ELISA, enzyme-linked immunosorbent assay, is a technique that helps quantify and detect solution substances, like antibodies, protein, peptides, and hormones. The plate-based technique is known with many other names in the industry, such as enzyme immunoassay or EIA.
In this technique, we immobilize the target macromolecule (referred to as antigen) on the microplate (or the solid surface). A complex is formed with an antibody, which is connected to the reporter enzyme used in the experiment. To detect the outcome, we measure reporter enzyme activity by incubating the plate with the proper substrate, which helps generate a measurable product or result. An important phase of ELISA assay is antigen-antibody interaction.
Experts believe ELISA assay to be a powerful technique used for quantifying any given protein inside a mixture. It was originally designed by Engvall and Perlmann back in 1971. Since then, the technique has become popular for identifying and quantifying protein with the help of specific antibodies.
Usually, an ELISA assay is performed in a 384-well or 96-well polystyrene plate to link proteins and antibodies. This immobilization and binding of reagents through the ELISA assay makes the technique simple yet efficient to perform.
By immobilizing the ELISA reactants on the surface of a microplate, it becomes simpler to separate non-bound and bound material at the time of the test. We can utilize antibodies with high affinity and wash off non-bound material while using the technique, which makes it an extremely valuable tool to measure a given analyte from a crude mixture.
What are the types of ELISA assay?
- The method that was originally developed is Direct ELISA. It immobilizes antigen on a microtiter plate, where enzyme wrapped antibodies react with substrates to offer results in the form of color development.
- The second method used for ELISA assay is Indirect ELISA. It involves two antibodies, where the primary one is antigen-coated and added to the microtiter. Then, a secondary antibody (with enzyme) is added to the bound antigen-antibody.
- The third method is Sandwich ELISA assay, which is widely utilized to detect macromolecules or toxins. In Sandwich ELISA too, there are two antibodies. The first one is coated on a multi-well, which allows antigen immobilization. The secondary antibody helps with antigen detection.
- The last type is the Competitive ELISA assay. In this, the primary antibody creates a complex from the sample. Once this complex settles, we add a secondary antibody to the complex. But, remember that we would be able to detect primary antibodies if and only if there’s no binding of antigen. Hence, in this case, antigen and the secondary antibody competes.
Advantage Of Using ELISA Assay
ELISA assay test is beneficial in various use cases. For example, we use Indirect ELISA to find HIV infection in an individual and Sandwich ELISA to find toxins.
Find some benefits of using ELISA assay:
ELISA assays are highly sensitive, as it uses enzyme for testing. We already understand that an enzyme can be used to create multiple catalytic reactions for chromogenic reactions. This is possible because enzymes are widely known organic catalysts that amplify the results. We are using only a trace of antibody or antigen, sometimes in just a nanogram or microgram, but we still receive results for antibody quantification.
We have the power to automate ELISA assay tests, which means that the entire technique is cost-effective. Further, it is possible to use one ELISA assay for around 96 tests at one time. Here, we are saving time, money, and effort. This benefit also makes ELISA assay non-challenging and hassle-free.
ELISA assay is simple to perform if we compare it with other immunoassay techniques. For this reason, ELISA has a wide range of applications. For example, we use the technique to identify the infection markers in the blood, such as in HIV. In many regions and countries, using blood screening is mandatory, as we can’t send infected blood for transfusion and other health functions.
You may find applications of ELISA assay in the food industry as well. For example, knowing the toxicology or allergen profile for different drug categories.
In ELISA assay, we also receive strong specificity. This is because of the antigen or antibody selection. This is to say that antigen or antibody binding can only happen inside the binding site of the given antibody or antigen epitope.
We know a complementary relationship forms between antigen-binding site and epitope of the given antibody for both spatial configuration and chemical structure. For this reason, the reaction happening between antibody and antigen is highly specific.
We don’t need to pre-treat the sample for the ELISA test or go through other complex procedures to use it for simultaneous analysis. This is the reason why we receive better efficiency in the case of the ELISA test.
ELISA tests are found to be extremely environment-friendly or eco-friendly. The technique doesn’t involve using a large amount of organic solvent or radioactive substances.
When compared to different immunoassay methods, ELISA assay is more specific and accurate. This means that you can use this when you need highly specific, sensitive, favorable, and strong results. Check out more details of the ELISA test method above and understand its uses and benefits.