Studies on the Sm and RNP nuclear antigens of human autoimmune rheumatic and connective-tissue diseases

1981 ◽  
Vol 9 (1) ◽  
pp. 144-145
Author(s):  
SOHRAB DAHI ◽  
GEORGE NAXAKIS ◽  
CHRISTINE M. WALLIS ◽  
ALEXANDER J. MacGILLIVRAY
Keyword(s):  
Connective Tissue ◽  
Connective Tissue Diseases ◽  
Nuclear Antigens

scholarly journals Detection of Specific Antinuclear Reactivities in Patients with Negative Anti-nuclear Antibody Immunofluorescence Screening Tests

2002 ◽  
Vol 48 (12) ◽  
pp. 2171-2176 ◽  
Author(s):  
Ilse EA Hoffman ◽  
Isabelle Peene ◽  
Eric M Veys ◽  
Filip De Keyser
Keyword(s):  
Connective Tissue ◽  
Confidence Interval ◽  
Connective Tissue Disease ◽  
Indirect Immunofluorescence ◽  
Connective Tissue Diseases ◽  
Clinical Suspicion ◽  
Screening Tests ◽  
Strongly Correlated ◽  
Nuclear Antigens

Abstract Background: For detection of anti-nuclear antibodies (ANAs) and antibodies to extractable nuclear antigens (ENAs), samples frequently are screened with indirect immunofluorescence (IIF); further determination of anti-ENA antibodies is performed only when the result is positive. However, because anti-ENA reactivities are found in samples with low fluorescence intensities, we determined anti-ENA antibodies in samples with negative IIF and thus calculated the sensitivity of IIF for specific ANAs. Methods: We collected 494 samples consecutively referred by rheumatologists for routine ANA testing. IIF on HEp-2 and HEp-2000 (HEp-2 cells transfected with Ro60 cDNA) and line immunoassay (LIA) for the detection of specific ANAs were performed on all samples. Results: Fluorescence intensities and patterns on HEp-2 were strongly correlated with those on HEp-2000 [Spearman ρ = 0.852 (P <0.001) and 0.838 (P <0.001), respectively]. Sixty-eight of 494 samples were positive on LIA, of which only 72% (confidence interval, 68–76%) were detected with HEp-2 and 75% (confidence interval, 70–78%) with HEp-2000. Of 291 samples negative on both substrates, 12 were positive on LIA. Connective tissue diseases were diagnosed in four of these patients and suspected in at least three others. Conclusion: The HEp-2 and HEp-2000 substrates perform comparably for fluorescence intensities and patterns and for detecting specific ANAs, but some patients with negative IIF show reactivity on LIA. We recommend testing for fine reactivities, regardless of the IIF result, when the clinical suspicion for rheumatic connective tissue disease is high.

Staining of antinuclear antibodies and antibodies against removable nuclear antigens in connective tissue diseases

2020 ◽  
Vol 48 (1) ◽  
pp. 18-25
Author(s):  
José Enrique Oliva Menacho ◽  
Jorge Luis Arroyo-Acevedo ◽  
José Arturo Oliva-Candela ◽  
Marco Antonio García-Hjarles ◽  
Lester Domínguez-Huarcaya
Keyword(s):  
Connective Tissue ◽  
Antinuclear Antibodies ◽  
Connective Tissue Diseases ◽  
Nuclear Antigens

scholarly journals Development and validation of a novel counter-immunoelectrophoresis assay for the detection of antibodies against extractable nuclear antigens

2021 ◽  
Author(s):  
◽  
Reuben Wallis
Keyword(s):  
Connective Tissue ◽  
Likelihood Ratio ◽  
Connective Tissue Diseases ◽  
Positive Likelihood Ratio ◽  
Negative Likelihood Ratio ◽  
Rater Agreement ◽  
Diagnostic Specificity ◽  
Nuclear Antigens ◽  
Sera Samples ◽  
Assay Conditions

<p>The identification of autoantibodies is a primary diagnostic marker for the diagnosis of some autoimmune diseases. The sera of patients with connective tissue diseases commonly contain autoantibodies that target nuclear antigens. As such these antibodies are called antinuclear antibodies (ANAs). Furthermore, the clinical identification of ANAs in patient sera to specific nuclear antigens is a primary tool for the diagnosis of connective tissue diseases. For this purpose specific extractable nuclear antigens (ENAs) are used. The most commonly employed laboratory technique for the detection of ENA antibodies is the enzyme linked immunoabsorbant assay (ELISA). ELISA is a simple technique that can be automated and provides high diagnostic sensitivity for the detection of ENA antibodies comparatively to a counter immunoelectrophoresis (CIE) assay. However, compared to CIE ELISA lacks diagnostic specificity. Therefore there is a demand for an assay with high diagnostic specificity as a secondary diagnostic test for the detection of ENA antibodies. The central aim of this project was to develop and validate a novel CIE assay that used fluorescently labelled ENAs to detect ENA antibodies in patient serum.   Development of the assay began with comparative testing of fluorescently labelled and unlabelled antigens to confirm that the immunochemical properties of the antigen remained intact. The CIE assay conditions were optimised for the detection of four ANAs SSA, SSB, RNP/Sm, and Sm using their corresponding ENAs. Assay conditions optimised were flurophore type, gel composition, buffer composition, antigen concentration, the running time, and analytical specificity. Evaluation of 281 clinical sera samples known to have previously tested positive test by ANA indirect immunofluorescence were performed using the optimised CIE assay and ELISA. The inter-rater agreement between the CIE assay and ELISA results was determined. Clinical details were used to retrospectively classify the clinical sera samples into clinical categories and case groups. This data was used for the diagnostic comparison of the CIE assay and ELISA results.  There was strong inter-rater agreement between the SSA CIE assay and ELISA results. The diagnostic specificity and positive likelihood ratio values of the SSA CIE assay were superior to those of the ELISA, while diagnostic sensitivity and the negative likelihood ratio values were approximately the same. There was strong inter-rater agreement between the RNP/Sm CIE assay and ELISA. However, it was observed that the statistical measures of assay accuracy (diagnostic specificity and sensitivity, positive likelihood ratio, and negative likelihood ratio) for the RNP/Sm ELISA were superior to those of CIE assay. The diagnostic specificity of the SSB and Sm CIE assays were higher than that of their respective ELISAs. This was also true for the positive likelihood ratio values of the SSB and Sm CIE assays when compared to their respect ELISAs.  The SSA CIE assay results suggest that this assay maybe a suitable replacement for ELISA as a diagnostic tool for the identification of anti-SSA antibodies and Sjogren’s syndrome. The SSB and Sm CIE assay results suggest that these assays would be suitable as secondary confirmatory diagnostic assays for the identification of their respective ENA antibodies. However, further performance evaluation of the assays within a routine diagnostic laboratory is required.</p>

scholarly journals Heterogeneous Nuclear Ribonucleoprotein H1, a Novel Nuclear Autoantigen

2009 ◽  
Vol 55 (5) ◽  
pp. 946-954 ◽  
Author(s):  
Karolien Van den Bergh ◽  
Herbert Hooijkaas ◽  
Daniel Blockmans ◽  
René Westhovens ◽  
Katrijn Op De Beéck ◽  
...  
Keyword(s):  
Connective Tissue ◽  
Clinical Laboratory ◽  
Connective Tissue Diseases ◽  
Nuclear Target ◽  
Serum Samples ◽  
Heterogeneous Nuclear Ribonucleoprotein ◽  
Proteomic Approach ◽  
Nuclear Antigens ◽  
Diagnostic Sensitivity And Specificity ◽  
Nuclear Ribonucleoprotein

Abstract Background: Serum samples from patients with autoimmune connective tissue diseases that show a finely speckled antinuclear antibody (ANA) on indirect immune-fluorescence often have antibodies against unknown nuclear target antigens. To search for such autoantigens we applied a proteomic approach using sera from patients with a high ANA titer (≥640) and finely speckled fluorescence but in whom no antibodies to extractable nuclear antigens (ENA) could be identified. Methods: Using an immunoproteomics approach we identified heterogeneous nuclear ribonucleoprotein H1 (hnRNP H1) as a novel nuclear target of autoantibody response. Results: Recombinant rat hnRNP H1 reacted in Western blot analyses with 48% of 93 sera from patients with primary Sjögren syndrome and with 5.2% of 153 sera from patients with other connective tissue diseases (diseased controls). For comparison, the diagnostic sensitivity and specificity of anti–Sjögren syndrome A (SSA) antibodies for primary Sjögren syndrome in the same patient cohort were 88.2% and 76.3%, respectively. Interestingly, 5 of 11 primary Sjögren syndrome patients with no anti-SSA or anti-SSB antibodies had anti–hnRNP H1 antibodies. Anti–hnRNP H1 antibodies were preabsorbed by hnRNP H1, as demonstrated by indirect immunofluorescence. In an evaluation of the presence of anti–hnRNP H1 antibodies in 188 consecutive samples submitted to the clinical laboratory with positive ANA (titer ≥160), anti–hnRNP H1 antibodies were found in 3 of 7 (2 primary and 5 secondary) Sjögren syndrome patients and in 8.3% of the diseased controls. Conclusions: HnRNP H1 is a newly discovered autoantigen that could become an additional diagnostic marker.

scholarly journals Development and validation of a novel counter-immunoelectrophoresis assay for the detection of antibodies against extractable nuclear antigens

2021 ◽  
Author(s):  
◽  
Reuben Wallis
Keyword(s):  
Connective Tissue ◽  
Likelihood Ratio ◽  
Connective Tissue Diseases ◽  
Positive Likelihood Ratio ◽  
Negative Likelihood Ratio ◽  
Rater Agreement ◽  
Diagnostic Specificity ◽  
Nuclear Antigens ◽  
Sera Samples ◽  
Assay Conditions

<p>The identification of autoantibodies is a primary diagnostic marker for the diagnosis of some autoimmune diseases. The sera of patients with connective tissue diseases commonly contain autoantibodies that target nuclear antigens. As such these antibodies are called antinuclear antibodies (ANAs). Furthermore, the clinical identification of ANAs in patient sera to specific nuclear antigens is a primary tool for the diagnosis of connective tissue diseases. For this purpose specific extractable nuclear antigens (ENAs) are used. The most commonly employed laboratory technique for the detection of ENA antibodies is the enzyme linked immunoabsorbant assay (ELISA). ELISA is a simple technique that can be automated and provides high diagnostic sensitivity for the detection of ENA antibodies comparatively to a counter immunoelectrophoresis (CIE) assay. However, compared to CIE ELISA lacks diagnostic specificity. Therefore there is a demand for an assay with high diagnostic specificity as a secondary diagnostic test for the detection of ENA antibodies. The central aim of this project was to develop and validate a novel CIE assay that used fluorescently labelled ENAs to detect ENA antibodies in patient serum.   Development of the assay began with comparative testing of fluorescently labelled and unlabelled antigens to confirm that the immunochemical properties of the antigen remained intact. The CIE assay conditions were optimised for the detection of four ANAs SSA, SSB, RNP/Sm, and Sm using their corresponding ENAs. Assay conditions optimised were flurophore type, gel composition, buffer composition, antigen concentration, the running time, and analytical specificity. Evaluation of 281 clinical sera samples known to have previously tested positive test by ANA indirect immunofluorescence were performed using the optimised CIE assay and ELISA. The inter-rater agreement between the CIE assay and ELISA results was determined. Clinical details were used to retrospectively classify the clinical sera samples into clinical categories and case groups. This data was used for the diagnostic comparison of the CIE assay and ELISA results.  There was strong inter-rater agreement between the SSA CIE assay and ELISA results. The diagnostic specificity and positive likelihood ratio values of the SSA CIE assay were superior to those of the ELISA, while diagnostic sensitivity and the negative likelihood ratio values were approximately the same. There was strong inter-rater agreement between the RNP/Sm CIE assay and ELISA. However, it was observed that the statistical measures of assay accuracy (diagnostic specificity and sensitivity, positive likelihood ratio, and negative likelihood ratio) for the RNP/Sm ELISA were superior to those of CIE assay. The diagnostic specificity of the SSB and Sm CIE assays were higher than that of their respective ELISAs. This was also true for the positive likelihood ratio values of the SSB and Sm CIE assays when compared to their respect ELISAs.  The SSA CIE assay results suggest that this assay maybe a suitable replacement for ELISA as a diagnostic tool for the identification of anti-SSA antibodies and Sjogren’s syndrome. The SSB and Sm CIE assay results suggest that these assays would be suitable as secondary confirmatory diagnostic assays for the identification of their respective ENA antibodies. However, further performance evaluation of the assays within a routine diagnostic laboratory is required.</p>

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