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Multiplex Assays for Accurate, Consistent Analysis

Luminex Flexmap 3D multiplex instrument

Need Maximum Analyte Flexibility and Selection?

Profile and validate your biomarker candidates with Luminex® Discovery and Luminex High Performance multiplex immunoassays.   

  • Multiplex up to 50 analytes per sample.
  • Choose from more than 450 analytes in 3 flexible formats.
  • Customize panels to fit your needs.
  • Rigorously validated for accuracy, reproducibility, and lot-to-lot consistency.
Automated ELISA system Ella runs high throughput multiplex assays

Need High Throughput and Fast Validation?

Bring a higher level of precision and automation to your immunoassay workflow with Simple Plex assays on the Ella platform.

  • Get consistent, reproducible results in just 90 minutes.
  • Generate triplicate data with only 25 µL of sample volume.
  • Reduce human error and variability with hand-free automation.
  • Choose from a menu of more the 200 fully validated assays.
Multiplex assay options including multiplex western system, multiplex arrays and automated multiplex immunoassays

 

Types of Multiplex Assays

 

Proteome Profiler antibody arrays
Screening/Exploration

Antibody Arrays

Providing broad, cost-effective early-stage screening, antibody arrays offer a quick and inexpensive analysis of many analytes simultaneously—in less time than it takes to perform a Western blot.   

Similar to an ELISA but with expanded multiplexing capabilities, these semiqualitative antibody arrays can analyze hundreds of different proteins simultaneously with substantially less time and effort.  Rather than a plate, antibody arrays use a membrane as a substrate. Capture antibodies are pre-spotted in duplicate with high-quality capture antibodies for multiplexing.  

With superior specificity, low background noise, and no cross-reactivity, you can count on this assay for clear and consistent data. 

  • Fast, economical analysis
  • No specialized equipment needed
  • Measure more than 100 analytes at once
  • Quickly analyze complex signaling responses.
Luminex Multiplex Assays
Precise, Accurate Biomarker Discovery

Luminex Multiplex Assays

Luminex multiplex assays are designed to maximize multiplexing capacity and flexibility while maintaining assay specificity. The bead-based Luminex format allows you to better tailor assay selection to your individual research needs while overcoming many common multiplexing challenges. 

Utilizing the same sandwich principles of traditional ELISAs, color-coded microspheres, or beads, are internally dyed with different proportions of red and infrared fluorophores that correspond to a distinct spectral signature, or bead region. Antibodies specific to the desired analyte are coupled to a unique bead region and are incubated with the sample.

Luminex multiplex assays allow you to quantify up to 50 biomarkers with less sample volume than a traditional plate-based ELISA.

  • Accurate bead-based assays
  • Multiplex up to 50 analytes simultaneously 
  • Customize panels to fit your needs
  • Rigorously validated for accuracy, reproducibility, and lot-to-lot consistency
Automated ELISA systems Ella runs single, multianalyte and multiplex immunoassays
Fully Automated ELISA

Cartridge-Based Immunoassays

With multi-analyte cartridge-based assays, you get the same specificity of a single-analyte ELISA but with greater sensitivity and a broader dynamic range. The automated immunoassay cartridges deliver results in just 90 minutes with better sensitivity and reproducibility than plate-based assays.  

Each cartridge accommodates up to 16, 32, or 72 samples, and each well is sampled three times to give you all the results in triplicate. The antibodies as well as the fluorescent detector are pre-functionalized in the cartridge, eliminating the need to run a standard curve.

Disposable cartridge kits come in a variety of multi-analyte and single-analyte configurations and include everything you need to run your automated ELISA. The reduction in manual steps enables a high level of reproducibility and data quality—all with a method that's fast and easy to transfer. 

  • Highly sensitive results in only 90 minutes
  • Choose from over 150 validated targets
  • Hands-free automation with high-throughput
  • Available in single and multi-analyte cartridge formats

The Latest in Multiplexing Technology

Luminex Metabolic assay image showing Luminex bead antibody binding metabolic biomarker that is part of Luminex metabolic panel

Human Metabolic Luminex Panel

Flexible, premixed Metabolic Luminex panels can be configured to quantify up to 12 metabolic biomarkers per 50 uL sample in less than a day, helping to save time and precious samples while accelerating your workflow. Rigorous quality control processes help eliminate false-positive data. 

  • Measure up to 12 metabolic biomarkers simultaneously
  • 4.5-hour assay length, no overnight incubation
  • Small sample volume (50 uL)

 

Simple plex cartridge 32x8

Multiplex Assays on Ella

Simple Plex multiplex assays combined with Ella's speed and automated workflow allow you to create large datasets quickly with minimal manual intervention. The 32 sample multiplex cartridge offers your choice of either 5, 6, 7, or 8 immunoassays in duplicate from just 25 µL of sample.  

  • Reduce setup time with factory-calibrated standard curves
  • Get the same assay results no matter the cartridge format  
  • Maximize throughput with up to 256 reportable results in just 90 minutes
Webinar with tips for Utilizing RePlex Sequential Immunoassays and Total Protein Normalization on Simple Western

RePlex on Simple Western

With the RePlex™ features on Jess™ and Abby™, you can run two immunoassays within the same capillary to get all your rich protein characterization data from just one sample.

  • Quantify expressed phosphorylated target and total target levels
  • Normalize your data with total protein expression data in the same capillary
  • Gain greater immunoprobe selection flexibility
  • Get more data points per sample
  • Save time and money on consumables
Detection of anti-BCMA CAR-T cells, Inteferon-g expression by RNAscope multiplex fluorescence, and CD3+ T cells by immunofluorescence

RNAscope™ Multiplex Fluorescent V2 Assay

The RNAscope Multiplex Fluorescent v2 assay provides exceptional sensitivity while allowing simultaneous detection of up to four RNA targets within a single sample using up to four target probes.

  • Nuclei can be counter-stained and visualized using DAPI fluorescent dye 
  • Sequential assay workflow provides flexibility to visualize any probe in any of the TSA-linked fluorophore channels.

Multiplex Immunoassay Applications

Bio-techne can help you fast-track your discovery stages with automated and innovative solutions.

Immunology

Investigate and quantify immune responses using our wide range of multiplex assays and automated platforms that are specifically designed to simplify experiments and accelerate results.

iPSC Therapies – Cell and Gene Therapy Symposia Series

Cell and Gene Therapy

Rigorously characterize your mid-process and final cell products. Get your therapies to patients quickly and safely using our highly reliable and reproducible assays and analytical platforms. 

Cancer & Immuno-Oncology

Cancer Research

Identify and track your cancer biomarker candidates using our gold-standard analytical instruments and tools. Gain the superior throughput and high-quality data you need for reliable biomarker validation.

Other Multiplex Assay Options

Jess and Peggy Sue instruments

Gel-Free, Blot-Free Westerns

Capillary-Based Assays

With the capillary-based Simple Western™ platform, you can detect multiple proteins in the same capillary using two or more antibodies, enabling a variety of strategies for the detection of multiple proteins in each sample.  With Simple Western, you get rich and quantitative multiplexed protein characterization data with just 3 µL of sample, and it’s fully automated so you can design your next experiment while Simple Western does the work for you.

Single-Cell Analysis

Single-Cell Western on Milo measures protein expression in ~1000 single cells so you can quantify heterogeneity of your target at the single-cell level and detect up to 12 targets per cell with a variety of multiplex strategies.

Find out more about Simple Westerns.

highly sensitive molecular detection

Highly Sensitive Molecular Detection

RNA-ISH Assays

Recent advances in RNA in situ hybridization (ISH) assay technology provide exceptional sensitivity, allowing single-molecule detection of RNA targets at the single-cell level. RNA-ISH assays are ideal for simultaneous detection studies of any genes in nearly any tissue type using fluorescent labels. 

The assays are available with various levels of plexing, allowing you to:

  • Plex RNA and protein targets on the same slide for multi-omic analysis in situ’
  • Find cells of interest by using cell-specific markers
  • Confirm cells by simultaneous detection of additional targets
  • Characterize cellular heterogeneity of complex tissues  

Raising the Bar in Multiplex Performance

Deep multiplexing expertise

Deep Multiplexing Expertise

Multiplexing calls for carefully chosen compromises that allow analytes to play well together while still producing accurate, meaningful results. Bio-Techne has deep and broad-based experience developing and manufacturing immunoassays. Extensive technical expertise and rigid quality control help ensure we build the most reliable assays with superior precision, accuracy, sensitivity, and reproducibility.  

Unparalleled Assay Quality

Unparalleled Assay Quality

Confidence in multiplex immunoassay data hinges on superior accuracy and precision that is only possible from using high-quality raw materials. Bio-Techne antibodies, proteins, and specialized diluents are the foundation of all our immunoassays.  All of our assays and complete kits go through stringent quality control testing to make sure they remain consistent, and your data is reproducible.  

Antibody for Custom Assay Solutions

Custom Assay Solutions

Looking for a unique analyte or multiplex panel? Bio-Techne experts will work closely with you to produce multiplex assays that are tailored to fit your experimental needs. Custom assay services include new assay development, qualification, assay modification, and species testing. When your needs change, you can move seamlessly between platforms with consistency in your results.

Luminex bead based multiplex assays

What are the Advantages of Multiplex Assays?

Growing demand for immunoassays and surging testing volumes has put pressure on scientists to rein in costs and optimize sample usage by designing multiplex assays to cover several analytes simultaneously. Factor in the time saved by running the equivalent of 20 or more different assays in a single immunoassay workflow, and the benefits of multiplexing are clear.

The ability to measure multiple targets in a single assay allows researchers to detect hundreds of analytes in a single run while being judicious with precious sample volumes. From screening biomarkers for further evaluation to validating results for drug development, multiplex assays provide several important benefits, helping to:

  • Conserve limited sample volume
  • Lower your cost per data point
  • Improve accuracy due to less sample handling
  • Optimize productivity
  • Save time and resources

How Can Automation Help?

Some of the latest developments in multiplexing include new automated platforms designed to eliminate user error and accelerate time-to-results. Automation not only improves the efficiency of critical steps along the discovery continuum but also helps remove many potential sources of variability to provide more accurate and reproducible results.

Simple Plex™ assays running on the Ella™ platform are a good example, With Simple Plex, you get the same reproducible answers across multiple users and multiple sites, regardless of the user. The assay works like a traditional ELISA except everything is pre-loaded into the cartridge. You simply dilute your sample and insert the cartridge into Ella. Each assay is run the same way every time—with no manual steps, washes, or reagent additions. 

While different labs require different degrees of automation, the underlying need for greater precision, throughput, and reproducibility remains a core driver across research sectors.

Scientist Brian Astry does immunoassay research

Scientist  Mandy Kubik does immunoassay research

What Are the Key Features of the Best Multiplex Immunoassays?

Confidence in immunoassay data depends on the accuracy and precision that is only possible from using high-quality antibody reagents. This means selecting antibodies with not only high analyte affinity but also specificity, which is the ability to distinguish between the analyte and other structurally similar components.

While multiplexing offers numerous efficiency advantages, it also introduces several technical challenges that make assay design more complicated. One of the biggest challenges is controlling for cross-reactivity and interference between all the antibodies and proteins in the assay.  Do you get the same results from IL-6 when run alone and when with 20 other analytes? 

Single-analyte assays are designed to be analyte-specific—meaning that every component, from buffer to antibody pairs and more, are custom-tailored for that specific analyte. Assays incorporating multiple analytes do not have this luxury.

Multiplexing requires carefully integrated balances that allow analytes to work consistently together to produce consistent, accurate results. Interfering factors, no matter the mechanism, can result in either reduced or elevated signals relative to the actual concentration of target analyte.  

While validation protocols help ensure that antibodies are producing appropriate results, if the assay isn’t carefully developed and properly designed, achieving optimal performance will be difficult.

At Bio-Techne, we take great care to select the antibodies and proteins that can be plexed harmoniously to reduce interference and maximize performance across all analytes. Meanwhile, our rigorous quality control and validation efforts confirm the performance meets required standards and specifications lot after lot.

Ensuring Assay Precision and Accuracy

One challenge in multiplex assay design is creating a buffer system that is optimized to perform well with other analytes in the panel. Since it is not possible to customize the buffer to any individual analyte, optimizing buffers for peak performance, such as linearity or recovery, and managing matrix effects are key steps to ensuring sensitivity, precision, and accuracy of the resulting immunoassay.

There are primarily two reasons for dilutions. In some assays most samples read above the standard curve, thus requiring a dilution for analyte levels to fall within the range of the assay. A second reason for dilution is to limit interference due to factors in complex matrices.

In the best scenario, an assay diluent works well with a broad range of analytes, allowing scientists to multiplex anywhere from a few to several dozen analytes within a single test. To avoid incorrect dilutions in multiplex immunoassays, you need to know the relative abundance of the analytes to be tested.

Higher-abundance analytes may require a different sample dilution than those of lower abundance. Grouping the analytes by common dilution is the most efficient means of creating multiplex panels.

No matter what dilution samples are run at, you should always derive the same concentration once back calculated, giving you consistency in results. As you dilute the sample, the interfering factors also dilute out, making the target analyte more accessible, and the concentration of the target analyte increases.

Scientist Valeriu Bortnov does immunoassay research
Luminex High Performance Assay - Icon

Multiplex with Speed and Efficiency

While developing a multiplex assay presents several complexities compared to single analyte, plate-based ELISAs, the ability to generate high-quality data while dramatically reducing time and costs needed for testing provides researchers with a powerful advantage. From narrowing down drug candidates in preclinical studies to evaluating therapeutic responses during a clinical trial, multiplex assays provide several important benefits, including the ability to:

  • Maximize limited test samples: collect data from just 25μl or less of undiluted sample.
  • Minimize experimental variability: derive multiple data points from a single manipulation.
  • Optimize productivity: reduce sample preparation and processing time. 
  • Improve efficiency: examine multiple analytes in a single sample.

Meeting immunoassay performance goals is an ongoing challenge across research fields. The good news is that today’s scientists have access to an abundance of external experts who have deep technical experience and access to vast libraries of antibodies to help develop their assays. Engaging with an experienced supplier like Bio-Techne and paying attention to the technical factors outlined here will significantly improve your chances of developing high-quality, highly effective multiplex assays.