Clinical and serological evaluation of a novel CENP-A peptide based ELISA

Mähler, Michael; Maes, Liesbeth; Blockmans, Daniel Engelbert; Westhovens, René; Bossuyt, Xavier; Riemekasten, Gabriela; Schneider, Sandra; Hiepe, Falk; Swart, Andreas; Gürtler, Irmgard; Egerer, Karl; Fooke, Margaret; Fritzler, Marvin J.

doi:10.1186/ar3029

Cited by 25 publications

(18 citation statements)

References 40 publications

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“…Altogether, these results strongly suggest that CENP-A Abs are generated independently from anti-CENP-B Abs and that the previously reported motif G/APXRX is not responsible for any cross-reactivity between anti-CENP-A and -CENP-B Abs [18], [32]. Supporting this conclusion are the results by Mahler et al [33], who reported a lack of cross-inhibition between anti-CENP-A and anti-CENP-B Abs, using an amino terminal CENP-A-derived peptide (sequence not reported) and recombinant CENP-B as inhibitor.…”

Section: Discussionsupporting

confidence: 77%

Autoantibodies Recognizing the Amino Terminal 1-17 Segment of CENP-A Display Unique Specificities in Systemic Sclerosis

et al. 2013

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Centromere-associated protein A (CENP-A), a common autoimmune target in a subset of systemic sclerosis patients, appears to have no role to explain why its corresponding auto-antibodies are more frequently found in the limited than the diffuse form of systemic sclerosis. Therefore, we investigated the fine specificity of anti-CENP-A antibodies as a first step to understanding their role in systemic sclerosis pathology. We focused on the amino-terminal portion of CENP-A spanning amino acids 1 to 17 (Ap1-17), which represents, along with Ap17-30, an immunodominant epitope of the protein. Peptide Ap1-17 was used to purify antibodies from 8 patients with systemic sclerosis. Anti-Ap1-17 antibodies specifically reacted with human CENP-A but did not cross-react with CENP-B or Ap17-30. Panning of a phage display peptide library with anti-Ap1-17 antibodies from 2 patients identified two novel, partially overlapping motifs, <5Rx(st)xKP10> and <9KPxxPxR15> as the result of the alignment of specific phage clone insert sequences. Anti-Ap1-17 IgG from the 8 patients had different reactivities to isolated phage clone insert sequences. Scanning the Swiss-Prot database revealed a large number of different types of proteins containing the two Ap1-17 antigenic motifs. These data show that anti-CENP-A1-17 antibodies are generated independently from anti-CENP-B antibodies and display great heterogeneity in their specificity by recognizing different motifs within that peptide sequence. This finding, along with the widespread interspecies and human tissue distribution of the two motifs, suggests that the number of motif-expressing proteins which can be the potential target of these antibodies is markedly higher than that estimated from the peptide-based epitope spreading model.

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Section: Discussionsupporting

confidence: 77%

Autoantibodies Recognizing the Amino Terminal 1-17 Segment of CENP-A Display Unique Specificities in Systemic Sclerosis

et al. 2013

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“…In a previous study dcSSc and lcSSc subsets were associated with particular organ manifestations, but in this analysis the clinical distinction appeared superseded by an antibody-based classification in predicting some SSc-related complications [33]. In addition, the discrimination between SSc and controls as measured by the AUC (0.73) derived from ROC analysis using anti-Th/To (Rpp25) antibody results was similar to the AUC (0.67) generated by ACA [34]. …”

Section: Discussionsupporting

confidence: 54%

Rpp25 is a major target of autoantibodies to the Th/To complex as measured by a novel chemiluminescent assay

et al. 2013

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IntroductionAutoantibodies to the Th/To antigen have been described in systemic sclerosis (SSc) and several proteins of the macromolecular Th/To complex have been reported to react with anti-Th/To antibodies. However, anti-Th/To has not been clinically utilized due to unavailability of commercial tests. The objective of the present study is to evaluate the newly developed ELISA and chemiluminescent immunoassay (CLIA) to measure autoantibodies to Rpp25 (a component of the Th/To complex) using immunoprecipitation (IP) as the reference method.MethodsThe first cohort consisted of 123 SSc patients including 7 anti-Th/To positive samples confirmed by IP. Additional seven anti-Th/To positive samples from non-SSc patients were also tested. For evaluation of the QUANTA Flash Rpp25 CLIA (research use only), 8 anti-Th/To IP positives, a cohort of 70 unselected SSc patients and sera from various disease controls (n = 357) and random healthy individuals (n = 10) were studied.ResultsAnti-Rpp25 antibodies determined by ELISA were found in 11/14 anti-Th/To IP positive but only in 1/156 (0.6%) negative samples resulting in a positive percent agreement of 78.6% (95% confidence interval [CI] 49.2, 95.3%) and a negative percent agreement of 99.4% (95% CI 96.4, 100.0%). To verify the results using a second method, 53 samples were tested by ELISA and CLIA for anti-Rpp25 reactivity and the results were highly correlated (rho = 0.71, 95% CI 0.56, 0.81; P < 0.0001). To define the cutoff of the CLIA, anti-Th/To IP positive and negative sera were tested using the anti-Rpp25 CLIA. At the cutoff selected by receiver operating characteristic (ROC) analysis 8/8 (100.0%) of the anti-Th/To positive sera but only 2/367 (0.5%) of the controls were positive for anti-Rpp25 antibodies. The positive and negative percent agreements were 100.0% (95% CI 63.1, 100.0%) and 99.5% (95% CI 98.0, 99.9%), respectively. In the disease cohorts 2/70 (2.9%) of the SSc patients were positive for anti-Rpp25 antibodies compared to 2/367 (0.5%) of the controls (P = 0.032). ROC analysis showed discrimination between SSc patients and controls with an area under the curve value of 0.732 (95% CI 0.655, 0.809).ConclusionRpp25 is a major target of autoantibodies to the Th/To autoantigen complex. Further studies are needed to evaluate the clinical utility of the new assays.

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“…The CENP-A ELISA (Dr. Fooke Laboratorien GmbH), a CE-certified peptide-based assay, was performed according to the manufacturer's instructions as described 14 . The CENP-A ELISA (Dr. Fooke Laboratorien GmbH), a CE-certified peptide-based assay, was performed according to the manufacturer's instructions as described 14 .…”

Section: Methodsmentioning

confidence: 99%

“…While a few studies have examined the prevalence of CENP-A and -B in SSc 14,15 , little has been published on the clinical correlates of the individual CENP-A or CENP-B autoantibodies. While a few studies have examined the prevalence of CENP-A and -B in SSc 14,15 , little has been published on the clinical correlates of the individual CENP-A or CENP-B autoantibodies.…”

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confidence: 99%

Clinical Correlates of CENP-A and CENP-B Antibodies in a Large Cohort of Patients with Systemic Sclerosis

et al. 2012

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Clinical and serological evaluation of a novel CENP-A peptide based ELISA

Cited by 25 publications

References 40 publications

Autoantibodies Recognizing the Amino Terminal 1-17 Segment of CENP-A Display Unique Specificities in Systemic Sclerosis

Autoantibodies Recognizing the Amino Terminal 1-17 Segment of CENP-A Display Unique Specificities in Systemic Sclerosis

Rpp25 is a major target of autoantibodies to the Th/To complex as measured by a novel chemiluminescent assay

Clinical Correlates of CENP-A and CENP-B Antibodies in a Large Cohort of Patients with Systemic Sclerosis

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