ELISA Principle And Procedure Presentation

Introduction to ELISA Principle and Procedure
ELISA stands for Enzyme-Linked Immunosorbent Assay. It is a widely used technique in the field of immunology and biochemistry. ELISA is used to detect and quantify specific proteins or antibodies in a sample.	Image Source
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Basic Principle of ELISA
ELISA is based on the specific binding of an antigen and an antibody. The antigen of interest is immobilized on a solid surface, such as a microplate. The immobilized antigen will bind to specific antibodies present in the sample.	Image Source
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Types of ELISA
Direct ELISA: The antigen is immobilized, and a labeled antibody directly detects it. Indirect ELISA: The antigen is immobilized, and a primary antibody binds to it. A secondary labeled antibody then detects the primary antibody. Sandwich ELISA: The antigen is captured between two antibodies - a capture antibody and a detection antibody - both specific to the antigen.	Image Source
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Steps of ELISA - Coating
The first step is to immobilize the antigen on a solid surface, typically a microplate well. The antigen is diluted in a buffer and added to the well. The plate is then incubated to allow the antigen to bind to the surface.	Image Source
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Steps of ELISA - Blocking
After coating, the plate is washed to remove any unbound antigen. The next step is to block any remaining uncoated surface to prevent nonspecific binding. A blocking agent, such as BSA or milk, is added to the wells and incubated.	Image Source
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Steps of ELISA - Primary Antibody
The primary antibody, specific to the antigen, is added to the wells. The primary antibody will bind to the antigen present on the coated surface. Incubation allows for the formation of antibody-antigen complexes.	Image Source
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Steps of ELISA - Secondary Antibody
If using an indirect or sandwich ELISA, a secondary antibody is added. The secondary antibody is specific to the primary antibody and is labeled with an enzyme. The secondary antibody will bind to the primary antibody-antigen complex.	Image Source
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Steps of ELISA - Enzyme Substrate
After incubation with the secondary antibody, the plate is washed again. A substrate specific to the enzyme label is added. The enzyme catalyzes a reaction, producing a detectable signal, such as a color change.	Image Source
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Steps of ELISA - Signal Detection
The signal generated by the enzyme-substrate reaction is measured using a spectrophotometer. The intensity of the signal is proportional to the amount of antigen or antibody present in the sample. A standard curve is used to quantify the unknown sample based on the signal intensity.	Image Source
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Applications of ELISA
ELISA is used in medical diagnostics to detect diseases, such as HIV or COVID-19. It is also used in research to study protein-protein interactions and antibody production. ELISA has applications in agriculture, environmental monitoring, and food safety testing.	Image Source
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Conclusion
ELISA is a versatile and widely used technique to detect and quantify proteins or antibodies. Understanding the principle and following the procedure are crucial for accurate results. The development of ELISA has revolutionized various fields, enabling rapid and sensitive detection of target molecules.	Image Source
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