Development of a custom pentaplex sandwich immunoassay using Protein-G coupled beads for the Luminex® xMAP® platform.
File(s)
Published version
Author(s)
McDonald, JU
Ekeruche-Makinde, J
Ho, MM
Tregoning, JS
Ashiru, O
Type
Journal Article
Abstract
Multiplex bead-based assays have many advantages over ELISA, particularly for the analyses of large quantities of samples and/or precious samples of limited volume. Although many commercial arrays covering multitudes of biologically significant analytes are available, occasionally the development of custom arrays is necessary. Here, the development of a custom pentaplex sandwich immunoassay using Protein G-coupled beads, for analysis using the Luminex® xMAP® platform, is described. This array was required for the measurement of candidate biomarkers of vaccine safety in small volumes of mouse sera. Optimisation of this assay required a stepwise approach: testing cross-reactivity of the antibody pairs, the development of an in-house serum diluent buffer as well as heat-inactivation of serum samples to prevent interference from matrix effects. We then demonstrate the use of this array to analyse inflammatory mediators in mouse serum after immunisation. The work described here exemplifies how Protein G-coupled beads offer a flexible and robust approach to develop custom multiplex immunoassays, which can be applied to a range of analytes from multiple species.
Date Issued
2016-02-24
Date Acceptance
2016-02-22
Citation
Journal of Immunological Methods, 2016, 433, pp.6-16
ISSN
1872-7905
Publisher
Elsevier
Start Page
6
End Page
16
Journal / Book Title
Journal of Immunological Methods
Volume
433
Copyright Statement
Crown Copyright © 2016 Published by Elsevier B.V. This is an open access article under the Open Government
License (OGL) (http://www.nationalarchives.gov.uk/doc/open-government-licence/version/3/).
License (OGL) (http://www.nationalarchives.gov.uk/doc/open-government-licence/version/3/).
Sponsor
Commission of the European Communities
Identifier
PII: S0022-1759(16)30033-3
Grant Number
115308
Subjects
Beads
Biomarkers
Protein G
Sandwich immunoassay
Serum
xMAP®
Publication Status
Published