Introduction

ELISA: is an abbreviation for enzyme linked immunosorbent assay

ELISA: is a biochemical technique used mainly in immunology to detect the presence of Ab or Ag in a sample also used as a diagnostic tool in medicine and

plant pathology quality control check in various industries

General steps and components

The main steps and components involved in ELISA running:

1. Solid phase: represented by microtiter plate which must have certain characteristics like

high level of adsorption of proteinious substances

the wells are flat in shape

2. Coating step: with Ag or Ab

3. Incubation: to allow the reaction between Ab and Ag substrate and enzyme etc…..

Less time the reaction is not complete More time affect Ab sensitivity

4. Washing: to remove excess Ab or Ag and remove non well attached Ab or Ag

Soft washing : the excess still in wells and bind conjugate and gives fouls results

Aggressive washing: affect binding of Ab and Ag

5. Concentration 1. Antibodies2. Antigen

6. Conjugates: are Ab of a certain spp against injected Ab from another spp and it is also called antispecies that is covalently bond to Enzyme

The main types of enzymes that can be used in this test are:

1. Horseradish preoxidase 2. Alkaline phosphatase3. β – Galactosidase4. Urease

7. Substrate “chromogene”: produce the color when react with the enzyme

8. Stopping solution: stop the reaction between the enzyme and the substrate

9. Washing buffer: used for washing step

10. Sample buffer: used for making dilution for the samples

Types of ELISA

The main methods of ELISA:

1. Direct ELISA

2. Indirect ELISA

3. Sandwich ELISA

4. competitive ELISA

Direct ELISA

Indirect ELISA

Direct sandwich ELISA

Indirect sandwich ELISA

Direct competitive ELISA for Ag

Direct Competitive ELISA for Ab