Multiplexed assays for simultaneously detecting several biomarkers in a single sample are becoming popular in clinical diagnostic research. An advantage of multiplexing in diagnostics is that patterns of several biomarkers have better predictive value compared to detection of single analyte using ELISAs. Several commercial multiplexing platforms are vying for success in this emerging market and include Luminex bead based platform, Meso Scale Discovery’s Multi Array Technology and protein array platforms from Whatman, GentelBio and others. Though various assay platforms are available their rigorous validations using large cohorts are lacking.
A recent report in Clinical Chemistry by Fu et al. compares the five multiplexed cytokine assays for reproducibility, reliability, robustness, accuracy, and throughput. The assay platforms compared were
- MULTI-ARRAY (Meso Scale Discovery),
- Bio-Plex (Bio-Rad Laboratories),
- A2 (Beckman Coulter),
- FAST Quant (Whatman Schleicher & Schuell BioScience), and
- FlowCytomix (Bender MedSystems)
Their conclusion: “The MULTI-ARRAY and Bio-Plex assays (based on Luminex Platform) had the best performance with the lowest limits of detection, and the MULTI-ARRAY system had the most linear signal output over the widest concentration range”
The study is useful but doesn’t go into details about the reasons behind the success and failure of a specific platform. This is obviously not the first study of its kind. Another study (I wrote about it few months back) using protein arrays concluded that based on FDA guideline of acceptable intra- and inter- assay precision to be <30%, substantial portion of measurements will be rejected.
Another study relevant to current study by Fu et al. was reported by Nechansky et al. and compared Cytokine assay from three commercial sources. All three assays were based on Luminex platforms. The authors concluded “– that performing an assay “according to the manufacturer’s instructions” does not necessarily guarantee reliable quantitative data—and even the collection of qualitative data may be questionable.” This study raises the question whether the conclusion of the current study will change if the Luminex based cytokine kits from other vendors are used.
These reports show the challenges facing the multiplexed assays that still remain to be solved.