Journal of Biosciences

, Volume 25, Issue 1, pp 47–54 | Cite as

An enzyme immunoassay for detection of Japanese encephalitis virus-induced chemotactic cytokine



Japanese encephalitis virus (JEV) induces human peripheral blood monocytes to secrete a chemotactic cytokine [human macrophage-derived factor (hMDF)] which causes chemotaxis of neutrophils. The only known assay for hMDF cannot quantify its level in samples, so an enzyme immunoassay has been standardized for detection of hMDF and hMDF-specific antibodies in test samples. The reported enzyme linked immunosorbent assay (ELISA) was found to be sensitive (89%), specific (91%), accurate (92.2%) and reproducible and was able to detect a minimum concentration of 23 ng hMDF/ml in test samples. The chemotactic factor could be detected in JEV inoculated mouse sera and JEV infected culture fluids. Significant finding of the test was the detection of hMDF in sera of human cases of JE.


Enzyme immunoassay human macrophage-derived factor Japanese encephalitis virus 

Abbreviations used


Enzyme linked immunosorbent assay


Japanese encephalitis virus


human macrophage-derived factor


Hank’s balanced salt solution


cerebrospinal fluid


minimum essential medium


foetal calf serum


phosphate buffered saline


normal saline




horse raddish peroxiidase


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  1. Adams D H and Lloyd A R 1997 Chemokines: leukocyte recruitment and activation cytokines;Lancet 349 490–502PubMedCrossRefGoogle Scholar
  2. Appassakij H, Bunchuin N, Sarasombath S, Rangpitarangsi B, Manatsathit S, Komolpit P and Sukosol T 1987 Enzyme-linked immunosorbent assay for detection ofSalmonella typhi protein antigen;J. Clin. Microbiol. 25 273–277PubMedGoogle Scholar
  3. Baggiolini M 1998 Chemokines and leucocyte traffic;Nature (London) 392 565–568CrossRefGoogle Scholar
  4. Beech J T, Bainbridge T and Thompson S J 1997 Incorporation of cells into an ELISA system enhances antigen-driven lymphokine detection;J. Immunol. Methods 205 163–168PubMedCrossRefGoogle Scholar
  5. Benveniste E N 1992 Inflammatory cytokines within the central nervous system: sources, function, mechanism and action;Am. J. Physiol. 263 C1-C6PubMedGoogle Scholar
  6. Chaturvedi U C, Mathur A, Tandon P, Natu S M, Rajvanshi S and Tandon H O 1979 Variable effect on peripheral blood leukocytes during JE virus infection of man;Clin. Exp. Immunol. 38 492–498PubMedGoogle Scholar
  7. de Gruchy G C 1976Clinical hematology in medical practice 3rd edition (Oxford: ELBS and Blackwell Scientific Publications) pp 359–403Google Scholar
  8. Gillis S and Williams D E 1998 Cytokine therapy: lessons learned and future challenges;Curr. Opinion Immunol. 10 501–503CrossRefGoogle Scholar
  9. Ida N, Sakurai S, Hosoi K and Kunitomo T 1992 A highly sensitive enzyme-linked immunosorbent assay for the measurement of interleukin-8 in biological fluids;J. Immunol. Methods 156 27–38PubMedCrossRefGoogle Scholar
  10. Ishiguro A, Suzuki Y, Inaba Y, Fukushina K, Komiyama A, Koeffler F and Shimbo T 1997 The production of IL-8 in cerebrospinal fluid in aseptic meningitis of children;Clin. Exp. Immunol. 109 426–430PubMedCrossRefGoogle Scholar
  11. Johnson R T, Burke D S, Elwell M, Leake C J, Nisalak A, Hoke C H and Lorsomrudee W 1985 Japanese encephalitis: Immunocytochemical studies of viral antigen and inflammatory cells in fatal cases;Ann. Neurol. 18 567–573PubMedCrossRefGoogle Scholar
  12. Jung T, Bews J P A, Enssle K H, Wagner K, Neumann C and Heusser C H 1998 Detection and discrimination between total and free human interleukin-4 and free soluble interleukin-4 receptor by ELISA;J. Immunol. Methods 217 41–50PubMedCrossRefGoogle Scholar
  13. Khanna N, Agnihotri M, Mathur A and Chaturvedi U C 1991 Neutrophil chemotactic factor produced by Japanese encephalitis virus-stimulated macrophages;Clin. Exp. Immunol. 86 299–303PubMedCrossRefGoogle Scholar
  14. Khanna N, Mathur A and Chaturvedi U C 1994 Regulation of vascular permeability by macrophage derived chemotactic factor produced in Japanese encephalitis;Immunol. Cell Biol. 72 200–204PubMedCrossRefGoogle Scholar
  15. Khanna N, Mathur A, Bharadwaj M and Chaturvedi U C 1997 Induction of hypoglycaemia in Japanese encephalitis virus infection: the role of T-lymphocytes;Clin. Exp. Immunol. 107 282–287PubMedCrossRefGoogle Scholar
  16. Khanna N, Srivastava S, Mathur A and Chaturvedi U C 1993 Stimulation of neutrophil respiratory burst and degranulation by Japanese encephalitis virus-induced macrophage derived factor;Int. J. Exp. Pathol. 74 339–345PubMedGoogle Scholar
  17. Ledur A, Fitting C, David B, Hamberger C and Cavaillon J M 1995 Variable estimates of cytokine level produced by commercial ELISA kits: results using international cytokine standards;J. Immunol. Methods 186 171–179PubMedCrossRefGoogle Scholar
  18. Liles W C and Van Voorhis W C 1995 Nomenclature and biologic significance of cytokines involved in inflammation and the host immune response;J. Infect. Dis. 172 1573–1580PubMedGoogle Scholar
  19. Lopez-Cortes L F, Cruz-Ruiz M, Gomez-Mateos J, Vicianna-Fernandez P, Martinez-Marcos F J and Panchon J 1995 Interleukin-8 in cerebrospinal fluid from patients with meningitis of different etiologies: Its possible role as neutrophil chemotactic factor;J. Infect. Dis. 172 581–584PubMedGoogle Scholar
  20. Lowry O H, Rosebrough N J, Farr A L and Randall R J 1951 Protein measurements with the Folin-phenol reagent;J. Biol. Chem. 193 265–275PubMedGoogle Scholar
  21. Mantovani A 1999 The chemokine system: redundancy for robust outputs;Immunol. Today 20 254–257PubMedCrossRefGoogle Scholar
  22. Mathur A, Bhardwaj M, Kulshreshtha R, Rawat S, Jain A and Chaturvedi U C 1988 Immunopathological study of spleen during JEV infection in mice;Br. J. Exp. Pathol. 69 423–432PubMedGoogle Scholar
  23. Mathur A, Chaturvedi U C, Tandon H O, Agarwal A K, Mathur G P, Nag D, Prasad A and Mittal V P 1982 Japanese encephalitis in Uttar Pradesh, India during 1978;Inidan J. Med. Res. 75 161–169Google Scholar
  24. Mathur A, Khanna N and Chaturvedi U C 1992 Breakdown of blood-brain barrier by virus induced cytokine during Japanese encephalitis virus infection;Int. J. Exp. Pathol. 73 603–611PubMedGoogle Scholar
  25. Mathur A, Kumar R, Sharma S, Kulshreshtha R, Kumar A and Chaturvedi U C 1990 Rapid diagnosis of Japanese encephalitis by immunofluorescent examination of cerebrospinal fluid;Indian J. Med. Res. A91 1–4Google Scholar
  26. Sekido N, Mukaida N, Harada A, Nakanishi I, Watanabe Y and Matsumisha K 1993 Prevention of lung perfusion injury in rabbits by a monoclonal antibody against interleukin-8;Nature (London) 365 654–657CrossRefGoogle Scholar
  27. Sharma S, Mathur A, Prakash, V, Kulshreshtha R, Kumar R and Chaturvedi U C 1991 JEV latency in peripheral blood lymphocytes and recurrence of infection in children;Clin. Exp. Immunol. 85 85–89PubMedGoogle Scholar
  28. Srivastava S, Khanna N, Saxena S K, Singh A, Mathur A and Dhole T N 1999 Degradation of Japanese Encephalitis virus by neutrophils;Int. J. Exp. Pathol. 80 17–24PubMedCrossRefGoogle Scholar
  29. Tsang MLS and Weatherbee J A 1996 Cytokine assays and their limitations;Aliment. Pharmacol. Ther. Suppl. 10 55–61Google Scholar
  30. Voller A, Bidwell E and Bartle A 1976The enzyme linked immunosorbent assay (ELISA) (Alexandria Dynatech Laboratories) 9–33Google Scholar

Copyright information

© Indian Academy of Sciences 2000

Authors and Affiliations

  1. 1.Postgraduate Department of MicrobiologyKG’s Medical CollegeLucknowIndia

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