Last Updated on
February 8, 2023
By
Excedr
Enzyme-linked immunosorbent assay (ELISA) antibodies are crucial elements of the ELISA assay that are used to study proteins. Before proceeding to understand the antibody roles, it’s essential to learn more about the ELISA test.
ELISA is one of the most frequently used assays in labs for the detection and quantitative analysis of soluble substances, such as peptides, antibodies, and hormones. It’s also known as enzyme immunoassay (EIA).
The ELISA assay works on the principle of specific antibody and antigen interaction. It consists of the immobilization of antigen on a solid surface and then complexing it with an antibody attached to a reporter enzyme. Then, using different detection methods, the reporter enzyme activity is measured via incubation with the appropriate substrate.
Though, different types or formats of ELISA assay have been developed by today, which include:
But, they all follow the same basic steps, consisting of:
The analysis of ELISA reactions is mainly possible because of the power interaction between antigens and antibodies. And, this article expands more on ELISA antibodies, their functions, roles in the ELISA method, and applications in industries.
ELISA antibodies are one of the main heroes of this method. The antibody pairs with the specific antigen and facilitates the study of specific proteins of molecules in different samples.
By definition, antibodies are proteins that provide organisms immunity against foreign invaders, such as bacteria and viruses (known as antigens). The antibodies are also known as immunoglobulin (Ig).
There are two light chains and two heavy chains in each of the four polypeptides of antibodies. In ELISA, antibodies are classified based on whether they bind directly to antigens or another antibody:
These antibodies are also known as matched antibody pairs. To reduce the background noise during the ELISA reaction and produce effective results, it’s recommended to use the dilutions of antibodies.
Today, many companies offer ready-to-use ELISA kits with suitable reagents for the detection of a variety of molecules, such as chemokines, cytokines, and growth factors.
A primary antibody binds directly to the specific antigen, which is immobilized on the surface of the well plate.
To produce primary antibodies, one host species is required, such as a rabbit, mouse, goat, or chicken. In the first step of production, the host will be immunized against an antigen, producing a primary antibody.
A secondary antibody binds to an antibody-target protein complex. It’s usually labeled and helps in the detection of target antigens. Specifically, this type of assay is crucial for analyzing the number of specific proteins present in small amounts by signal amplification.
A secondary antibody is produced by injecting a primary antibody into another species of host. For example, if the primary antibody is generated in a mouse, the secondary antibodies can be generated in a goat, a rabbit, or a chicken.
Both the primary and secondary antibodies can either be monoclonal or polyclonal antibodies.
Monoclonal antibodies are a homogenous population of antibodies that are produced from identical B-cell clones, generated in a single parent cell. It interacts with only a single specific epitope of a specific antigen.
Polyclonal antibodies are a heterogenous population of antibodies that are produced in different clones of B-cells in the body. They can identify and bind to a variety of epitopes on a single antigen.
Immunoassays are powerful techniques used in life sciences labs to study proteins, measure their amount in a given sample, and analyze their functions and localization in organisms. They work on the principle of specific antigen-antibody interaction.
Two immunoassays that detect the antigen-antibody interaction include:
ELISA antibodies have extensive usage in Immunology and Life Sciences labs. Some of the applications are listed below:
Additionally, the primary antibodies also have applications in the distribution, metabolism, absorption, and excretion of a variety of therapeutic agents.
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ELISA antibodies are proteins involved in the immunoassays to facilitate the studies of proteins, peptides, and hormones in labs. They are two types: primary antibody, which directly binds to the antigen of interest; and secondary antibody, which is conjugated to an enzyme and binds to the primary antibody.
Both, primary and secondary antibodies, can be either monoclonal (binding to one specific epitope of the target antigen) or polyclonal (having the affinity for the same antigen but different epitope) antibodies.
Working with these molecules in high-throughput workflows requires researchers to work with high-quality lab-grade reagents and high-tech equipment. For labs that are just starting out, this might prove too expensive and cause them financial hardship.
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