Development of a suspension array assay in multiplex for the simultaneous measurement of serum levels of four eosinophil granule proteins

Abstract

The concentrations of major basic protein (MBP), eosinophil cationic protein (ECP), eosinophil derived neurotoxin (EDN) and eosinophil peroxidase (EPO) have been associated with eosinophilic disease severity. Whereas a variety of techniques have been used to measure individual eosinophil granule protein concentration, none of these methods efficiently measures MBP, ECP, EDN and EPO simultaneously. A multiplex suspension array system was developed to simultaneously measure the concentration of MBP, ECP, EDN and EPO in serum. The assay showed excellent inter- and intra-assay reliability, and serum levels of MBP, ECP and EDN from eosinophilic subjects analyzed by ELISA and multiplex were highly correlated (r = 0.8579; P < 0.0001, r = 0.6356; P = 0.0006 and r = 0.8600; P < 0.0001, respectively, Spearman rank correlation). Moreover, the multiplex assay required 500-fold less serum than a single ELISA to achieve comparable sensitivity. Absolute eosinophil count and eosinophil surface expression of the activation marker, CD69, were significantly correlated with concentrations of MBP, EDN and EPO, but not ECP, in serum from eosinophilic subjects. Furthermore, subjects with eosinophilic gastrointestinal disorder and normal peripheral absolute eosinophil counts (< 0.5 × 109/L) had significantly increased concentrations of MBP (P < 0.0001), ECP (P < 0.0001), EDN (P = 0.0001) and EPO (P < 0.0001) compared to normal donors. In summary, the eosinophil granule protein multiplex assay provides a rapid and reliable way to measure eosinophil granule protein levels and should prove useful in assessing patterns of degranulation in patients with eosinophilic disorders.

Posttreatment Reactions After Single-Dose Diethylcarbamazine or Ivermectin in Subjects With Loa loa Infection

“…Serum levels of eosinophil granule proteins, cytokines, and chemokines were measured by suspension array multiplex immunoassays, using a previously described assay [12] and Millipore kits (Milliplex MAP Human T-Helper 17 Cytokine Panel and Cytokine/Chemokine Panels I and II, EMD Millipore). Specific mediators measured and minimal detectable levels are given in the Supplementary Data.…”

Association between eosinophilia and renal prognosis in patients with pathologically proven cholesterol crystal embolism

“…Eosinophils show the highest inflammatory activity when GM-CSF is released. Makiya et al reported that the eosinophil count and the CD69 expression correlated with the granule protein concentration [10]. Liapis et al showed that activated eosinophils contributed to thrombotic microangiopathy (TMA) [11].…”

Drug-Induced Eosinophilia Secondary to the Cotrimoxazole Administration in Acute Brucellosis

“…Since blood eosinophil numbers do not necessarily indicate the extent of eosinophil involved in the affected tissues, eosinophilic tissue infiltration and end-organ manifestations can occur in the absence of peripheral eosinophilia [5]. In such cases, measurement of eosinophil granule proteins in serum, sputum, or other body fluids can help to monitor the disease activity [7].…”