A new diagnostic algorithm for pattern-oriented autoantibody testing according to the ICAP nomenclature: A pilot study.

The commercial tests currently available as second-level tests to detect ANA sub-specificities are generally used independently from the ANA immunofluorescence (IIF) pattern. The aim of this study was to evaluate the efficacy of the use of a customizable pattern-oriented antigenic panel by immunoblot (IB) using the International Consensus on ANA Patterns (ICAP) classification scheme, in order to introduce a novel and updated autoimmune diagnostic flowchart. 710 sera referred for routine ANA testing were selected on the basis of the ANA pattern according to the ICAP nomenclature (nuclear speckled AC-2,4,5; nucleolar AC-8,9,10,29; cytoplasmic speckled AC-18,19,20) and on an IIF titer ≥1:320. They were then assayed by three experimental IB assays using a panel of selected antigens. ICAP-oriented IB detected 515 antibody reactivities vs. 457 of traditional anti-ENA in the nuclear speckled pattern group, 108 vs. 28 in the nucleolar pattern group, and 43 vs. 34 in the cytoplasmic speckled pattern. This pilot study may lead the way for a new approach introducing an ICAP pattern-oriented follow up testing as a valid alternative to the existing standard panels, thus enabling more patients with autoimmune rheumatic disease to be accurately diagnosed.

Anti-HMGCR antibodies as a biomarker for immune-mediated necrotizing myopathies: A history of statins and experience from a large international multi-center study.

In an effort to find naturally occurring substances that reduce cholesterol by inhibiting 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR), statins were first discovered by Endo in 1972. With the widespread prescription and use of statins to decrease morbidity from myocardial infarction and stroke, it was noted that approximately 5% of all statin users experienced muscle pain and weakness during treatment. In a smaller proportion of patients, the myopathy progressed to severe morbidity marked by proximal weakness and severe muscle wasting. Remarkably, Mammen and colleagues were the first to discover that the molecular target of statins, 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR), is an autoantibody target in patients that develop an immune-mediated necrotizing myopathy (IMNM). These observations have been confirmed in a number of studies but, until today, a multi-center, international study of IMNM, related idiopathic inflammatory myopathies (IIM), other auto-inflammatory conditions and controls has not been published. Accordingly, an international, multi-center study investigated the utility of anti-HMGCR antibodies in the diagnosis of statin-associated IMNM in comparison to different forms of IIM and controls. This study included samples from patients with different forms of IIM (n=1250) and patients with other diseases (n=656) that were collected from twelve sites and tested for anti-HMGCR antibodies by ELISA. This study confirmed that anti-HMGCR autoantibodies, when found in conjunction with statin use, characterize a subset of IIM who are older and have necrosis on muscle biopsy. Taken together, the data to date indicates that testing for anti-HMGCR antibodies is important in the differential diagnosis of IIM and might be considered for future classification criteria.

The ANA-reflex test as a model for improving clinical appropriateness in autoimmune diagnostics.

Reflex tests are widely used in clinical laboratories, for example, to diagnose thyroid disorders or in the follow-up of prostate cancer. Reflex tests for antinuclear antibodies (ANA) have recently gained attention as a way to improve appropriateness in the immunological diagnosis of autoimmune rheumatic diseases and avoid waste of resources. However, the ANA-reflex test is not as simple as other consolidated reflex tests (the TSH-reflex tests or the PSA-reflex tests) because of the intrinsic complexity of the ANA test performed by the indirect immunofluorescence method on cellular substrates. The wide heterogeneity of the ANA patterns, which need correct interpretation, and the subsequent choice of the most appropriate confirmatory test (ANA subserology), which depend on the pattern feature and on clinical information, hinder any informatics automation, and require the pathologist's intervention. In this review, the Study Group on Autoimmune Diseases of the Italian Society of Clinical Pathology and Laboratory Medicine provides some indications on the configuration of the ANA-reflex test, using two different approaches depending on whether clinical information is available or not. We further give some suggestions on how to report results of the ANA-reflex test.

Diagnostic accuracy and predictive value of extended autoantibody profile in systemic sclerosis.

Systemic sclerosis (SSc) is a heterogeneous autoimmune disorder characterized by microvascular injury, fibrosis of the skin and other organs, and presence of antinuclear autoantibodies (ANA) with a prevalence varying from 80 to 98%. The ANA classically detected in SSc include anti-centromere (ACA) and anti-topoisomerase I (ATA), which are positive in 50-60% of the patients. Even if other autoantibodies, such as anti-fibrillarin (AFA), anti-RNA polymerase III (RNAP III), anti-PMScl, anti-Th/To, and anti-hUF/NOR-90, are almost specific for SSc, until recently they were not routinely looked for, since the techniques for their identification were not suitable for routine use. In recent years, the advances in the knowledge of the biochemistry and of the immunoreactive sites of the autoantigens led to the development of new immunoassays using recombinant proteins as autoantigens. We evaluated a new multiplex line immunoblot assay (LIA) for the simultaneous detection of 13 different SSc-associated autoantibodies, in a cohort of 210 SSc Italian patients. The sensitivity and the specificity of this assay were as follows: 30.5% and 97.3% for ACA (anti-CENP-B), 29.5% and 96% for ACA (anti-CENP-A), 20% and 99.3% for ATA, 5.7% and 99.3% for anti-RNAP III (RP-155), 5.2% and 100% for anti-RNP III (RP-11), 6.7% and 98% for anti-PMScl (PMScl-100), 10.9% and 93.3% for anti-PMScl (PMscl-75), 3.3% and 98.7% for anti-Th/To, 0.48% and 100% for AFA, 4.8% and 96.7% for anti-hUF/NOR-90, 4.7% and 96% for anti-Ku, 0.95% and 100% for anti-Platelet-Derived Growth Factor Receptor, and 18.1% and 50% for anti-Ro-52, respectively. These results, which are similar to those obtained in other studies using traditional techniques, show that the LIA assay can be considered a more rapid and a more practical method than immunoprecipitation assays for studying SSc-related antibodies in the diagnostic work-up of SSc patients.