[A novel electrochemiluminescence method for detecting insulin autoantibody].

To establish the electrochemiluminescence (ECL) method for insulin autoantibody (IAA) detection in China and preliminarily evaluate its application value. Proinsulin was labeled with Sulfo-tag and biotin, and then incubated with IAA in serum. The Meso Scale Discovery (MSD) streptavidin plate was used to capture the labeled antigen-antibody complex, which was subsequently detected by MSD electrochemiluminescence machine. IAA index ≥ 0.005 was used as a positive threshold. After optimizing the detection conditions, the correlation and consistency of ECL method and microplate radioimmunoassay were compared. Patients with type 1 diabetes mellitus (T1DM, 55) and first-degree relatives (216) from the Department of Metabolism and Endocrinology of the Second Xiangya Hospital of Central South University from 2016 to 2018, and healthy volunteers (413) were selected as subjects, and their serum samples were tested to evaluate the clinical application value of ECL. The optimized conditions included as follows: (1) The optimal concentration of biotin-proinsulin and Sulfo-tag-proinsulin was 800 ng/ml. (2) Serum acidification was achieved by incubating for 45 min. (3) Adding 35 μl diluted antigen, followed by adding 15 μl Tris-HCl to the reaction system, and then incubated with acidified serum overnight to obtain a higher signal-to-noise ratio (S/N). Twenty-two samples were repeatedly detected by ECL method at intervals of 3 months. There was no significant difference in IAA index (0.095) and the positive or negative results was completely consistent. The agreement rate between ECL method and microplate radioimmunoassay for IAA was 93.7% (119/127) (Kappa value: 0.858), and the index showed a significant positive correlation (correlation coefficient 0.749, 0.001). The area under the curve (AUC) of ECL assay was 0.764 (95: 0.669 to 0.858). The positive rate of IAA in T1DM patients by ECL method was 52.73% (29/55), which was significantly higher than 0.76% (2/263) in healthy controls. The positive rate of IAA in T1DM first-degree relatives group was 0.93% (2/216), and there was no significant difference compared with healthy control group (0.854). ECL method of IAA has the features of high sensitivity and specificity, simple operation and no radioactive contamination, indicating its appreciable clinical application value.