Variance in multiplex suspension array assays: Intraplex method improves reliability

Abstract

Background. Flow cytometry based suspended microarray assays are susceptible to many sources of variance; multi-well replication and inter-instrument reproducibility is uncertain. Method and results. An "intraplex" method was developed in order to minimize differences in sample readings between instruments. A full intraplex assay consists of a set m of microparticle set classifications assaying for the same analyte, with each of the m classifier sets having different sensitivity to analyte, and n classifier sets replicating each of the m levels of sensitivity, where m > 1 (generally m > 4 would be used). Conclusion. The intraplex method can compensate adequately for the sources of variance that have been identified in suspended microarray assays. It requires no changes to current equipment in use, and is a superior method of constructing precision assays. Additionally, Luminex® users may want to consider the evidence that shows that despite calibration to the same standard, two instruments may not give similar results for all concentrations of analytes. © 2007 Hanley; licensee BioMed Central Ltd.