Sensitive and specific immunoassays to detect rabbit IL-8 and MCP-1: cytokines that mediate leukocyte recruitment to the lungs.
The alpha- and beta-chemokines such as IL-8 and MCP-1 direct the recruitment of neutrophils and monocytes into the lungs and other tissues. In order to study the roles of IL-8 and MCP-1 in animals models, specific reagents are required that provide accurate measurements of these cytokines in biological fluids. Here we describe the development of sensitive and specific immunoassays for rabbit IL-8 and rabbit MCP-1, and the validation of these assays in rabbit plasma and bronchoalveolar lavage fluid. The sensitivity of each assay in 0.25 ng/ml for IL-8 and 0.1 ng/ml for MCP-1. The rabbit IL-8 assay does not crossreact with rabbit GRO, another alpha-chemokine, and crossreacts only weakly with human IL-8. The rabbit MCP-1 assay does not crossreact with human MCP-1. Anticoagulants interfere with the detection of IL-8 and MCP-1 in plasma, although. EDTA has the least inhibitory effect. Heat-sensitive inhibitors in normal rabbit serum interfere with the detection of IL-8 and MCP-1, although autoantibodies to IL-8 and MCP-1 were not detected. Rabbit erythrocytes bind IL-8 and MCP-1, but erythrocyte contamination of bronchoalveolar lavage fluid causes only a small error in the detection of IL-8 and MCP-1, unless the number of erythrocytes approaches the number found in blood. These assays provide sensitive and specific means to detect IL-8 and MCP-1 in rabbit plasma and bronchoalveolar lavage fluid, and demonstrate the importance of using species-specific reagents in animal studies.
Medical Research Service, Seattle Veterans' Affairs Medical Center, WA 98108, USA.
O
O
Kajikawa
R B
RB
Goodman
M C
MC
Johnson
K
K
Konishi
T R
TR
Martin
D000818:Animals; D001323:Autoantibodies / Q000032:analysis; D001992:Bronchoalveolar Lavage Fluid / Q000737:chemistry; D018932:Chemokine CCL2 / Q000032:analysis* / Q000276:immunology; D005260:Female; D006358:Hot Temperature; D007118:Immunoassay / Q000379:methods; D007249:Inflammation / Q000503:physiopathology; D016209:Interleukin-8 / Q000032:analysis* / Q000276:immunology; D016676:Macrophages, Alveolar / Q000737:chemistry; D011817:Rabbits; D011994:Recombinant Proteins
D001323:Autoantibodies; D018932:Chemokine CCL2; D016209:Interleukin-8; D011994:Recombinant Proteins