B-217 Newly developed ELISA for detection of circulating calprotectin in serum and plasma

Abstract

Circulating calprotectin is a biomarker that reflects activation of the innate immune system. Measuring its concentration in serum or plasma is relevant for diagnosis and monitoring of systemic inflammations, autoimmune diseases, and infections. Here we introduce a newly developed enzyme-linked immunosorbent assay (ELISA) from Euroimmun by presenting results of circulating calprotectin measurements in samples from healthy individuals and patients. Furthermore, the results of the ELISA are compared with those of a newly developed calprotectin chemiluminescence assay (ChLIA) from Immunodiagnostic Systems Ltd (IDS). To determine the reference interval of the Circulating Calprotectin ELISA, calprotectin levels were measured in 136 paired serum and K3 EDTA samples from healthy adults (74 male, 62 female, age range: [19-71] years). For assessment of inflammatory status, calprotectin concentrations were measured in a panel comprising 441 serum samples from 136 healthy adults, 25 healthy children, 80 rheumatoid arthritis (RA) patients, 50 systemic lupus erythematodes (SLE) patients, 50 Sjögren’s syndrome (SS) patients, 50 systemic sclerosis (SSc) patients, and 50 idiopathic inflammatory myopathy (IIM) patients with both the Circulating Calprotectin ELISA (Euroimmun) and the IDS Fluid Calprotectin ChLIA. Quantitative results (n=305) of both immunoassays were compared, and possible bias was estimated with Passing-Bablok regression. The upper 95% reference limit of the Circulating Calprotectin ELISA was higher for serum (2.29 µg/mL) than for plasma samples (1.31 µg/mL). Compared to healthy individuals, calprotectin concentrations were elevated in patient sera. Passing-Bablok regression indicated small constant and proportional differences between calprotectin concentrations measured with ELISA versus ChLIA (n=305, regression: y=-0.13+1.33x, 95% confidence interval (CI) intercept: [-0.14, -0.10], CI slope: [1.32, 1.34]). In comparison, the calprotectin concentrations were lower when measured with ELISA. Calprotectin concentrations measured with the Circulating Calprotectin ELISA (Euroimmun) aid in the serological assessment of inflammation status in rheumatic diseases. Despite the high agreement between the Circulating Calprotectin ELISA (Euroimmun) and the IDS Fluid Calprotectin ChLIA, the results showed differences. It is therefore necessary to emphasize that the quantitative results must be evaluated with respect to the assay-specific reference intervals. Soon, the newly developed ELISA will be applied in clinical studies set out to determine reference values, likelihood ratios, and correlation with disease states of different systemic inflammations. Future research will determine whether circulating calprotectin is a more specific biomarker of inflammation than C-reactive protein (CRP).