A cell-based assay for detection of anti-fibrillarin autoantibodies with performance equivalent to immunoprecipitation
A cell-based assay for detection of anti-fibrillarin autoantibodies with performance equivalent to immunoprecipitation
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Anti-fibrillarin autoantibodies are useful for the diagnosis and prognosis of systemic sclerosis (SSc).Anti-fibrillarin produces a clumpy nucleolar pattern in trip in the woods sierra nevada indirect immunofluorescence assay on HEp-2 cells (HEp-2 IFA).Here we develop and validate a reliable cell-based anti-fibrillarin assay (Fibrillarin/CBA) for use in clinical diagnostic laboratories.A TransMembrane Signal was fused to the human fibrillarin gene (TMS-fibrillarin).
HEp-2 cells overexpressing transgenic TMS-fibrillarin at the cytoplasmic membrane were used as IFA substrate in the Fibrillarin/CBA.Sixty-two serum samples with nucleolar pattern in the HEp-2 IFA (41 clumpy; 21 homogeneous/punctate) were tested for anti-fibrillarin using Fibrillarin/CBA, immunoprecipitation (IP), line-blot and ELISA.In addition, samples from 106 SSc-patients were evaluated with Fibrillarin/CBA and the results were correlated with disease phenotypes.Thirty-eight of 41 samples with the clumpy nucleolar pattern (92.
7%) were positive in the Fibrillarin/CBA, while all 21 samples with other nucleolar patterns were negative.Fibrillarin/CBA bullet bill shirt results agreed 100% with IP results.Among the 38 Fibrillarin/CBA-positive samples, only 15 (39.5%) and 11 (29%) were positive for anti-fibrillarin in line-blot and ELISA, respectively.
Higher frequency of diffuse cutaneous SSc (dcSSc) phenotype (72.7% vs 36.8%; p=0.022), cardiac involvement (36.
4% vs 6.5%; p=0.001) and scleroderma renal crisis (18.2% vs 3.
3% p = 0.028) was observed in SSc patients with positive compared to negative Fibrillarin/CBA result.Performance of Fibrillarin/CBA in the detection of anti-fibrillarin autoantibodies was comparable to the gold standard IP.Positive Fibrillarin/CBA results correlated with disease phenotypes known to be associated with anti-fibrillarin autoantibodies, underscoring the clinical validation of this novel assay.