Luminex-Based Immunoassay Systems for Biomedical Labs

By Deliana InfanteReviewed by Megan Craig, M.Sc.

Luminex-based immunoassays are advanced diagnostic tools that simultaneously detect multiple proteins and genes, enhancing efficiency in research, clinical, and industrial settings. Leveraging multiplex technology, they provide valuable insights into disease progression and drug toxicity, driving innovation in personalized medicine and biotechnology.

What is Luminex and the xMAP Technology?

Luminex is a biotech company that offers innovative multiplexing technologies for the detection and measurement of RNA and proteins.¹

In 2021, Luminex was adquired by DiaSorin, an Italian company leader in the development, production, and distribution of immunological and molecular diagnostics.^2-3 With Luminex's acquisition, DiaSorin is now in possession of the xMAP technology, one of the most versatile high-tech used in the biopharmaceutical industry, in public health surveillance and academic research.

Luminex xMap technology is a microplate-formatted multiplexed immunoassay system based on beads. Depending on the system architecture, it can concurrently detect up to 500 targets in a single sample.⁴ Secreted protein and mRNA expression levels can be measured on a Luminex instrument, noticing that the limits of sensitivity, specificity to a target, and quality control are overcome by xMAP compared to other methods.⁴

The technique offers several advantages, including the flexibility to singleplex or multiplex, simultaneous measurement of multiple targets in a single sample, reduced cost and labor through multiplexing, real-time tracking of biological interactions, and smaller sample requirements compared to other techniques like ELISA.⁴

Additionally, using beads with a larger surface area than a flat surface enhances the technique's sensitivity. Liquid reaction kinetics can increase the detection range and provide faster, more reproducible results compared to planar arrays.⁴

This platform is versatile and can be used for analyzing proteins (including phosphoproteins), signaling molecules, and nucleic acids.⁴ Luminex offers a wide variety of kits for detecting diseases, cytokines, growth factors, and more.⁴

Assays kits are available for different species, such as humans, monkeys, mice, and rats.⁴ Customized assays can also be designed. Depending on the experiment design, proteins and nucleic acids can be bound to either magnetic or non-magnetic beads.⁴

How Do Luminex-based Immunoassays Work?

Luminex xMAP-based Immunoassays work with a bead-based multiplexing system that can detect and quantify multiple analytes in a single sample using beads with unique spectral addresses.⁴

These beads have color codes that are created by internally labeling them with varying ratios of two fluorophores.⁴ Different molecules, such as proteins or nucleic acids that are specific to the selected targets can be attached to these beads. In the kits, each antibody is covalently attached to a different bead with a unique spectral code.⁴

The samples are incubated with the beads. Then, targets in the samples bind to their specific antibodies or capture molecules.⁴ Similar to a sandwich ELISA, the beads are washed and incubated with a combination of biotinylated antibodies against all targets.⁴

After an additional wash, the beads are incubated with the reporter streptavidin.⁴ A dual laser system is utilized to detect both the presence and intensity of the reporter associated with each bead, providing information about the identity of the target molecule and its concentration in the sample.⁴

This technology effectively detects variations in the concentrations of numerous targets under different conditions.⁴

Clinical and Research Applications of Luminex-based Immunoassays

The potential applications of this technique are not limited to cytokines. The surface of beads can be conjugated with antibodies, receptors, peptides, and oligonucleotides to facilitate immunoassays, receptor-ligand assays, enzymatic assays, DNA assays, and other applications.⁵ This allows for protein expression profiling, gene expression profiling, infectious disease diagnosis, biomarker discovery, disease surveillance, drug discovery, vaccine development, microRNA analysis, HLA testing, and more.⁵

During the COVID-19 pandemic, Multiplex Luminex technology has proven to be a valuable tool for detecting antibodies to SARS-CoV-2.⁶ These assays are utilized for epidemiology and sero-surveillance to monitor the pandemic's progression, distinguish individuals who have developed vaccine-induced immunity from those who have been naturally infected, and identify potential donors of convalescent plasma for therapeutic purposes.⁶

The Luminex xMAP SARS-CoV-2 Multi-Antigen IgG Assay, which can detect and identify antibodies against three SARS-CoV-2 antigens, was used for this.⁶ It is a versatile kit that can be customized to evaluate other sample types, antibody isotypes, and potential neutralizing antibody responses.⁶

Luminex-based Immunoassays Equipment

Several Luminex-based instruments are available for multiplexing purposes. However, the most widely used instruments are those in the Luminex product pipeline, including the FLEXMAP 3D System⁷, MAGPIX System⁸, Luminex 200™ System⁹, and xMAP INTELLIFLEX¹⁰.

Among these four instruments, the FLEXMAP 3D System is the most advanced and versatile multiplexing platform. It has a fast read time and is compatible with 96- and 384-well plates.⁷ It is easy to integrate with front-end automation systems and has multiplexing capability for up to 500 analytes in a single sample.⁷

The second equipment, the MAGPIX System, is the most affordable of all models and has a compact fluorescent detection system suitable for medium-throughput multiplex immunoassays.⁸ The system can read up to 50 distinct magnetic bead regions with only 20 μl per sample, providing a higher throughput of up to 4,800 results in one hour.⁸

The Luminex 200™ System is known for being user-friendly.⁹ Its compact analyzer allows for up to 100 assays to be performed simultaneously in a single well of a microtiter plate, and both magnetic and non-magnetic beads can be used with this equipment.⁹

Additionally, the xMAP INTELLIFLEX is an advanced multiplexing platform with a compact footprint.¹⁰ It enables fast read times for both 96- and 384-well plate formats and possesses dual-reporting capabilities with a second violet laser and 4.5 logs of dynamic range to collect two parameters per analyte.¹⁰ Additionally, it has a side eject door compatible with automation to maximize throughput.¹⁰

All of these features streamline startup and shutdown protocols and minimize maintenance requirements.^7-10

These instruments have numerous applications in clinical diagnostics, genomics, proteomics, drug discovery, and academic research.^7-10 These applications range from SNP analysis and drug-target validation to genetically inherited disease screening and antibody epitope mapping, among others.^7-10

The choice of instrument depends on the specific application. For example, the xMAP INTELLIFLEX is an advanced piece of equipment with a small footprint, high multiplexing capabilities, and efficient data collection.¹⁰ However, it is not suitable for clinical applications and is better suited for research purposes.¹⁰

Further Reading

• All Antibodies Content
• What are Therapeutic Antibodies?
• What are Bispecific Antibodies?
• Creating a Nanobody Library with PCR
• Proving Monoclonality with PCR