{'fraction_non_alphanumeric': 0.02735562310030395, 'fraction_numerical': 0.010638297872340425, 'mean_word_length': 5.562678062678063, 'pattern_counts': {'":': 0, '<': 0, '<?xml version=': 0, '>': 0, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 0, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
Enzyme‐linked immunosorbent assays for diagnosis of immune‐mediated necrotizing myopathy

Immune-mediated necrotizing myopathy (iNM), characterized by necrosis and regeneration of muscle fibers in the absence of prominent inflammatory cells, is the fourth category of inflammatory myopathy, besides polymyositis, dermatomyositis and inclusion body myositis. Autoantibodies are considered to be important in the pathogenesis of iNM. Autoantibodies against signal recognition particle (SRP) or 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) are regarded as typical of iNM. These iNMassociated autoantibodies are useful in diagnosis and in considerations of immunosuppressive therapy, because there is usually a lack of immunological findings in muscle histology. Signal recognition particle is a cytoplasmic ribonucleoprotein consisting of six proteins and 7S RNA. SRP regulates the translocation of proteins across the endoplasmic reticulum membrane during protein synthesis. A case series accumulated from the field of rheumatology showed that anti-SRP antibodies were associated with treatment resistance and refractory myositis. Recently, anti-SRP antibodies have been observed to entail a broad variety of neuromuscular phenotypes, collected under the heading “anti-SRP myopathy.” Anti-SRP myopathy affects individuals of various ages, predominantly females. It has been reported that 66% of SRP-positive patients have prominent muscular atrophy in their limb and trunk muscles. Muscle weakness and atrophy progress more slowly, potentially leading to a misdiagnosis of muscular dystrophy. Anti-SRP detection is useful for differential diagnoses among a variety of myopathies. RNA immunoprecipitation is an original detection method for anti-SRP antibodies. One disadvantage is that RNA immunoprecipitation requires a complicated technique, and is carried out at only a limited number of facilities. In addition, antibody titers are not determined by RNA immunoprecipitation. Other conventional detection assays, including enzymelinked immunosorbent assay (ELISA) and line blot assay using a recombinant SRP protein, are typically used. Statins are associated with myotoxic effects including myalgia, elevation of creatine kinase and transaminases, and weakness. These problems usually disappeared after the discontinuation of statin treatment. However, the muscular symptoms of some statin-exposed patients do not improve after the cessation of statins, and the symptoms progress to inflammatory myopathy requiring immunosuppressive therapy. In 2011, Mammen et al. identified autoantibodies to HMGCR in patients with statininduced myopathy. Although anti-HMGCR antibodies were originally identified by protein immunoprecipitation assay using radiolabeled culture cell extracts, many researchers agree that ELISA is a preferred standard method. We established antiHMGCR ELISA with a recombinant protein, and showed the association between anti-HMGCR antibodies and iNM. Anti-HMGCR antibodies have also been detected in patients without statin treatment. Anti-SRP and anti-HMGCR antibody studies are not used in routine clinical practice because there is no convenient, widely accepted assay, such as radioimmunoassay for anti-acetylcholine receptor antibodies. Our goal is to establish accurate quantitative assays for iNM-associated autoantibodies. We have developed new ELISA for anti-SRP and antiHMGCR with some modifications of previous studies (Cosmic Corporation, Tokyo, Japan). The optical density at 450 nm was read with an automatic plate reader (Molecular Devices, Tokyo, Japan). Values were determined from the results of optical density at 450 nm using a standard curve obtained with serial concentrations. The cut-off value was calculated from the mean + 5 9 SD of 240 disease–control sera, and arbitrarily set to 1 IU/mL. Figure 1 shows the results of anti-SRP and antiHMGCR ELISA. We studied sera from 40 patients with SRP-positive iNM, 40 with HMGCR-positive iNM, 40 with seronegative myositis, 40 with dermatomyositis, 40 with inclusion body myositis, 40 with myasthenia gravis and 40 with muscular dystrophy. Positivity for the anti-SRP antibody was observed in 39 of 40 patients with SRP-positive iNM, which were determined by the RNA immunoprecipitation. In contrast, none of the 240 patients with other diseases had antiSRP antibodies. One patient with low titer of anti-SRP antibody was 40-year-old man. He had moderate limb weakness and muscle atrophy, an increased level of ceratine kinase (6834 IU/L) and iNM muscle biopsy, which were compatible with anti-SRP myopathy.