Clinical optimization and multicenter validation of antigen-specific cut-off values on the INNO-LIAr ANA Update for the detection of autoantibodies in connective tissue disorders

Objectives

Optimization and validation of antigen-specific cut-off values for SmB, SmD, RNP-70k, RNP-A, RNP-C, SSA/Ro52, SSA/Ro60, SSB/La, Cenp-B, Topo-I, Jo-1, ribosomalP and histones to achieve 98% specificity for each of the markers.

Patients and methods

The INNO-LIAr ANA Update is a qualitative test detecting antibodies to several different antigens, most of which are recombinantly made, with the exception of SSA/Ro60 and histones (natural), and SmD and ribosomal P (synthetic). The LIA-SCAN ANA provides a quantitative read-out of the INNO-LIAr ANA Update results. The cut-off value of the different antigen lines was optimized using an in-house set of 955 samples. The assay specificity was validated at multiple sites using a different set of 330 samples obtained from 158 apparently healthy blood donors, 100 patients with a variety of infections, 20 with Wegener's granulomatosis, 20 with inflammatory bowel disease, 20 with primary antiphospholipid syndrome, and 12 with psoriatic arthritis. The INNO-LIAr ANA Update reactivity using the optimized cut-off was tested in 147 patients with scleroderma, 93 with Sjögren's disease, 40 patients with systemic lupus erythematosus (SLE), 40 with rheumatoid arthritis (RA), 39 with mixed connective tissue disease, and 19 with myositis. The clinical diagnosis was considered as the gold standard.

Results and Conclusions

The optimized cut-off values resulted in an average specificity of over 98% for all LIA markers in the validation set of 330 samples. The pattern of reactivity for the different LIA ANAs in the 378 samples from the target patient groups corresponded to the sensitivities reported in the literature. In conclusion, the INNO-LIAr ANA Update shows uniformly high specificities combined with sensitivities very similar to those of reference assays, in a single test format.