The use of interleukin-2 receptor expression as a marker of cell-mediated immunity in goats experimentally infected with Mycobacterium avium ssp. paratuberculosis.
The purpose of the present work was to demonstrate cell-mediated immune response to paratuberculosis in experimentally infected animals, using quantification of interleukin-2 receptor (IL-2R) expression on activated lymphocytes by means of in vitro stimulation with Mycobacterium avium ssp. paratuberculosis-derived purified protein derivative (PPDp). A whole-blood technique was developed, and optimal conditions for quantification of IL-2R expression on caprine lymphocytes, using monoclonal antibodies (anti-bovine IL-2R-alpha) and low cytometrical analysis, were determined. Different PPDp-antigen concentrations and incubation times were compared. The whole-blood method was also compared to the more traditional IL-2R assay using peripheral blood mononuclear cultures (Hesketh et al., 1993). Cross-reactivity to Mycobacterium avium was studied at different mycobacteria-PPD concentrations. An immune response could be demonstrated in animals infected with Mycobacterium avium ssp. paratuberculosis. We found that a PPDp concentration of 10microgml(-1) together with an incubation time of 72h, gave the best results using the whole-blood method. The whole-blood method eliminates many laborious steps involved in lymphocyte separation, and the effects of all the constituents of blood are expressed in a way which corresponds more to in vivo conditions. The risk of selecting subpopulations of lymphocytes during cell separation is avoided.
Whist
S K
SK
Institute of Pharmacology, Microbiology and Food Hygiene, The Norwegian School of Veterinary Science, Postbox 8146 Dep., N-0033, Oslo, Norway. solvejg.k.whist@veths.no
Storset
A K
AK
Larsen
H J
HJ
D000818:Animals; D000907:Antibodies, Bacterial / Q000032:analysis; D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial / Q000494:pharmacology; D015415:Biomarkers / Q000032:analysis; D003208:Concanavalin A / Q000494:pharmacology; D005434:Flow Cytometry / Q000662:veterinary; D015511:Goat Diseases / Q000276:immunology*; D006041:Goats; D007111:Immunity, Cellular; D008213:Lymphocyte Activation / Q000187:drug effects; D008297:Male; D009162:Mycobacterium avium / Q000276:immunology; D016927:Mycobacterium avium subsp. paratuberculosis / Q000276:immunology*; D010283:Paratuberculosis / Q000276:immunology*; D015375:Receptors, Interleukin-2 / Q000096:biosynthesis*; D013601:T-Lymphocytes / Q000276:immunology*
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D015415:Biomarkers; D015375:Receptors, Interleukin-2; D003208:Concanavalin A