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Concanavalin A- Sandwich-ELISA for the detection of antibodies against Newcastle disease virus (NDV) in chicken

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Title Concanavalin A- Sandwich-ELISA for the detection of antibodies against Newcastle disease virus (NDV) in chicken
 
Creator T.R. Kannaki, E. Priyanka and Santosh Haunshi
 
Subject Chicken
Newcastle Disease
Con A
 
Description Not Available
Concanavalin A (Con A), a lectin interacts with carbohydrate moieties of viruses and provide stable and sensitive detection
when used as a capture agent. Indirect ELISA methods need purified Newcastle disease virus (NDV) or recombinant
antigens for adsorption, whereas use of Con A as capture agent will enable the use of non-purified and non-concentrated
virus as antigen replacing costly and time-consuming virus purification step. Con A based sandwich ELISA with nonpurified
NDV whole virus antigen with single serum dilution format has been developed in this study. The optimum
concentrations of the capture agent, Con A and non-purified antigen preparations were determined by checker-board titration.
Briefly, microplates were coated with predetermined optimum concentration of ConA (0.5 mg/ml; 50μg per well) and
incubated for 18h at 4C. After washing, allantoic fluid with Newcastle disease virus (NDV) LaSota (HA titre, 210) at a
constant predetermined dilution (1:1; 50μl) was coated and incubated for 45 min at 37C, followed by blocking with 2 %
bovine serum albumin for 45 min at 37C. The antigen coated plates were used in the detection of antibody titre against
NDV in serum samples at single serum dilution of 1: 500. Then, wells were added with goat anti-chicken IgG horseradish
peroxidase conjugate and incubated for 1h at 37C, followed by addition of TMB substrate and the plates were read
spectrophotometrically at 450 nm. ELISA antibody titres were determined by standard serial dilution of positive sera and
endpoints were calculated by a subtraction method. By using positive negative threshold curve (PNT), intercept and slope
of the standard curve were calculated. Total of 271 random chicken serum samples were analyzed for antibodies against
NDV by Haemagglutination inhibition assay (HI), indirect ELISA and compared with the Con A- S- ELISA developed in
this study. The Con A-S-ELISA showed a high coefficient of correlation (r=0.85, n=271, P
Not Available
 
Date 2019-12-05T05:23:37Z
2019-12-05T05:23:37Z
2019-01-31
 
Type Research Paper
 
Identifier Not Available
DOI: 10.18805/ijar.B-3711
http://krishi.icar.gov.in/jspui/handle/123456789/27772
 
Language English
 
Relation Not Available;
 
Publisher Agriculture Research Communication centre