Evaluation of a Novel Line‐Blot Immunoassay for the Detection of Antibodies to Extractable Nuclear Antigens

Abstract

We have evaluated the performance of a novel line-blot immunoassay (LIA; Mikrogen) and compared results with those obtained by CIE (in-house), ELISA (Pharmacia Diagnostics), and FEIA (Pharmacia Diagnostics). Sera from systemic lupus erythematosus (SLE) patients (n = 123), systemic sclerosis patients (n = 25), and healthy controls (n = 40) were analyzed for the presence of antibodies to RNP, Sm, SSA, SSB, CENP-B, Scl-70, and Jo-1. Reading of LIA results, as compared with a cutoff control, was performed by automatic analysis of the test strips. Because LIA enables recognition of separate subunits of RNP (68, A, and C), Sm (B and D), and SSA (52 and 60), at least two of the RNP antigens and either one of the Sm or SSA antigens should be detected for considering the test RNP, Sm, or SSA-positive, respectively. LIA had the highest sensitivity in patients with autoimmune connective tissue diseases: 131 specificities (not PO, PCNA, or histones), as compared with ELISA (121), FEIA (119), and CIE (80). However, LIA revealed three positive reactions in healthy controls; other assays were completely negative. LIA is better than CIE, but similar to ELISA and FEIA, in terms of detecting systemic sclerosis-associated antibodies (CENP-B and Scl-70). Furthermore, LIA had the highest sensitivity (17.9%) for the SLE-specific anti-Sm antibodies, as compared with ELISA (11.4%), CIE (8.1%), and FEIA (5.7%). Finally, anti-SSA antibodies were far more prevalent by LIA in the systemic sclerosis samples because of anti-SSA52 reactivity. The clinical relevance of the latter finding remains to be determined. In conclusion, LIA is suitable for routine evaluation of autoantibodies to extractable nuclear antigens.

Isolated atrioventricular block of unknown origin in the adult and autoimmunity: diagnostic and therapeutic considerations exemplified by 3 anti-Ro/SSA–associated cases

“…Further support of this view, also substantiated by previous data demonstrating a higher sensibility of Western blot in detecting anti-Ro/SSA-52kD with respect to immunoenzymatic methods, 14 another case of anti-Ro/SSA-52kD–positive (blot only) III°AVB fully reversing after immunosuppressive therapy(acquired form) has been recently described 10 …”

Update on autoantibodies in systemic sclerosis

“…Anti-CENPs have been typically detected by IIF, but more recently ELISA [28] and line immunoassays have been used [44,45]. [43] retrospectively reviewed 120 anti-CENP-B positive patients whose sera were screened by IIF and then anti-CENP-B detected by ELISA.…”

Borrelia burgdorferiComplement Regulator-Acquiring Surface Protein 1 of the Lyme Disease Spirochetes Is Expressed in Humans and Induces Antibody Responses Restricted to Nondenatured Structural Determinants

“…Similar results were obtained when recombinant CRASP-1 was subjected without boiling and reduction to SDS-PAGE (data not shown), indicating that SDS treatment might be the critical factor responsible for the loss of the immunogenic epitopes of CRASP-1. In order to reveal whether sera from Lyme disease patients contain antibodies with specificities to nondenatured determinants of CRASP-1, we have used two appropriate test systems, i.e., a line immunoassay (7,17) and an ELISA, in which nondenatured forms of recombinant Borrelia proteins serve as target molecules. The integrity of the three recombinant spirochetal proteins used, i.e., CRASP-1, VlsE, and OspC, was verified by Coomassie staining (Fig.…”