Development of an indirect ELISA and study of immune response to Clostridium chauvoei in cattle
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Date
2023-08-17
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LUVAS Hisar
Abstract
In the present study, an Indirect ELISA was developed to demonstrate antibodies
against Black quarter (B.Q.) vaccine antigen. The B.Q. vaccine antigen was retrieved from
the vaccine formulation by dialysis to remove the formalin and was used as antigen. The
hyper immune serum raised in rabbits against B.Q. vaccine antigen was used as positive
control in ELISA development. The optimized dilutions of the reagent used in ELISA were 5
µg/ml for coating antigen; 1:200 dilution for hyper-immune rabbit serum; 1:200 dilution for
pre-immune Rabbit serum; 1:200 dilution for Cattle serum and 1:8000 dilution for Protein G
HRP conjugate. The cut-off value (mean+3SD) for the developed ELISA calculated using the
naive calf serum (n=37) was found to be 0.347135.
In this study, the animals kept at Animal Farm, Department of Animal Genetics and
Breeding were divided into three groups with 10 animals in each group: Group I (calves; < 9-
month age), Group II (heifers; 18-to-26-month age), and Group III (adults; > 26-months age).
In Group I 70% of the calves; in Group II 90% of the heifers and in Group III 100% of the
adults showed one month post vaccination immune response (in terms of OD450) above cut-off
value. However, the kinetics of the immune response studied over a period of six months was
found to be aberrant which required to be further investigated. Based on the above study, we
can conclude that some preliminary base had been developed and further improvement and
refinement is required in the ELISA so that we can calculate sero-conversion in vaccinated
animals in terms of antibody titer