The interference of IgM rheumatoid factor in enzyme-linked immunosorbent assays of rubella IgM and IgG antibodies. with or without the addition of the flagellin epitope to the sonicate, we found the most advantageous combination was IC as the source of antibodies and sonicate plus the flagellin epitope as the antigen. In a blinded study of sera obtained from patients with early and later-phase Lyme disease, EMIBA with the enhanced antigenic preparation compared favorably with other serologic assays, especially for the confirmation of early disease. Lyme disease (LD) is usually a multisystem inflammatory disorder (49) due to infection with the spirochete sensu lato, which is usually transmitted by ticks in the complex (5, 50). Soon after the tick bite the pathognomonic skin lesion erythema migrans (EM) occurs in 50 to 75% of patients (49) (and perhaps in as many as 90% of patients [20]), and this is usually often accompanied by nonspecific constitutional symptoms much like those of a viral syndrome. If EM is present, the diagnosis of LD can be made without serologic confirmation. However, in the absence of EM (or if the lesion is not identified properly), the symptoms of early LD are nonspecific and objective indicators of disease may appear only later in the infection. Thus, in the absence of EM the early and prompt diagnosis of early LD may be difficult to make on purely clinical grounds. Due to the great public concern over LD, the number of blood assessments performed exceeds the number of confirmed cases of LD by a factor of 100 (4). False-positive enzyme-linked immunosorbent assay (ELISA) results are common, often resulting in an incorrect diagnosis. It may take 6 to 8 8 weeks for seroconversion to occur, so serologic confirmation of new contamination may be delayed (49). The current recommendation is usually to confirm all positive or equivocal ELISA results by immunoblotting (8a) due to the high frequency of false-positive ELISA results (i.e., a positive ELISA result for a patient without LD). In our experience misinterpretation of the results of Western blot analysis by practioners contributes to the overdiagnosis of LD (45a). Since early antibiotic treatment of LD is more effective in curing the infection and preventing progression to later disease, our challenge was to devise a serologic assay capable of detecting antibodies at the earliest time after contamination while severely limiting false-positive responses. We BAY-598 have focused on detecting immunoglobulin M (IgM) antibodies, which occur during the initial humoral immune response, MAPK3 and in order to optimize the assay, we decided to incorporate the best of a number of different assays. The IgM capture format was selected in order to eliminate potential false-positive results from rheumatoid factor (9, 29, 52) and competition that might otherwise occur in a direct ELISA due to the relatively greater serum IgG concentration (9, 52). Since previous studies have shown that circulating specific anti-IgM antibodies are frequently sequestered within antigen-antibody immune complexes (ICs) (42C44), especially at an early stage of contamination, we chose to compare two sources of serum antibodies: IgM free in serum versus IgM bound in BAY-598 ICs. In order to detect low levels of IgM we had previously chosen biotinylated antigen, much like Hansen et al. (24), and an enzyme-avidin complex (8) to amplify the transmission while preserving a low background (7). This combination of techniques, termed the enzyme-linked IgM capture IC biotinylated antigen assay (EMIBA), was used to test a lender of serum samples obtained from the Lyme Disease Center at the Robert Solid wood Johnson Medical School. It was found to have better sensitivity (98%; 95% confidence interval [CI], 90 to 100%) than IgM ELISA (66%; 95% CI, 53 to 77%) or IgG ELISA (58%; 95% CI, 45 to 70%) and IgM immunoblotting (58%; 95% BAY-598 CI, 45 to 70%) or IgG immunoblotting ELISA.