Korean Journal of Veterinary Service (한국동물위생학회지)

The Korean Society of Veterinary Service (한국동물위생학회)
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Development of monoclonal antibody capture ELISA for the detection of antibodies against transmissible gastroenteritis virus

  • Oh, Yeonsu (Department of Veterinary Pathology, College of Veterinary Medicine and Institute of Veterinary Science, Kangwon National University) ;
  • Tark, Dongseob (Korea Zoonosis Research Institute, Chonbuk National University)
  • Received : 2018.11.09
  • Accepted : 2019.02.25
  • Published : 2019.03.30
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Abstract

Transmissible gastroenteritis (TGE) is a disease confined to pigs of all ages, and can be a significant cause of economic loss in breeding herds, primarily because of the very high piglet mortality. The causative agent is a coronavirus, an enveloped positive strand RNA virus and closely related but non-enteropathogenic porcine respiratory coronavirus (PRCV). Although the TGEV has declined with its innocent relative, PRCV, further genome changes could not be excluded. Therefore, the herd-level immunity against this virus is important for the prevention of disease and should be carefully monitored. The aim of this study is to develop monoclonal antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) which can rapidly and accurately determine a large numbers of serum samples for surveillance purpose, and to compare the ELISA with a TGEV-specific serum neutralization test. The MAC-ELISA was sufficiently achieved, and the comparison with the virus-specific serum neutralization assays for 713 sera from pig farms showed a high correlation ($r^2=0.812$, P<0.001). The specificity and sensitivity of MAC-ELISA for the serum neutralization test 91.9% and 91.6%, respectively, which means that the antibody detected by the MAC-ELISA could be said to be protective antibodies. In conclusion, the developed MAC-ELISA would be very helpful in evaluating protective antibodies against TGEV.

Keywords

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Fig. 1. Determination of the optimal monoclonal antibody (Mab) concentration. Dilution factors of each component are as follows: 1:100 for recombinant spike protein, 1:40 for porcine serum, 1:1,000 for conjugate.

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Fig. 2. Determination of the optimal molar concentration of Zn acetate for concentration of recombinant spike protein. Dilution factors of each component in MAC-ELISA for optimizing Zn acetate concentration are as follows: 1:4,000 for monoclonal antibody, 1:80 for recombinant spike protein, 1:40 for porcine serum, 1:1,000 conjugate.

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Fig. 3. Determination of the optimal antigen concentration. Dilution factors of each component in MAC-ELISA for optimizing recombinant spike protein are as follows: 1:4,000 for monoclonal antibody, 1:40 for porcine serum, 1:1,000 conjugate.

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Fig. 4. Determination of the optimal serum and conjugate concentration. A; Dilution factors of each component in MAC-ELISA for optimizing serum concentration are as follows: 1:4,000 for monoclonal antibody, 1:100 for recombinant spike protein, 1:1,000 conjugate. B; Dilution factors of each component in MAC-ELISA for optimizing conjugate concentration are as follows: 1:4,000 for monoclonal antibody, 1:100 for recombinant spike protein, 1: 40 for test serum.

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Fig. 5. Correlation between virus neutralization test and monoclonal antibody capture ELISA (MAC-ELISA). Dilution factors of each component in MAC-ELISA are as follows: 1:4,000 for monoclonal antibody, 1:100 for recombinant spike protein, 1:40 for porcine serum, 1:1,000 conjugate.

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Fig. 6. Stability of the monoclonal antibody capture ELISA kit containing plate and other components during 7 months.

Table 1. Comparative analysis between the virus neutralization (VN) test and and MAC-ELISA to detect anti-TGEV antibodies in field porcine serum samples

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References

  1. Benfield DA, Haelterman EO, Burnstein T. 1978. An indirect fluorescent antibody test for antibodies to transmissible gastroenteritis virus of swine. Can J Comp Med 42: 478-482.
  2. Bohac J, Derbyshire JB, Thorsen J. 1975. The detection of transmissible gastroenteritis viral antigens by immunodiffusion. Can J Comp Med 39(1):67-75.
  3. Bohac J, Derbyshire JB. 1976. The detection of transmissible gastroenteritis viral antibody by immunodiffusion. Can J Comp Med 40: 161-165.
  4. Cesbron JY, Capron A, Ovlague G, Santoro F. 1985. Use of a monoclonal antibody in a double-sandwich ELISA for detection of IgM antibodies to Toxoplasma gondii major surface protein(P30). J Immunol Methods 83: 151-158. https://doi.org/10.1016/0022-1759(85)90068-7
  5. Cho HJ, Bohac JG. 1985. Sensitivity and specificity of an Enzyme-linked immunosorbent assay for the detection of infectious bovine rhinotracheitis viral antibody in cattle. Can J Comp Med 49: 189-194.
  6. Cho HJ, Masri SA, Deregt D, Yeo SG, Thomas EJG. 1991. Sensitivity and specificity of an enzyme-linked immunosorbent assay for the detection of bovine virus antibody in cattle. Can J Vet Res 55: 56-59.
  7. Correa I, Jimenez G, Sune C, Bullido MJ, Enjuanes L. 1988. Antigenic structure of the E2 glycoprotein from transmissible gastroenteritis coronavirus. Virus Res 10: 77-94. https://doi.org/10.1016/0168-1702(88)90059-7
  8. Delmas B, Gelfi J, Laude H. 1986. Antigenic structure of transmissible gastroenteritis virus. II. Domains in the peplomer glycoprotein. J Gen Virol 67: 1405-1418. https://doi.org/10.1099/0022-1317-67-7-1405
  9. Derbyshire JB, Jessett DM, Newman G. 1969. An experimental epidemiological study of porcine transmissible gastroenteritis. J Comp Pathol 79: 445-452. https://doi.org/10.1016/0021-9975(69)90064-4
  10. Doyle LP, Hutching LM. 1946. A transmissible gastroenteritis in pigs. J Am Vet Med Assoc 108: 257-259.
  11. Gebauer F, Posthumus WP, Correa I, Sune C, Smerdou C, Sanchez CM, Lenstra JA, Meloen RH, Enjuanes L. 1991. Residues involved in the antigenic sites of transmissible gastroenteritis coronavirus S glycoprotein. Virol 183: 225-238. https://doi.org/10.1016/0042-6822(91)90135-X
  12. Godet M, Grosclaude J, Delmas B, Laude H. 1994. Major receptor-binding and neutralization determinants are located within the same domain of the transmissible gastroenteritis coronavirus spike protein. J Virol 68: 8008- 8016. https://doi.org/10.1128/JVI.68.12.8008-8016.1994
  13. Harada K, Kumagai T, Sasahara J. 1967. Studies on transmissible gastroenteritis in pigs. III. Isolation of cytopathogenic virus and its use for serological investigation. Natl Inst Anim Health Quart 7: 127-137.
  14. Hohdatsu T, Eiguchi Y, Ide S, Baba K, Yamagishi H, Kume T, Matumoto M. 1987. Evaluation of an enzyme-linked immunosorbent assay for the detection of transmissible gastroenteritis virus antibodies. Vet Microbiol 13(1): 93-97. https://doi.org/10.1016/0378-1135(87)90103-9
  15. Jang YE, Cho SH, Kim BH, Ahn JM, Kang SY. 1998. Production and characterization of monoclonal antibodies against porcine transmissible gastroenteritis virus. Korean J Vet Res 38(2): 336-344.
  16. Komaniwa H, Makabe T, Fukusho A, Shimizu Y. 1986. Isolation of transmissible gastroenteritis virus from feces of diarrheic pigs in roller culture of CPK cells in the presence of trypsin. Nihon Juigaku Zasshi 48(6): 1245-1248. https://doi.org/10.1292/jvms1939.48.1245
  17. Lanza I, Rubio P, Munoz M, Carmenes P. 1993. Comparison of a monoclonal antibody capture ELISA (MACELISA) to indirect ELISA and virus neutralization test for the serodiagnosis of transmissible gastroenteritis virus. J Vet Diagn Invest 5: 21-25. https://doi.org/10.1177/104063879300500106
  18. Laude H, Rasschaert D, Delmas B, Godet M, Gelfi J, Charleyet B. 1990. Molecular biology of transmissible gastroenteritis virus. Vet Microbiol 23: 147-154. https://doi.org/10.1016/0378-1135(90)90144-K
  19. Nelson LD, Kelling CL. 1984. Enzyme-linked immunosorbent assay for the detection of transmissible gastroenteritis virus antibody in swine sera. Am J Vet Res 45: 1654-1657.
  20. Pensaert M, Callebaut P, Vergote J. 1986. Isolation of a porcine respiratory, non-enteric coronavirus related to transmissible gastroenteritis. Vet Q 8(3):257-261. https://doi.org/10.1080/01652176.1986.9694050
  21. Pritchard GC, Paton DJ, Wibberley G, Ibata G. 1999. "Transmissible gastroenteritis and porcine epidemic diarrhea in Britain." Vet Rec 144(22):616-618. https://doi.org/10.1136/vr.144.22.616
  22. Reynolds DJ. 1982. Application of ELISA to the study of viruses. Development and application of ELISA for veterinary diagnosis and research on enteric and other vises. pp112-123. In: The ELISA: enzyme-linked immunosorbent assay in veterinary research and diagnosis, ed. Wardley RC, Crowther JR. Martinus Nijhoff, The Hague, The Netherlands.
  23. Rukhadze GG, Aliper TI, Sergeev VA. 1989. Isolation of peplomer glycoprotein E2 of transmissible gastroenteritis virus and application in enzyme-linked immunosorbent assay. J Clin Microbiol 27: 1754-1758. https://doi.org/10.1128/JCM.27.8.1754-1758.1989
  24. Saif LJ, Wesley RD. 1999. Transmissible gastroenteritis and Porcine respiratory coronavirus. pp 295-325. In: Straw BE, D'Allaire S, Mengeling WL, Taylor DI(ed.). Diseases of swine. 8th ed. Iowa State University Press, Ames, Iowa.
  25. Siddel S, Wege H, Ter Neulen V. 1983. The Biology of coronaviruses. J Gen Virol 64: 761-776. https://doi.org/10.1099/0022-1317-64-4-761
  26. Stone SS, Kemeny LJ, Jensen MT. 1976. Partial characterization of the principal soluble antigens associated with the coronavirus of transmissible gastroenteritis by complement fixation and immunodiffusion. Infect Immun 13: 521-526. https://doi.org/10.1128/IAI.13.2.521-526.1976
  27. Sune C, Jimenez G, Correa I, Bullido MJ, Gebauer F, Smerdou C, Enjuanes L. 1990. Mechanisms of transmissible gastroenteritis coronavirus neutralization. Virol 177: 559-569. https://doi.org/10.1016/0042-6822(90)90521-R
  28. Tark DS. 1998. Studies on the analysis of the spike gene of porcine transmissible gastroenteritis virus isolated in Korea and the use of recombinant spike protein. Ph. D. Thesis, Chonnam National University, Kwangju, Korea.

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