This document provides an introduction to enzyme-linked immunosorbent assay (ELISA), covering its basic principles, procedures, advantages, and applications. ELISA is an antibody-antigen based assay developed in 1971 to detect the presence and quantify concentrations of antigens, antibodies, and other proteins. It works by utilizing antibodies and enzyme-linked substrates to produce a detectable color change in direct proportion to the presence of an antigen or antibody in a sample. Key advantages of ELISA include its speed, sensitivity, specificity, ability to process many samples at once using small sample sizes, and production of easy-to-read colorimetric results. ELISA kits now exist for detecting a wide range of targets like viruses, hormones, toxins, allerg

1. Enzyme Linked Immunosorbent Assay
(ELISA)
Applied Food Safety Education Laboratory Course III
NFS 492/592
Hi-Yung Hsueh, Extension Graduate Assistant
Joan Hegerfeld-Baker, Extension Food Safety Specialist
July, 2011

2. Introduction of ELISA
• Introduced by Engvall and Pearlmann in 1971.
• Main points covered in Lecture
– Basic principles of ELISA
– Advantages of using ELISA
– Uses of ELISA widely used

3. Basic Principles of ELISA
• Based on Basic Immunology Response
• Lock and Key Concept
– Antigen (key): substance when introduced into the body produces
antibodies
– Antibody (lock): protein in the body that is used by immune system to
identify and neutralize foreign targets (referred to as antigens)
– Key fits into the lock
• Enzyme conjugate substrates
– Enzyme that converts colorless substrates to a colored product
– Bound to the antibody that is part of the antibody-antigen complex
Or
– Bound to a secondary antibody that binds with the antibody-antigen
complex

4. Antibody-Antigen Complex

5. Conducting an ELISA test
Measuring amount of Antibodies in solution:
– Antigens (Ag) fixed to a solid surface
• (immobilized)
• 96-wells plastic plate
– Antibodies (Ab) in solution to be tested
• i.e. urine or serum
– Enzyme-conjugated Anti-immunoglobulin
• Antibody against the antibodies being tested for.
• Enzyme linked (conjugated) to the Ab
– Substrate – binds to enzyme and produces color
– Color intensity proportional to bound enzyme-Ab

6. Conducting ELISA (cont.)
• Quantitative & Qualitative Measurements
• Four common ELISA tests – based on the
binding structure between the Antibody and
Antigen.
1. Direct-ELISA
2. Indirect-ELISA
3. Sandwich-ELISA
4. Competition-ELISA

7. Common ELISA tests
• Direct-ELISA
– Enzyme conjugated Ab is directly bound to the Ag
• Indirect-ELISA

8. Common ELISA tests (contd.)
• Sandwich-ELISA
• Competition-ELISA
Competition between two antibodies for one antigen, or
two antigens for one antibody.

9. Advantages of ELISA
• Fast—90 samples tested in 2-3 hr
• Sensitivity (up to 10 pg/mL)
• Specificity (sample with high concentration contaminants
• Many samples can be processed at once
• Small sample size required (10 μL ~ 100μL )
• Colorimetric results – easily observed and measured
(spectrophotometer)
• Test for presence of Ag or Ab
• Flexible usage for research design
• Easy to learn, simple procedure

10. Hundreds of ELISA Kits Manufactured
• Viral Contamination
• Hormone levels
• Infections
• Specific disease factors
• Drugs
• Allergens in food
• Residues in food
• Toxins in food
• Others

11. End of part I