ELISA Technique: Types, Principles, Procedure & Advantages

ELISA Technique is a simple and efficient technique that is used to check the presence of antigens in the blood samples. It needs a solid stationary medium, a liquid medium, and a well plate to hold the solid stationary support. 

• ELISA stands for Enzyme-Linked Immunosorbent Assay.
•  It is used to determine the presence or concentration of antibodies, hormones, proteins, and peptides in the given sample.
• It relies on antibodies to check for antigens in the body.
• ELISA technique is used to detect the presence of ligands in a liquid sample.
• Assays are the basic procedures commonly used in chemical laboratories.
• It determines the sample's purity, constituents, environment, biology, etc.
• Another name for the ELISA technique is the EIA test.
• ELISA is most commonly used in plant pathology and biotechnology.

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Key Terms: ELISA Technique, Antigen, Antibodies, Blood Samples, Hormones, Proteins, Ligand, Peptides

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ELISA Technique Principle

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ELISA Technique works on the principle of antigen-antibody binding. The principle requires two primary components, an analyte or ligand which is needed to be estimated quantitively and qualitatively.

• It require a liquid or solid support media, on which the analyte is adsorbed.
• The solid support media may act as an antigen, or an antibody, depending on the type of the ELISA method used.
• This solid support is coated on the surface of the wells as a uniform layer.
• In ELISA Technique, the analyte to be estimated is converted into a liquid sample using appropriate solvents.
• This liquid mixture is then poured onto the solid support media.
• The mixture is stored in 96 welled plates.
• This solid and liquid mixture is made to interact for a suitable period of time.
• The analyte binds with the solid support and separates on filtration, or any other separating procedure.

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Types of ELISA Techniques

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ELISA Technique can be broadly divided into three branches, which are as follows:

Indirect ELISA Technique

In the Indirect ELISA Technique, the antigen is poured into the wells of the plate. It is attached to walls of microtitre plate.

Process for Indirect ELISA Tecnique

The process is as follows:

• The antibodies are added to the wells precoated with antigen.
• The mixture is allowed to stand for some time.
• During this time period, the antigen and antibodies react to form a complex.
• These antibodies are called primary antibodies.
• The non-bonded primary antibodies are washed away.
• The antigen-antibody complex is then treated with a secondary antibody-enzyme conjugation.
• This enzyme then displaces the antigen and binds with the primary antibodies.
• Finally, a substrate is added which gives the enzyme-primary antibody complex a blue color.

Sandwich ELISA Technique

In the Sandwich ELISA Technique, the antibody is poured into the wells of the plate. In this microtitre plate is coated with antibody.

Process for Sandwich ELISA Tecnique

The process is as follows:

• The antigens are added to the wells precoated with antibodies.
• The mixture is allowed to stand for some time.
• During this time period, the antigen and antibodies react to form a complex.
• These antibodies, pre-coated on the wells, are called primary antibodies.
• The antigen-antibody complex is then treated with a secondary antibody-enzyme conjugation.
• This secondary then displaces the primary antibodies and binds with the antigen,
• Finally, a substrate is added which gives the enzyme-antibody-antigen complex a blue color.

Competitive ELISA Technique

In the Competitive ELISA Technique, the antigen is poured into the wells of the plate, and the coated wells are then treated with an antigen-antibody mixture.

Process for Competitive ELISA Tecnique

The process is as follows:

• The antibody-antigen complex is added to the wells precoated with antigen.
• The mixture is allowed to stand for some time.
• During this time period, the antigen and antibodies react to form a complex.
• These antibodies are called primary antibodies.
• The non-bonded primary antibodies are washed away.
• The antigen-antibody complex is then treated with a secondary antibody-enzyme conjugation.
• This enzyme then displaces the antigen and binds with the primary antibodies.
• Finally, a substrate is added which gives the enzyme-primary antibody complex a blue color.

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The table below showcase the difference between types of ELISA Techniques on different parameters along with the procedure followed for each type.

ELISA Experiment

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Material Required for ELISA Technique

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The material required for ELISA Techniques are as follows:

• Coating Wells:  In this process, 96 or higher well plates are used.
• Analyte to be Evaluated: It will evaluate the antigen, antibodies, peptides, and proteins
• Supporting Media: It includes polysaccharides, antigens, antibodies, etc.
• Colouring Agent: It is responsible for imparting the solution a blue color.
• Spectrophotometer: It is used to detect the concentration works on the principle of absorption of light rays by the sample.

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Advantages of ELISA Technique

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The advantages of conducting an ELISA technique is as follows:

• It accurately detects a particular disease.
• ELISA Technique is a type of quick test whose results are available quickly.
• The test can easily be carried out for complex samples.
• ELISA Technique uses direct and indirect methods to give highly responsive results.

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Applications of ELISA Technique

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The various applications of ELISA Technique are as follows:

• ELISA Technique can be used to track the outspread of diseases in communities.
• It can easily determine whether an individual require a vaccine or not.
• The technique can easily detect for food allergies.
• ELISA Technique can easily be used to estimate hormone level in the body.

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Things to Remember

• ELISA is abbreviated as Enzyme-Linked Immunosorbent Assay.
• It is one of the most common assay techniques used in science.
• ELISA Technique checks for the presence of antibodies, hormones, proteins, and peptides.
• It is broadly divided into three types, depending on their functioning.
• Direct ELISA, Indirect ELISA, and Sandwich ELISA are three types of Elisa Technique.
• ELISA is used to detect an antibody to HIV by using indirect or sandwich ELISA. 
• It also can detect an antigen for human chorionic gonadotropin hormone (HCG).
• The component is excreted in the urine of a pregnant woman by using direct ELISA.

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