Indian Journal of Animal Research

  • Chief EditorK.M.L. Pathak

  • Print ISSN 0367-6722

  • Online ISSN 0976-0555

  • NAAS Rating 6.50

  • SJR 0.263

  • Impact Factor 0.4 (2024)

Frequency :
Monthly (January, February, March, April, May, June, July, August, September, October, November and December)
Indexing Services :
Science Citation Index Expanded, BIOSIS Preview, ISI Citation Index, Biological Abstracts, Scopus, AGRICOLA, Google Scholar, CrossRef, CAB Abstracting Journals, Chemical Abstracts, Indian Science Abstracts, EBSCO Indexing Services, Index Copernicus
Indian Journal of Animal Research, volume 53 issue 12 (december 2019) : 1656-1660

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

T.R. Kannaki, E. Priyanka, Santosh Haunshi
1Avian Health Lab, Directorate of Poultry Research, Rajendranagar, Hyderabad-500 030, Telangana, India.
Cite article:- Kannaki T.R., Priyanka E., Haunshi Santosh (2019). Concanavalin A- Sandwich-ELISA for the detection of antibodies against Newcastle disease virus (NDV) in chicken. Indian Journal of Animal Research. 53(12): 1656-1660. doi: 10.18805/ijar.B-3711.
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 non-purified 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<0.01) and an agreement of ê=0.99 with the commercially available Indirect-ELISA. The relative sensitivity and specificity were 98% and 85% respectively in comparison to HI test. Hence, the Con A-S-ELISA is a simple, easy and effective for monitoring serum antibody levels against NDV.
  1. Alexander, D.J., (1991). Newcastle disease and other avian paramyxovirus infections, In: Diseases of Poultry, 9th edition. [Calnek, B.W. (Ed.)], Iowa State University Press, Ames, pp. 496–519.
  2. Alexander, D.J., Bell, J.G., Alders, R.G., (2004). A Technology Review: Newcastle Disease - With Special Emphasis on Its Effects on Village Chickens. FAO Animal Production and Heath Paper, Food and Agriculture Organization of the United Nations, Rome, Italy 161-163.
  3. Allan, W.H., Gough,R.E., (1974). A standard haemagglutination inhibition test for Newcastle disease. (1) Comparison of macro and micro methods. Vet. Rec. 95 (6): 120-123.
  4. Becht, H., Rott, R., Klenk, H.D., (1972). Effect of Concanavalin A on cells infected with enveloped RNA viruses. J. Gen. Virol. 14(1):1-8.
  5. Bronzoni, R.V., Fatima, M., Montassier,S., Pereira, G.T., Gama, N.M., Sakai, V.,Montassier H.J., (2005). Detection of infectious bronchitis virus and specific anti- viral antibodies using a Concanavalin A-Sandwich-ELISA. Viral Immunol. 18(3):569-78.
  6. Brown, J., Resurreccion, R.S., Dickson, T.G., (1990).The relationship between the hemagglutination-inhibition test and the enzyme-    linked immunosorbent assay for the detection of antibody to Newcastle disease. Avian Dis. 34(3):585-7.
  7. Chowdhury, M.MI., Islam, M.T., Aktar, A., Bhuiyan, M.K.J., Kamal, M.M., Haque, M.A., et al (2011). Comparative performance of in-house and commercially available ELISA Kits for the detection of antibody against newcastle disease virus vaccine. Progress Agric. 22(1 & 2): 55 – 64.
  8. de Oliveira, E.S., Silva, K.R., Fernando, F.S., Gonçalves, M.C., Fernandes, C.C., et al (2013). A liquid-phase-blocking concanavalin A enzyme-linked immunosorbent assay for the detection of antibodies against Newcastle disease virus in serum of free-    ranging pigeons. J. Vet. Diagn. Invest. 25(6):720-726.
  9. Errington, W., Steward, M., Emmerson, P.T., (1995). A diagnostic immunoassay for Newcastle disease virus based on the nucleocapsid protein expressed by a recombinant baculovirus. J. Virol. Methods 55(3):357-65.
  10. Makkay, A,M., Krell, P.J.,Nagy, E., (1999). Antibody detection-based differential ELISA for NDV-infected or vaccinated chickens versus NDV HN-subunit vaccinated chickens. Vet. Microbiol. 66(3):209-22.
  11. Miers, L.A., Bankowski, R.A., Zee, Y.C., (1983). Optimizing the enzyme-linked immunosorbent assay for evaluating immunity of chickens to Newcastle disease. Avian Dis. 27(4):1112-25
  12. Mohan, C.M., Dey, S., Rai, A., Kataria, J.M., (2006). Recombinant haemagglutinin neuraminidase antigen-based single serum dilution ELISA for rapid serological profiling of Newcastle disease virus. J. Virol. Methods 138(1-2):117-22.
  13. Pradhan, S.K., Kamble, N.M., Pillai, A.S., Gaikwad, S.S., Khulape, S.A., Reddy, M.R., Mohan, C.M., Kataria, J.M.,Dey, S., (2014). Recombinant nucleocapsid protein based single serum dilution ELISA for the detection of antibodies to infectious bronchitis virus in poultry. J Virol. Methods 209:1-6.
  14. R R., Roy, P., (2014). Recombinant hexon antigen based single serum dilution ELISA for rapid serological profiling against fowl adenovirus-4 causing hydropericardium syndrome in chickens. J Virol. Methods 207:121-7.
  15. Ramadass, P., Parthiban, M.,Thiagarajan, V., Chandrasekar,M.,Vidhya, M., Raj, G. D., (2008). Development of single serum dilution ELISA for detection of infectious bursal disease virus antibodies. Vet.Arhiv. 78: 23-30.
  16. Rivetz, B., Weisman, Y., Ritterband, M., Fish, F.,Herzberg, M., (1985). Evaluation of a novel rapid kit for the visual detection of Newcastle disease virus antibodies. Avian Dis. 29(4):929-42.
  17. Snyder, D.B., Marquardt, W.W., Mallinson, E.T., Russek, E., (1983). Rapid serological profiling by enzyme-linked immunosorbent assay. I. Measurement of antibody activity titer against Newcastle disease virus in a single serum dilution. Avian Dis. 27(1):161-70.
  18. Tizard, L., (1982). An Introduction to Veterinary Immunology. Saunder Company Published. Sussex. England 

Editorial Board

View all (0)