Anti-signal recognition particle autoantibody ELISA validation and clinical associations

Abstract

Objective: The aim of this study was to develop and validate a quantitative anti-signal recognition particle (SRP) autoantibody serum ELISA in patients with myositis and longitudinal association with myositis disease activity.

Methods: We developed a serum ELISA using recombinant purified full-length human SRP coated on ELISA plates and a secondary antibody that bound human IgG to detect anti-SRP binding. Protein immunoprecipitation was used as the gold standard for the presence of anti-SRP. Serum samples from three groups were analysed: SRP(+) myositis subjects by immunoprecipitation, SRP(-) myositis subjects by immunoprecipitation and non-myositis controls. The ELISA's sensitivity, specificity, positive predictive value and negative predictive value were evaluated. Percentage agreement and test-retest reliability were assessed. Serial samples from seven SRP immunoprecipitation-positive subjects were also tested, along with serum muscle enzymes and manual muscle testing.

Results: Using immunoprecipitation, we identified 26 SRP(+) myositis patients and 77 SRP(-) controls (including 38 patients with necrotizing myopathy). Non-myositis control patients included SLE (n = 4) and SSc (n = 7) patients. Anti-SRP positivity by ELISA showed strong agreement (97.1%) with immunoprecipitation (κ = 0.94). The sensitivity, specificity, positive predictive value, and negative predictive value of the anti-SRP ELISA were 88, 100, 100 and 96, respectively. The area under the curve was 0.94, and test-retest reliability was strong (r = 0.91, P < 0.001). Serial samples showed that anti-SRP levels paralleled changes in muscle enzymes and manual muscle testing.

Conclusion: We developed a quantitative ELISA for detecting serum anti-SRP autoantibodies and validated the assay in myositis. Longitudinal assessment of SRP levels by ELISA may be a useful biomarker for disease activity.

Keywords: ELISA; anti-signal recognition particle (SRP) autoantibody; idiopathic inflammatory myopathy; immunoprecipitation; quantitative measure.

Fig. 1

Anti-SRP antibody serum levels by ELISA in myositis and control subjects IP: Protein immunoprecipitation; Ab: Autoantibody; IMNM: immune mediated necrotizing myopathy; Anti-SRP: anti-signal recognition particle.

Fig. 2

Receiver operating characteristic curve for anti-signal recognition particle antibodies detected by ELISA vs protein immunoprecipitation ROC: receiver operating characteristic.

Fig. 3

Longitudinal changes in anti-signal recognition particle 54-kDa subunit antibody levels and serum creatine kinase levels over time in seven subjects with positive anti-SRP by protein and RNA immunoprecipitation Anti-SRP54: anti-signal recognition particle 54-kDa subunit antibody; CPK: creatine phosphokinase (also creatine kinase); SRP: signal recognition particle.