Department of Dermatology, Faculty of Medicine, University of Tsukuba, Ibaraki, Japan.
Department of Dermatology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
J Dermatol. 2022 Apr;49(4):441-447. doi: 10.1111/1346-8138.16295. Epub 2021 Dec 29.
A line blotting assay (LB) is currently used to detect myositis-specific autoantibodies (MSAs) in patients with idiopathic inflammatory myopathies (IIMs), because of its simplicity; however, the sensitivity and specificity of this assay is low. The aim of this study is to evaluate the accuracy of the commercial LB in detection of antinuclear matrix protein 2 (NXP2) antibody. Seventy-seven serum samples from patients with IIMs, in which anti-NXP2 antibodies were detected through immunoprecipitation and western blotting (IP-WB) using K562 cell lysate, were enrolled. All samples were assessed by LB and IP-WB using recombinant human NXP2 whole protein (rNXP2) produced by insect cells, and the positive rates of each assay were compared. Thirty-two samples (41.6%) showed false-negativity by LB, which includes 11 samples with negative results by IP-WB using rNXP2. Relative intensities of IP-WB using cell lysate were significantly higher in the samples with positive results by both LB and IP-WB using rNXP2, compared to samples with positive by IP-WB using rNXP2 but negative by LB. Three of 11 samples with negative results by both LB and IP-WB using rNXP2 revealed high antibody titers. Further, differences in post-transcriptional SUMOylation were observed between recombinant and natural NXP2 proteins. In conclusion, the LB showed low sensitivity for detection of anti-NXP2 antibody, an effect exacerbated at low titers of anti-NXP2 antibodies. Moreover, there appears to be differences in the reactivities of antibodies to recombinant and natural NXP2 proteins with different post-transcriptional modifications.
线印迹检测（LB）目前用于检测特发性炎性肌病（IIM）患者中的肌炎特异性自身抗体（MSA），因其操作简单；然而，该检测方法的灵敏度和特异性较低。本研究旨在评估商业 LB 检测抗核基质蛋白 2（NXP2）抗体的准确性。共纳入 77 例通过免疫沉淀和 Western blot（IP-WB）使用 K562 细胞裂解物检测到抗 NXP2 抗体的 IIM 患者血清样本。所有样本均通过 LB 和使用昆虫细胞产生的重组人 NXP2 全长蛋白（rNXP2）进行 IP-WB 进行评估，并比较了每种检测方法的阳性率。LB 检测有 32 个样本（41.6%）出现假阴性，其中包括 11 个使用 rNXP2 的 IP-WB 检测为阴性的样本。与 LB 检测为阴性而 rNXP2 的 IP-WB 检测为阳性的样本相比，LB 和 rNXP2 的 IP-WB 检测均为阳性的样本的 IP-WB 检测使用细胞裂解物的相对强度更高。在 rNXP2 的 LB 和 IP-WB 检测均为阴性的 11 个样本中的 3 个显示出高抗体滴度。此外，在重组和天然 NXP2 蛋白之间观察到翻译后 SUMOylation 的差异。总之，LB 检测抗 NXP2 抗体的灵敏度较低，在抗 NXP2 抗体的滴度较低时效果更为明显。此外，似乎存在抗体对具有不同转录后修饰的重组和天然 NXP2 蛋白的反应性差异。
