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Discover How Multiplex Assays are Advancing Biomedical Research

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Top tips to optimize your analyses

9 May 2016
Sarah Thomas
Associate Editor

Editorial article

Top tips to help you optimize your multiplex assays

Sonali Nayak, Senior Manager for North America Life Science Technical Services at MilliporeSigma, spoke to SelectScience® about the advantages and applications of multiplex assays. Read on to discover the latest technological developments and top tips for setting up your multiplex assays.

Q: Please tell us about your role at MilliporeSigma.

A: As the manager for protein detection, cell analysis and sample preparation, I support the products and tools developed for biomedical research, including the first commercially available multiplexed cytokine assay for the Luminex® platform. I also collaborate in data analysis software development for multiplexed assays. My experience encompasses many areas across drug discovery and development with an emphasis on research diagnostics.

Q: What are the benefits of multiplex assays?

A: Multiple biomarkers may need to be measured for research or diagnostic purposes and conventional methodologies, such as RIAs, ELISAs or Western blotting are constrained due to:

  • sample volumes required
  • costs
  • time for standardization, validation & assay runs
  • data reliability of running multiple assays on one sample

Multiplex assays address all these issues. Luminex’s xMAP® technology coupled with MILLIPLEX® Immunoassay kits, enables simultaneous measurement of multiple analytes, in low sample volumes, using one assay run. Fluorescent detection also provides a wider dynamic range for higher assay sensitivity. Costs are reduced as fewer assay runs are required.

Q: What are the most important applications of multiplex assays?

A: These assay formats have been used in several aspects of preclinical research and are now used in diagnostics. In drug development, multiplex assays can be applied at the discovery stage during screening, for biomarker profiling in preclinical tests, for final ADME studies and in phase IV clinical trials. In new-born screening, only one capillary tube of blood is sufficient to generate data across multiple parameters. Dried blood spots, collected worldwide, have been tested for biomarker research and epidemiology studies.

Multiplex assays, such as MILLIPLEX® MAP, are facilitating drug discovery screening, by reducing the time and resources necessary to develop a commercially viable drug.

Q: What are your top tips to get the best multiplex assay results?

A: Several factors should be considered for optimum results:

  • Cross Reactivity

The antibody pairs used in a multiplexed panel should have negligible cross reactivity to the other analytes in the panel.

  • Sample Dilution / Treatment

The sample values for the chosen assays should fall within the dynamic range of the standard curves. For example, high abundant proteins may require a sample dilution, in which case they cannot be plexed with low abundant protein assays. Similarly, analytes that require sample extraction (by either acidification, column separation etc.) may need to be plexed separately.

  • Matrix Compatibility

Some sample types may have interfering matrix effects that reduce the recovery of the analyte in the assay. The use of a compatible matrix in the standard curve and /or buffers can compensate for these interferences. However, as the effect is different for each assay, only those assays, that give similar accuracy and linearity can be plexed.

  • Assay Format

Some applications only require a qualitative response, i.e. presence, absence or change in analyte concentration. These assays may need to be run separately from quantitative assay, for example cell signaling studies versus cytokine profiling.

Q: What developments could benefit multiplex assays in the future?

A: We have the tools to generate data from genomic and proteomic microarrays separately but the ability to combine these parallel resources would greatly enhance research. Development of more sophisticated software, capable of quickly analyzing and interpreting the large volumes of data that can be generated from just one sample would also improve data management.

Do you use MILLIPLEX Immunoassay Kits? Tell us your opinion, leave a review today.

Image: diamant24/Shutterstock

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ELISAEnzyme-linked immunosorbent assays (ELISA), also known as enzyme immunoassays (EIA), are used for the detection and quantification of proteins, peptides and antibodies in a sample. ELISA often comes in kit format, with pre-selected antibody pairs to detect specific proteins or biomarkers of diseases such as diabetes and obesity, or cardiovascular and neurological disorders. Detection is made possible with chemiluminescent, fluorescent or colorimetric substrates and detection instruments such as microplate / ELISA readers. Other kits available include: Enzyme-linked immunospot (ELISPOT) assays for the quantification of protein-producing cells and chemiluminescent immunoassays (CLIA) kits. Find the best ELISA kits and products in our peer-reviewed product directory: compare products, check customer reviews and receive pricing direct from manufacturers.Cell-Based AssaysCell-based assays are used to monitor the presence, quantity and activities of a desired cellular analyte including drug molecules or biomarkers. This can reveal information on cell health (apoptosis, cytotoxicity, viability and proliferation assays), cell metabolism, cell migration and cell signaling mechanisms. Find the best cell-based assay products, kits and equipment with our peer reviewed product directory: compare products, check customer reviews and receiving pricing direct from manufacturers.BiomarkersBiomarkers are biological markers which can be measured and evaluated to indicate a biological state. The use of biomarkers in research and diagnosis can indicate a normal or disease state or drug response of cells / tissues. Biomarkers include genetic markers, cell surface markers such as antigens, antibodies or receptors and secreted molecules such as cytokines. An assay system is required for identification of biomarkers. :Clinical ChemistryBiochemistry (or clinical chemistry) involves the analysis of bodily fluids using chemical tests. Techniques used include HPLC, chromatography, spectroscopy, mass spectrometry, immunochemical, electrophoresis, turbidometric / spectrophotometric assay, MRI and ISE analysis. Tests are often carried out on plasma or serum but urine (urinalysis) and fecal specimens are also processed.ImmunoassayImmunoassays are laboratory tests that utilize antibody-antigen interactions to detect specific substances. They are widely used in diagnostics for detecting diseases, drug testing, and environmental monitoring. These assays offer rapid, sensitive, and precise detection of biomarkers, making them invaluable in clinical and research labs. Browse our peer-reviewed product directory to compare immunoassay systems, read user reviews, and get pricing directly from manufacturers.MultiplexProtein BiologyThe analysis of protein expression, identity and function is vital for many areas of life science research and drug discovery. Some of the most commonly used techniques in protein analysis include Western blotting, electrophoresis and mass spectrometry.Drug DiscoveryDrug discovery is the process of identifying potential new medications, involving stages such as target identification, compound screening, and preclinical development. It relies on cutting-edge technologies like high-throughput screening, artificial intelligence, and molecular modeling to accelerate the identification of drug candidates. Drug discovery plays a pivotal role in developing new therapies for diseases ranging from cancer to rare genetic disorders. Browse our peer-reviewed product directory to find the latest drug discovery technologies, compare options, check customer feedback, and get pricing directly from manufacturers.Cell BiologyCell biology studies the structure, function, and behavior of cells. Understanding cellular processes is essential for research in areas such as cancer biology, stem cell research, and neurobiology. Techniques like flow cytometry, microscopy, and cell culture enable researchers to explore cellular mechanisms in detail. Browse our peer-reviewed product directory to find the best cell biology tools and equipment, compare products, check customer reviews, and get pricing directly from manufacturers.Molecular DiagnosticsMolecular diagnostics use an individual’s genetic code and gene expression to diagnose and monitor diseases. The technique is used increasingly in the field of infectious diseases and oncology, as well as areas such as coagulation, HLA typing and pharmacogenomics. Molecular diagnostics plays a pivotal role in personalized medicine.Dried Blood SpotsDrug DevelopmentDrug development refers to the process of bringing a new drug to market.Biomarker DiscoveryClinical biomarkers refer to substances related to known medical conditions that can be accurately measured <i>in vitro</i>. Biomarkers can be used to diagnose presence of a disease and indicate disease severity. The discovery of new biomarkers is incredibly valuable in the field of diagnostics.

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