RAPID AND SENSITIVE DETECTION OF ANTI-RO (SS-A) ANTIBODIES BY INDIRECT IMMUNOFLUORESCENCE OF 60KDA RO HEP-2 TRANSFECTANTS

Anti-Re (SS-A) antibodies are important diagnostic markers for primary Sjogren's syndrome and systemic lupus erythematosus, but their detect ion by indirect immunofluorescence (IF) in the diagnostic laboratory i s hindered by the low cellular abundance of 60kDa Ro protein (Ro60). T he approach we used to overcome this problem was to transfect and over -express the Ro60 gene into HEp-2 cells. In this study we have used a mixture of Ro60 transfectants and untransfected HEp-2 cells (HEp-Ro60) as a substrate for If-antinuclear antibody (ANA) testing in a hospita l laboratory. Screening of 240 routine serum specimens identified 14 R o transfectant-positive sera which were confirmed by counterimmunoelec trophoresis (CIE); 3 of these sera were ANA-negative on untransfected cells and regular HEp-2. A comparison of HEp-Re60 and regular HEp-2 sh owed strong concordance of the different ANA patterns between the 2 su bstrates. No increase in background staining was observed on the Ro tr ansfectants when reacted with normal human sera. A comparison between HEp-Ro60 and CIE for 53 sera from patients with primary Sjogren's synd rome showed that HEp-Ro60 were a sensitive and specific substrate for detection of anti-Ro antibodies. Masking of positive Ro transfectants was observed rarely in sera containing multiple ANA specificities, but the Ro60 staining on these transfectants was unmasked at higher serum dilutions. We conclude that HEp-Ro60 are a suitable substrate for IF- ANA in the routine laboratory and that they have the additional advant age over regular HEp-2 slides of being able to detect anti-Ro in ANA-n egative sera. HEp-RO60 are also a valuable confirmatory test for sera giving equivocal precipitin reactions or ELISA results.