How Do Immunoassay Tests Work


Immunoassay Tests are biochemical tests that detect disease and works on the principle of antigen-antibody reaction. It aims at measuring the presence and the concentration of an analyte. Researchers and health care professionals utilize immunoassays to detect different kinds of antibodies. 

Surmodics™ IVD provides immunoassay developers the critical components they need to build reliable, robust, and accurate immunoassay tests. Our broad portfolio includes the industry’s gold standard in protein stabilizers/blocking buffers designed to increase sensitivity and specificity while decreasing non-specific binding. We also offer best-in-class detection substrates for color (TMB, ABTS, PNPS, BCIB/NBT) and light (Chemiluminescent) development as well as a selection of accessory reagents including our high-performance diluents designed to reduce matrix interferences including Human Anti-Mouse Antibodies (HAMA) and Rheumatoid Factor (RF) interferences.

We also manufacture and supply a line of microarray slides and surface technology as well as supply high quality antigens and antibodies from our partner company, DIARECT™ located in Germany.

The Working Mechanism of Immunoassay Tests

Immunoassay tests work by using antibodies to detect small biological substances in the blood and other bodily fluids. The method follows the aspect that particular antigens binds to specific newly introduced antibodies, thereby stimulating an immune response. This refers to specificity, as antibodies are highly specific to analytes such as hormones, nucleoproteins, and peptides, among others. Provided the immunoassay used reagents are pure, detection of analytes becomes successful through the formation of antigen-antibody complexes. A color change occurs, which indicates the presence of the analyte in question. The observed color symbolizes the amount of targeted molecule present in the test solution.

ELISA Immunoassays

For an enzyme-linked immunosorbent assay, an ELISA reader is employed to measure the color contrast on the ELISA plate. The microplate reader uses the principles of spectrophotometry, fluorescence, and luminescence. It measures the amount of light absorbed, reflected, or emitted by an analyte at one wavelength. ELISA plates and tube kits contain known concentrations of the target analyte and negative control, which are the standards of interpretation. For the semi-quantitative reading, the ELISA reader focuses on the different shades of blue exhibited by the set standards. Alternatively, one can use a quantitative interpretation. This involves adding microwells to a plate reader to measure the optical densities of all the samples and standards simultaneously.

Surmodics IVD provides assay developers a one-stop-shop for immunoassay development, including the components needed for development of ELISA’s. This can be seen in our full line of dried protein stabilizers and blockers, sample or assay diluents, in-solution protein stabilizers, ELISA substrates, stop solutions and accessory reagents, and supply of DIARECT antigens and antibodies. 

Please review some of the key technical advantages of each of our immunoassay reagents. 

  • Dried Protein Stabilizers/Blockers (Immunoassay Stabilizers):
    • Surmodics IVD’s StabilCoat™, StabilBlock™ and StabilGuard™ Immunoassay Stabilizers are formulated to minimize non-specific binding interactions with the surface and stabilize the dried capture protein over time.
    • These reagents offer assay developers the gold standard for stability and blocking efficacy. They provide improved assay performance in a one-step process for streamlined manufacturing.
    • Learn More About Dried Protein Stabilizers/Blockers
  • Sample/Assay Diluents:
    • Surmodics IVD’s StabilZyme Stabilizers offer assay developers the gold standard for stability of protein conjugates at working strength concentration for ELISA/EIA, ELISpot, RIA and immunoblot/western blot applications.
    • Learn More About Sample/Assay Diluents
  • ELISA Substrates:
    • The signal generation step of immunoassay systems is critical to obtaining an accurate measurement of the target protein. Surmodics’ portfolio of BioFX™ colorimetric and chemiluminescent substrates offers the stability, low background and sensitivity needed to meet the demands of assay manufacturers.
    • Learn More About ELISA Substrates
  • Stop Solutions:
    • Gold Standard BioFX stop solutions for colorimetric substrates are available as dry blends or in liquid formulations including our Nova-Stop solution that provides minimal drift and a safer alternative to assay developers as it is non-corrosive to both skin and eyes.
    • Learn More About Stop Solutions
  • DIARECT Antigens and Antibodies:
    • DIARECT recombinant and native antigens have a proven track record for quality and sensitivity in ELISA, Western Blot and lateral flow applications. The lot-to-lot consistency of DIARECT antigens reduces development time and increases productivity of immunoassay developers.
    • Learn More About Antigens and Antibodies

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Chemiluminescent Immunoassay

This is an immunoassay that involves using chemiluminescent substances and an antigen sample to detect molecular material. It works on the basic principle of antigen-antibody specificity. The enzymes are used to convert a substrate to a different product, thereby leading to the emission of light of a specific color. The luminescence in an indication of the presence of an antigen. 

Chemiluminescent substrates in particular provide a higher degree of sensitivity than other substrates to enable lower limits of detection to be achieved in ELISA and membrane assays. The increase in sensitivity requires more vigilance during the optimization process through the proper selection of the substrate, antibody/antigen titers and blockers.

Please review the chemiluminescent reagents we offer below: 

Please review the list of our reagents below that can be used in lateral flow applications. 

  • Dried Protein Stabilizers and Blockers: 
    • StabilBlock (ST01), StabilGuard (SG01), StabilCoat (SC01)
      • Formulated to minimize non-specific binding interactions with the surface and stabilize the dried capture protein over time. StabilBlock® was recently developed to be the strongest blocker and optimized to improve overall assay sensitivity.
      • Learn More
  • Sample/Assay Diluents: 
    • Surmodics Assay Diluent Protein-Free (SM01)
      • Universal blocking agent that utilizes synthetic components to reduce matrix interference within immunoassay applications. The product was designed to be used within the existing assay protocol to proactively block interferences using a non-specific blocking mechanism. 
    • MatrixGuard™ Diluent (SM02)
      • This reagent is designed to be used within an existing assay protocol to proactively remove matrix interferences within discordant samples, while maintaining or even increasing signal-to-noise ratios.
    • Learn More 
  • Membrane/Precipitating Substrates: 
    • BCIP/NBT Blue One Component AP Membrane Substrate (BCIB) 
      • The BCIP/NBT substrate provides a dark blue precipitate upon reaction with alkaline phosphatase.
    • BCIP/NBT Purple One Component AP Membrane Substrate (BCID)
      • The BCIP/NBT substrate provides a dark purple precipitate upon reaction with alkaline phosphatase.
    • TMB Enhanced HRP Membrane Substrate (ESPM) 
      • BioFX TMB Enhanced HRP Membrane Substrate provides an intense staining pattern for increased sensitivity.
    • TMB One Component HRP Membrane Substrate (TMBM) 
      • BioFX TMB Super Slow Kinetic One Component HRP Microwell Substrate is a low-kinetic range substrate for ELISA applications.
    • DAB Membrane Substrate (DABM) 
      • BioFX DAB HRP Membrane Substrate is a two-component substrate system.
    • Learn More

Applications of Immunoassay Lateral Flow Devices: 

Clinical Analysis

Similar to other forms of immunoassays, a lateral flow device uses antibodies to detect antigens in a blood sample, serum, cells, tissues, urine and other clinical and biological samples. Common types of analytes tested include Thrombin, Alpha fetoproteins, HBs antigens, and pharmaceuticals, to mention a few. 

Protein Detection

Proteins are primary biomarkers in clinical analysis. Numerous proteins are linked to diseases such as cancer, HIV, Sickle Cell Anemia, Hepatitis, and so forth. 

Assay Blood Assessments

Assay blood test immunoassays are analysis carried out to determine the amount of analytes in a blood sample. These may be proteins, hormones, and drugs to mention a few. Types of assay blood tests include; blood glucose, calcium, cardiac enzymes, lipids, folate, erythrocyte sedimentation rate, Hcg, magnesium, and oestrogen examinations.