The Enzyme-Linked Immunosorbent Assay (ELISA) is one of the most sensitive immunological techniques that allow the detection of antigens or antibodies in the patient blood sample. The assay is performed in the microtitre plate and helps quantify and detect peptides, proteins, antibodies, and hormones. A target molecule is immobilized onto the polystyrene microplates, then combined with an enzyme-labelled antibody. The enzyme activity is measured by incubating it with a substrate to generate a quantifiable coloured product, then measured using an ELISA reader.
The assay is carried out in the following manner:
1. ELISA begins with a coating of a microtitre plate with a target antigen or antibody. Antigen or antibody molecule is adsorbed onto the polystyrene microtitre plate.
2. Following a series of washing steps, the unbound sites are blocked using a blocking agent.
3. The plate is incubated with enzyme conjugate antibodies after another round of washings.
4. All unbound antibodies are removed by washing several times and substrate to enzyme is added, producing the coloured reaction product.
5. The plate is read using an ELISA reader.
Washing steps are critical in ELISA assays for removing unbound material because any nonspecific binding can result in false-positive results.
Applications:
ELISA has several applications, e.g., diagnostics, food industry, vaccine development, immunology, toxicology, drug monitoring, pharmaceutical industry, and transplantation. Some of the applications are described as:
1. Diagnostics: The ELISA was the first screening test widely used for HIV because of its high sensitivity (Alhajj and Farhana, 2022). It helps detect autoantibodies and antibodies against infectious diseases (antibacterial, antiviral, antifungal). It also helps in screening the donated blood for possible viral contaminants. Detection of tumor markers, and hormone levels can be carried out using ELISA.
2. Food industry : ELISA has a vital role in the food industry. It helps identify the food allergens in milk, peanuts, walnuts, almonds, and eggs.
3. Vaccine development: ELISA has a significant advantage in vaccine development. It allows testing serum samples from immunized animals or humans for the presence of antibodies.
4. Pregnancy Test: ELISA can detect pregnancy-related proteins in pregnant women’s blood, saliva, or urine sample. This test is the first one prescribed by a gynaecologist to confirm the pregnancy. HCG is one of the most common hormones detected by ELISA in the first month of pregnancy.
5. Cancer Detection: Survival of a cancer patient depends on diagnosis at a very early stage.
Cancer biomarkers are complex biomolecular entities to target as analytes. ELISA is a promising technique for the identification of cancer biomarkers.
References:
• Enzyme Linked Immunosorbent Assay, StatPearls [Internet]. StatPearls Publishing. Alhajj, M., Farhana, A., 2022.
• Immunology__Janis_Kuby_5th Edition.pdf, n.d
• ELISA Principle, Procedure, Types, and Applications (https://blog.praxilabs.com/2021/09/20/elisa-principle/)
