Quantification of C1 esterase inhibitor in human serum by enzyme-linked immunosorbent assay: Correlation with turbidimetric immunoassay

C1 inhibitor of serine proteases (C1-INH) performs a regulatory function in the complement system and vascular permeability. Deficiency of C1-INH leads to various forms of angioedema, including hereditary angioedema (HAE). The cause of HAE is a genetically determined violation of the synthesis of C1-INH. A decrease in the level of C1-INH to 50% relative to the norm leads to an increase in the production of bradykinin, which is the basis for the diagnosis of HAE. The development of affordable ELISA for the quantitative determination of C1-INH is a popular direction for clinicians. During the development of a new kit for quantitative determination of C1-INH, two mouse monoclonal antibodies (mAb) with different epitope specificities were obtained. On their basis, a sandwich-type ELISA was developed. The specificity of the obtained mAb's was confirmed using the medical device “Berinert”. To prepare calibrators, C1-INH was affinity purified from human blood plasma using a sorbent with immobilized mAbs. The identity of the C1-INH protein was confirmed by PAGE electrophoresis, immunoblotting, and mass spectrometry on MALDI-TOF/TOF UltrafleXtreme mass spectrometer. To assess the quality indicators of developed reagents kit, studies were carried out in accordance with GOST R 51352-2013 and TU 21.20.23-041-01967164-2022. Values of quality indicators: accuracy — 93.53%; measurement linearity interval — 22.00-176.07 ng/mL. Using the developed ELISA test system, we examined 28 blood sera from healthy donors and 7 blood sera from patients with confirmed HAE. In the same samples, the content of C1-INH was determined by turbidimetric method, using the "Diagnostic reagents for in vitro immunochemical studies of specific blood proteins. Model: C1-esterase inhibitor (C1 EsteraseInhibitor)" (Aptec, Belgium). The correlation coefficient was 0.94 (p < 0.05). It was found that the diagnostic sensitivity and specificity of the developed ELISA is 100%. As a result of the study, an original ELISA test system for the quantitative determination of C1-INH was developed "Reagent kit for enzyme-linked immunosorbent assay of human C1-inhibitor (C1-inh PS)".