Enzyme-Linked Immunosorbent Assay (ELISA) An Overview

Introduction

Enzyme-Linked Immunosorbent Assay (ELISA) is a widely used biochemical technique for detecting and quantifying specific proteins, antigens, and antibodies. It is crucial in medical diagnostics, immunology, and research due to its high specificity and sensitivity.

Principles of ELISA

ELISA relies on antigen-antibody interactions, where an enzyme-linked antigen or antibody produces a measurable signal. Key steps include:

1. Coating – Immobilizing the antigen or antibody onto a microplate.
2. Blocking – Preventing non-specific binding.
3. Detection – Using enzyme-linked antibodies or antigens.
4. Substrate Addition – Producing a measurable signal.
5. Measurement – Optical density (OD) detection using a spectrophotometer.

Types of ELISA

1. Direct ELISA – Uses an enzyme-conjugated primary antibody.
2. Indirect ELISA – Employs a secondary antibody for signal amplification.
3. Sandwich ELISA – Captures and detects antigens with a pair of antibodies.
4. Competitive ELISA – Measures antigen-antibody binding competition.

Applications of ELISA

1. Medical Diagnostics

• Detection of infectious diseases (e.g., HIV, COVID-19, Hepatitis B & C).
• Identification of autoimmune disorders (e.g., rheumatoid arthritis).
• Cancer biomarker screening (e.g., PSA, CA 125, CEA).
• Allergy testing (IgE detection).

2. Food & Environmental Safety

• Identifying pathogens (e.g., Salmonella, E. coli).
• Detecting allergens in food.
• Screening for pesticides and toxins.

3. Pharmaceutical & Research

• Drug discovery and vaccine evaluation.
• Protein quantification in biological samples.

Advantages and Limitations

Advantages

• High specificity and sensitivity.
• Rapid and cost-effective.
• Scalable for high-throughput screening.

Limitations

• Potential for cross-reactivity.
• Requires specialized equipment.
• Dependent on antibody quality.

Advances in ELISA Technology

1. Multiplex ELISA – Simultaneous detection of multiple analytes.
2. Point-of-Care ELISA – Rapid, portable diagnostics.
3. Digital ELISA – Ultra-sensitive molecular counting.
4. Nanotechnology-Enhanced ELISA – Improved signal amplification.

Keywords

ELISA, Immunoassay, Antigen-antibody interaction, Diagnostics, Protein detection, Biosensors, Optical density measurement.

References

1. Engvall E, Perlmann P. Enzyme-linked immunosorbent assay (ELISA): Quantitative assay of immunoglobulin G. Immunochemistry. 1971;8(9):871-874. https://doi.org/10.1016/0019-2791(71)90454-X
2. Crowther JR. The ELISA Guidebook. Humana Press; 2009. https://doi.org/10.1007/978-1-59745-240-3
3. Lequin RM. Enzyme immunoassay (EIA)/enzyme-linked immunosorbent assay (ELISA). Clinical Chemistry. 2005;51(12):2415-2418. https://doi.org/10.1373/clinchem.2005.051532
4. Wild D. The Immunoassay Handbook. Elsevier; 2013. https://doi.org/10.1016/B978-0-08-097037-0.00001-1
5. Kaushik A, Jayant RD, Vashist A, Nair M. Nanotechnology-based immunosensors and rapid detection techniques for COVID-19. Biosensors and Bioelectronics. 2020;165:112-287. https://doi.org/10.1016/j.bios.2020.112287

This document provides a concise yet detailed overview of ELISA, its principles, applications, advantages, limitations, and recent advancements.