CiteAb’s introductory guide: multiplex immunoassays


Min Read

In this blog:

  • What types of multiplex immunoassays are there?
  • What methods of detection do these immunoassays use?
  • Discover further resources on multiplex immunoassays.

Adding data on multiplex immunoassays to CiteAb involved me and the rest of the team learning about what they are, why they are used and which types are available.

In an effort to help researchers and suppliers starting on a similar journey to us, in this blog we share a simple overview of the main types and detection methods as a framework for more detailed research.

Unlike singleplex immunoassays such as ELISA kits which detect one analyte at a time, multiplex immunoassays are antibody based assays which analyse multiple targets within a single reaction volume.

Although our data reveals singleplex immunoassays are still more highly cited than multiplex immunoassays, recently we have noticed an increase in size and competition within the multiplex immunoassay market – discussed in a recent blog post. So, what types of multiplex immunoassay are available in this growing market?

Types of multiplex immunoassay

Below, we broadly categorise multiplex immunoassays into two groups: planar/spatial based and suspension based. We then delve into examples of the methods of detection in each group.

Our data shows that suspension based multiplex immunoassays are more highly cited than planar/spatial based. They also have over three times the amount of products in our search. We will start by considering this more highly cited type. 

Suspension based 

Suspension methods work by immobilising each assay for individual analytes on a distinct bead or particle. 

Detection methods:

Examples of detection methods for this type of immunoassay include cytometric bead arrays and luminex technology. By citations, luminex based assays lead over cytometric bead arrays (using flow cytometry).

With luminex technology, fluorescent signals are used to differentiate targets. Specifically, combinations of red and near-infrared dyes encode the bead for detection, whilst phycoerythrin-labelled detector antibodies enable quantification. The dyes are read by a specialised flow cytometer. By using luminex based kits, more targets can be detected than cytometric bead arrays as two internal dyes are used.

A cytometric bead array is a flow cytometry application which works by sorting beads with internal dyes of different intensities. This method allows for a better dynamic range than methods such as ELISA which use colorimetric detection.

Planar/Spatial based

As the name would suggest, in this type of multiplexing the assays for each analyte are spatially separated. To achieve this, immobilisation occurs on a solid surface.

The solid surface is often slide based (with capture antibodies spotted on the surface), or microtitre based (a flat plate with a varying number of wells). The whole array is able to be analysed at once, with methods of automation available.

Detection methods:

Two popular methods of detection for this type of assay are electrochemiluminescence and chemiluminescence. Our data showed that chemiluminescence kits are more highly cited than kits using electrochemiluminescence.

The detection method of chemiluminescence works by having an enzyme conjugated to the detection antibody. A chemical reaction occurs if the analyte is present, generating light.

Electrochemiluminescence works by measuring light generated from electrical stimulation. Just as with chemiluminescence, this detection method enables a broad dynamic range to be achieved, as well as high sensitivity.

Find out more

To summarise, the two main types of multiplex immunoassay are planar/spatial based and bead-based. For these types, examples of detection methods include chemiluminescence and electrochemiluminescence for planar/spatial assays, and luminex or cytometric bead arrays for bead-based assays.

It is worth noting that in this overview we have not gone into the further considerations and details of these assays. Therefore, at the end of this blog we have compiled a list of references for further reading on how multiplex immunoassays work. If you know of any other resources on multiplex immunoassays that you found particularly useful, do let us know and we can add it below! 

Check out our reagent search tool where you can discover more about the multiplex immunoassays that we have citation data on. You can gain a fuller picture of how they have been used in research publications and the relevant suppliers in this research area. 

References  – Bead based assays compared to planar electrochemiluminescence methods. – An overview of multiplex immunoassays with specific details on Luminex technology. – Common questions around multiplexing answered by a variety of experts.

Tighe, Patrick J et al. “ELISA in the multiplex era: potentials and pitfalls.” Proteomics. Clinical applications vol. 9,3-4 (2015): 406-22. doi:10.1002/prca.201400130 – An overview of the types of multiplex immunoassays available (with a useful diagram) and the challenges associated with multiplexing. – A breakdown of the types of multiplex immunoassays.

  • Rhys and the CiteAb team
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