Multiplex bead array assays: Performance evaluation and comparison of sensitivity to ELISA☆

Abstract

The measurement of soluble cytokines and other analytes in serum and plasma is becoming increasingly important in the study and management of many diseases. As a result, there is a growing demand for rapid, precise, and cost-effective measurement of such analytes in both clinical and research laboratories. Multiplex bead array assays provide quantitative measurement of large numbers of analytes using an automated 96-well plate format. ELISAs (enzyme linked immunosorbent assay) have long been the standard for quantitative analysis of cytokines and other biomarkers, but are not well suited for high throughput multiplex analyses. However, prior to replacement of ELISA assays with multiplex bead array assays, there is a need to know how comparable these two methods are for quantitative analyses. A number of published studies have compared these two methods and it is apparent that certain elements of these assays, such as the clones of monoclonal antibodies used for detection and reporting, are pivotal in obtaining similar results from both assays. By careful consideration of these variables, it should be possible to utilize multiplex bead array assays in lieu of ELISAs for studies requiring high throughput analysis of numerous analytes.

Endometriosis-associated changes in serum levels of interferons and chemokines

“…We could also take in consideration as a limitation the detection sensitivity of multiplexed immunoassays. It was shown that while multiplexed immunoassays have sensitivity comparable to conventional ELISA, it is possible that the robustness may vary among different multiplex bead arrays (29).…”

Development of a custom pentaplex sandwich immunoassay using Protein-G coupled beads for the Luminex® xMAP® platform

“…Serum is a complex mix of proteins that may non-specifically bind to microspheres -known as matrix effects (Elshal and McCoy, 2006). These matrix effects can interfere with the performance of a xMAP® assay and may be observed as poor bead recovery, instrumentation clogging, low signals, or variable results.…”

Expression of heat shock proteins in medulloblastoma

“…The sensitivity of each analyte, based on the lower-used standard, is as follows: Akt (0.2 ng/ml), HSP27 phospho-Ser 82 (5.2 U/ ml), HSP27 phospho-Ser 15 (3.4 U/ml), HSP40 (0.2 ng/ml), HSP60 (3.4 ng/ml), HSP70 (0.2 ng/ml), HSP90-α (0.3 ng/ ml), and Akt phospho-Ser473 (2.0 U/ml). 8 For the analysis we used FACSCalibur Flow Cytometer equipped with 2 lasers (488 and 635 nm) and 6 parameters (Fsc, Ssc, and FL1-FL4).…”