title: Correlation of antigen-specific IFN-γ responses of fresh blood samples from Mycobacterium avium subsp. paratuberculosis infected heifers with responses of day-old samples co-cultured with IL-12 or anti-IL-10 antibodies.
abstract: Paratuberculosis is a chronic infection of the intestine of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). Early stage MAP infection can be detected by measuring cell-mediated immune responses using the interferon gamma (IFN-γ) assay. Whole blood samples are cultured overnight with specific MAP antigens followed by quantification of IFN-γ by ELISA. It is recommended that the time interval from sampling to culture does not exceed eight hours but addition of the co-stimulating cytokine interleukin 12 (IL-12) or anti-IL-10 antibodies to culture have been demonstrated to enhance IFN-γ responses of cultures stimulated with Johnin purified protein derivative (PPDj). Here we examined the correlation of IFN-γ production in response to PPDj and 15 recombinant antigens in day-old blood samples from heifers 10-21 months of age from a MAP infected herd with addition of either recombinant bovine IL-12 or anti-bovine IL-10 antibody with IFN-γ production in sample day samples. IFN-γ responses of sample day samples showed high correlation with responses to some antigens in day-old samples with addition of IL-12 or anti-IL-10 antibodies to cultures, indicating that day-old protocols can be applied as an alternative to the conventional IFN-γ protocol. Immunogenicity of the novel antigens was generally low for day-old samples. The most promising antigen using the day-old protocol with addition of IL-12 was latency protein LATP-2 as correlations, immunogenicity and diagnostic specificity collectively was high. The latency protein LATP-1 was the most promising antigen in the day-old protocol with addition of anti-IL-10 antibodies.
authors.affiliation: National Veterinary Institute, Technical University of Denmark, Bülowsvej 27, 1790 Copenhagen V, Denmark.
authors.forename: Heidi
authors.initials: H
authors.lastname: Mikkelsen
authors.forename: Claus
authors.initials: C
authors.lastname: Aagaard
authors.forename: Søren Saxmose
authors.initials: SS
authors.lastname: Nielsen
authors.forename: Gregers
authors.initials: G
authors.lastname: Jungersen
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology*; D000942:Antigens, Bacterial / Q000276:immunology*; D002417:Cattle; D018920:Coculture Techniques; D007371:Interferon-gamma / Q000096:biosynthesis*; D016753:Interleukin-10 / Q000502:physiology*; D018664:Interleukin-12 / Q000494:pharmacology*; D010283:Paratuberculosis / Q000276:immunology*
chemical_list: D000906:Antibodies; D000942:Antigens, Bacterial; D016753:Interleukin-10; D018664:Interleukin-12; D007371:Interferon-gamma