Analysis of human sera that are polyreactive in an addressable laser bead immunoassay

Abstract

Background: Following the introduction of addressable laser bead immunoassays (ALBIA) into our clinical laboratory, it was noted that certain sera would exhibit reactivity to numerous antigens in the array. To further understand the nature of this reactivity, we analyzed the reactivity of sequential sera that were identified over a 1 year period. Methods: Sera that demonstrated reactivity to 6 or more of the 8 antigens in an ALBIA kit (QuantaPlex 8: chromatin, Sm, RNP, Scl-70, ribosomal P protein, SS-A/Ro, SS-B/La, Jo-1) were tested for autoantibodies by indirect immunofluorescence (IIF) on HEp-2 cell substrates, for IgG, IgM and IgA rheumatoid factor, chromatin and ribosomal P protein by ELISA and by LINE immunoassay (LIA) and immunoblotting (IB). Results: In one calendar year, 40/4096 (0.8%) sera analyzed in a routine clinical laboratory setting demonstrated reactivity to 6 or more antigens in the QuantaPlex 8 kits. There was no common IIF pattern that could be attributed to the polyreactive sera. There was no apparent correlation of polyreactivity with IIF titers, indeed, 4/40 (10%) sera had a negative ANA at the screening dilution of 1/160. When subjected to IB, LIA and ELISA, polyreactivity to three or more antigens was confirmed for 12/40 (30%) of sera while 8/40 (20%) had reactivity to 1–2 antigens and 20 (50%) did not react with any antigens in these assays. Overall agreement of positive or negative tests between the ALBIA and IB, LIA and ELISA was 75% for chromatin, 50% for SS-A, 27.5% for Sm, 25% for Rib-P, 22.5% for RNP, 20% for Scl-70, 15% for Jo-1 and 7.5% for SS-B. 17/40 (42.5%) had a positive IgM, IgG or IgA rheumatoid factor, and 12/40 (30%) had all three isotype rheumatoid factors. Conclusions: On average, the agreement between ALBIA and other assays in this study of polyreactive sera was 30%. Approximately, one-half of sera that demonstrate reactivity to multiple autoantigens in a commercial ALBIA were confirmed to have reactivity to at least one autoantigen in another diagnostic assay and 30% could be regarded as polyreactive. Other sera, some of which had rheumatoid factor, appeared to have high background binding without demonstrating specific binding to any of the cognate antigens.