title: 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 concentrations 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 (<0.5×10(9)/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.
authors.affiliation: Eosinophil Pathology Unit, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United States. Electronic address: makiyam@niaid.nih.gov.
authors.forename: Michelle A
authors.initials: MA
authors.lastname: Makiya
authors.affiliation: Helminth Immunology Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United States.
authors.forename: Jesica A
authors.initials: JA
authors.lastname: Herrick
authors.affiliation: Eosinophil Pathology Unit, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United States.
authors.forename: Paneez
authors.initials: P
authors.lastname: Khoury
authors.affiliation: Mast Cell Biology Section, Laboratory of Allergic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United States.
authors.forename: Calman P
authors.initials: CP
authors.lastname: Prussin
authors.affiliation: Helminth Immunology Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United States.
authors.forename: Thomas B
authors.initials: TB
authors.lastname: Nutman
authors.affiliation: Eosinophil Pathology Unit, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United States.
authors.forename: Amy D
authors.initials: AD
authors.lastname: Klion
mesh_terms: D000818:Animals; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D047091:Eosinophil Granule Proteins / Q000097:blood* / Q000276:immunology; D004802:Eosinophilia / Q000097:blood* / Q000276:immunology; D005260:Female; D005767:Gastrointestinal Diseases / Q000097:blood* / Q000276:immunology; D006801:Humans; D008297:Male; D051379:Mice; D012680:Sensitivity and Specificity
chemical_list: D047091:Eosinophil Granule Proteins
keywords: Degranulation; ELISA; Eosinophil; Eosinophil granule proteins; Multiplex