Dot-Enzyme Linked Immunosorbant Assay for detection of Paramphistomum cervi antibodies in Rabbits
Tariq Ahamed1. Mohammad Lateef Reshi2, M..Z Chasti1, Syed Tanveer1, Abdul Baqui1. Zaffer Amin Shah1 Bashir Ahamed Fomida2 and O.K Raina3
1P.G Department of Zoology University of Klashmir (India) 2Department of Immunology and Microbiology SKIMS Soura, Kashmir (India) 3Department of Pariastology IVRI, Izatnagar, Barielly U.P. (India
ABSTRACT
A Dot-ELISA for detection of Paramphistomum cervi antibodies in rabbits was standardized. Soluble antigen was coated on nitrocellulose membrane in the form of dot and incubated at 37oC for 1 hr. Remaining sites were blocked by incubation with 3% skimmed milk powder, followed by incubation with anti-serum. Blood antibodies were detected by incubation with anti-rabbit IgG-HRP conjugate. Enzymatic reaction was carried out with H2O2 as substrate and TMB as the indicator dye. Positive results was recorded by the appearance of red dots. Optimum concentration of Ag was found to be 80ng/dot. Sera from all the immunized rabbits was titrated by checkerboard.