Enzyme-linked immunosorbent assay (ELISA) is a technique that uses enzymes such as β-galactosidase and alkaline phosphatase to detect antigen-antibody reactions in a sample. The technique is used in blood screening, microbiology, and veterinary screening. Additionally, it’s used as a quality control check in the food industries and as a diagnostic tool in medicine and plant studies.
ELISA is used for quantitative and qualitative analysis of a sample’s hormones, metabolites, proteins, and peptides. The process involves using specific antibodies that are immobilized to a solid surface, such as a microtiter plate. The sample is then added to the plate and incubated with the immobilized antibodies. The ELISA reader then analyzes the data to quantify the antigens and antibodies in the sample.
This article discusses the advantages of ELISA and why it should be a preferred method over other immunoassay methods.
ELISA follows the principle of the antibody-antigen reaction, which is highly specific. When an antibody and antigen are brought together, they bind to each other in a lock-and-key-like manner, forming an antigen-antibody complex. The binding between antibodies and antigens occurs in the epitope of the binding area in the antibody. This interaction is highly specific, with a chemical and spatial structure between the antigen binding site and the epitope.
ELISA takes advantage of this specificity by using antibodies specific to a particular antigen. In a typical ELISA, a sample containing the antigen of interest is added to a well-coated antibody specific to that antigen.
After washing away any unbound antigen, a detection antibody also specific to the antigen is added to the well. This detection antibody is usually conjugated with an enzyme, which catalyzes a reaction that produces a detectable signal, such as a color change.
Additionally, the components of an ELISA provide high specificity through negative controls. Negative controls don’t contain the antigen of interest. They ensure the test is not producing non-specific bindings or false positive results. Any background signal generated from the non-specific binding is subtracted from the signal generated.
One of the critical parameters of an ELISA is its sensitivity, which refers to the ability of the assay to detect low levels of the target molecule. Sensitivity in ELISAs can vary widely by the type of ELISA. In a competitive ELISA, the target antigen competes with a labeled antigen for binding to a limited amount of antibody.
Hence, the amount of labeled antigen that binds to the antibody is inversely proportional to the target antigen concentration in the sample. The labeled antigen is usually a smaller molecule than the target antigen. Therefore, it can be detected more efficiently, allowing for the detection of low molecular weight antigens with high sensitivity. This method can quantitate molecules with less than 10,000 Daltons in molecular weight because they’re immunogenic.
High sensitivity in ELISAs is a desirable parameter in immunoassays for several reasons.
First, it allows for detecting low levels of the target molecule. These levels can be important in many applications, such as detecting the early stages of the disease or monitoring the effectiveness of treatment. Second, high sensitivity can improve the accuracy and precision of the assay by reducing the likelihood of false negatives or positives. Finally, high sensitivity can increase the assay’s dynamic range, allowing for detecting a more comprehensive range of target molecule concentrations.
The ELISA procedure is excellent in reducing wastage. It uses microplates designed to handle small volumes of reagents and samples, reducing waste generated during the assay. Additionally, ELISA kits often come with pre-made reagents, eliminating the need for researchers to prepare reagents independently and reducing the chance of reagent waste.
ELISA also uses non-toxic reagents that are not harmful to the environment. For instance, the most commonly used ELISA substrate, 3,3’,5,5’-tetramethylbenzidine (TMB), is not harmful to the environment. The color change it produces during the assay poses no risk to human health or the environment. It doesn’t use radioactive components and high solvent solutions, making it a safe option in laboratory procedures.
Finally, ELISA does not require specialized equipment or instrumentation. The assay can be performed using essential laboratory equipment such as a microplate reader. This reduces the amount of energy required to run the assay and minimizes the carbon footprint of the experiment.
ELISA is a susceptible and specific procedure with many advantages over other immunoassay methods. It is eco-friendly, guarantees high specificity and sensitivity, and is cost-effective. Generally, using this method is highly effective in quality control checks, medicine and plan studies, and other medical procedures.
Having completed my Bachelor’s degree in medicine and currently pursuing a house job at a well reputed hospital in California, I decided to utilize my spare time in sharing knowledge with others through my blog. Apart from my time spent in the medical field, I love to read fiction novels and go on long drives.