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

Marvin j. Fritzler* , Fatemeh Behmanesh, Mark L. Fritzler

Deparment of Biochemistry and Molecular Biology, Faculty of Medicine, University of Colgary , 3330 Hospital Dr N.W., Calgary, AB, Canada T2N 4N1

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_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 ELSIA was 75% for chromation , 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 all three isotype rheumatoid factors.

KEYWORDS: Autoantibodies, Diagnostic Kits, Addressable laser bead, immunoassay, Luminex, Rheumatic diseases