Enzyme-linked immunosorbent assays (ELISA) are popular among medical researchers and physicians as a quality control measure and diagnostic tool. Apart from that, ELISA assays are also frequently used as analytical tools in the realm of biomedical research. In this role, ELISA essentially helps detect and quantify specific antibodies and antigens in a given sample, for the discovery and testing of drugs. Diagnostics, drug discovery, and quality control are therefore three of the primary areas where ELISA assays are used during biomedical studies and research projects.

ELISA is derived from, and shares elements with, radioimmunoassay (RIA). RIA was first discovered and used by scientists in the 1960s to measure and quantify endogenous plasma insulin. Later, RIA was developed into a novel technique meant for the detection and measurement of biological molecules (such as peptides, hormones, and proteins) present in minuscule quantities in any given sample. Despite their usefulness, however, RIA assays ran into some trouble because of the safety concerns associated with the use of radioactivity in the biomedical sector.

A modification, which involved using an enzyme instead of a radioisotope for the RIA assays, was proposed, in order to minimize safety concerns and maximize the efficacy of the process. This minor modification to the erstwhile RIA resulted in the creation of the modern enzyme-linked immunosorbent assay or ELISA. While the RIA did have the benefit of being slightly more sensitive than the ELISA, the latter is preferred for being safer, more practical, and having better secondary antibody selectivity.

The General Principles of ELISA

A basic concept in immunology, of an antigen binding to its particular antibody, is used by the ELISA assay to detect minuscule quantities of antigens in a fluid sample. These antigens can include hormones, peptides, and proteins, all of which are essential components needed in the process of drug development.

Enzyme-labeled antibodies and antigens are used, during an ELISA assay, to detect biological molecules. Glucose oxidase and alkaline phosphatase are the two enzymes which are most commonly used for this purpose.

In the fluid phase, the antigen is immobilized within 96-well microtiter plates. Inside these plates, the antigen being used is bound to a specific antibody. This bound antibody is then subsequently detected and attached to a secondary, enzyme-coupled antibody. A visible color change, also described as fluorescence, is yielded by a chromogenic substrate for the enzyme, indicating the presence of one or more antigens.

Such colorimetric reading can facilitate the assessment of both qualitative and quantitative measures. Fluorogenic substrates are often used for their higher sensitivity, which confers them the ability to accurately measure antigen concentration levels in any given sample.

Types of ELISA Assays

Over the years, various types of ELISA assays have been developed and employed in the biomedical sector. The primary step in any ELISA assay is the direct or indirect detection of one or more antigens in the sample, achieved by immobilizing or attaching the antigen to the well surface. The antigen-specific capture antibody can also be immobilized and attached directly to the surface of the well for similar results.

For robust and sensitive measurements, the antigen should ideally be selected specifically out of a sample of mixed antigens, so that the test antigen is obtained directly from the serum being tested. Some of the different types of ELISA assays commonly used by researchers are:

Indirect ELISA

Sandwich ELISA

Competitive ELISA

Multiple and Portable ELISA

Advantages of ELISA Assays

Some of the primary benefits of an ELISA assay have been listed below for your convenience.

ELISA is essentially a simple and easy to perform biochemical assay that makes use of enzyme-mediated color change, as seen in antibodies, to detect the presence of antigens such as proteins, hormones, peptides, etc.)

The detection of antigens or antibodies, even when present in very low concentrations in any given sample, can be facilitated by the sandwich or indirect ELISA methods.

The competitive ELISA can help researchers detect compositional differences in complex and highly sensitive antigen mixtures, even when the antibody being detected is present only in trace amounts.

Multiple and portable ELISA assays are a low-cost, ready-to-use testing kit that can be used to screen large populations effectively, particularly in settings where resources are scarce or limited.

If the process of binding the antigen or antibody to the plate is conducted properly, then the chances of a falsely high-positive result in the ELISA assay are almost non-existent.

In Conclusion

Both as a tool for biochemical research and diagnostics, ELISA is a powerful option available to researchers and scientists wishing to gain accurate, reliable results in a cost-effective and timely manner. This is because ELISA assays can facilitate the detection of various different biological molecules at low concentration levels. Despite its limitations, therefore, ELISA remains an important tool in the realms of both clinical diagnostics and research.



