title: Overcoming matrix matching problems in multiplex cytokine assays.
abstract: Failure to match assay matrices with samples in immunoassays can result in incorrect sample values being reported. For multiplex assays this presents particular problems, due to the need to find a matrix suitable for all the analytes. Here, we describe strategies adopted to overcome matrix problems identified in establishing a cytokine multiplex assay in human plasma. Standard analytes were diluted in plasma samples to identify representative plasma for assay development. Horse sera were screened to evaluate potential interference before using to adjust a matrix to match plasma samples. Suitability of the matrix match was confirmed by evaluating recovery of known concentrations of analytes from plasma. Individual plasmas modified the assay signal for some analytes to a variable extent, particularly for IL-1α and IL-1β. Addition of horse serum to assay buffer improved matching to plasma samples, although endogenous MCP-1 activity was apparent in one sample. Matching of plasma and assay matrices allowed recoveries within 10% to 20% of the expected values, unless the samples contained atypical interfering activity. Attention to choice of samples and diluent used for assay development is particularly important for measurement of sample analytes in cytokine multiplex assays.
authors.affiliation: Brain Injury Research Group, Clinical Sciences Building, Salford Royal Foundation Trust, Salford, M6 8HD, UK. Electronic address: margaret.hoadley@manchester.ac.uk.
authors.forename: Margaret E
authors.initials: ME
authors.lastname: Hoadley
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Hopkins
mesh_terms: D000818:Animals; D018932:Chemokine CCL2 / Q000097:blood*; D016207:Cytokines / Q000097:blood*; D006736:Horses / Q000097:blood; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D053582:Interleukin-1alpha / Q000097:blood*; D053583:Interleukin-1beta / Q000097:blood*; D010949:Plasma / Q000378:metabolism
chemical_list: C111835:CCL2 protein, human; D018932:Chemokine CCL2; D016207:Cytokines; D053582:Interleukin-1alpha; D053583:Interleukin-1beta
keywords: Assay matrix; Cytokines; IL; Immunoassay; MCP-1; Multiplex; TNF-α; interleukin; monocyte chemoattractant protein-1; tumour necrosis factor-α