to detect
early pregnancy, but none of them is optimal due to the test's inherent limitations in terms
of sensitivity, accuracy, specificity, speed, and convenience of use. Pregnancy associated
glycoproteins (PAGs) are potential pregnancy biomarkers in farm animals that are secreted
during days 22 to 28 after fertilization from conceptus to maternal circulation. A total of 12
PAG isoforms and their variants were identified in buffalo during the early stages of
pregnancy, i.e., BuPAG 1, 2, 4, 6, 7, 8, 9, 13, 15, 16, 18, and one novel isoform. In our study
we have used two isoforms, i.e. BuPAG-1 and BuPAG-7 to develop two major
immunodiagnostic assays that are enzyme linked immunosorbent assay (ELISA) and lateral
flow immunoassay (LFIA). The goal of the present study was to develop and validate a
Sandwich-ELISA which can detect pregnancy in maternal serum at 35 days of post
insemination. Therefore, serum samples of both pregnant and non-pregnant animals were
collected from cows and buffaloes from 0-60 days after insemination. The developed
BuPAG-1 & 7 based ELISA assays were able to identify pregnancy at day 35 p.i. in serum
with accuracy of 86-93%. This is time saving for farmers and more efficient for laboratories.
Since ELISA requires laboratories and technicians, which can’t be done at field level, so we
have also tried to optimize and develop BuPAG-1 and 7 based lateral flow immunoassay
(LFIA), which can give results in 2-5 minutes after applying serum samples with accuracy
of 75-80% at day 35 p.i. Being affordable, sensitive, specific, user-friendly, rapid, robust,
and equipment-free, this point of care testing can be considered as one of the most promising
pregnancy diagnostic tool for farmers.