ELISA, or Enzyme-Linked Immunosorbent Assay, is a laboratory technique used to detect and quantify substances like antibodies, antigens, proteins, and hormones in a sample.
How ELISA Works: A Breakdown
The ELISA method relies on the principle of antigen-antibody interactions and enzyme-linked detection. Here's a simplified overview of the process:
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Coating: A microplate well is coated with a target antigen or antibody.
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Blocking: A blocking buffer is added to prevent non-specific binding of other molecules to the plate.
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Sample Addition: The sample being tested (e.g., blood, serum, urine) is added to the well. If the target substance is present, it will bind to the antigen or antibody already on the plate.
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Enzyme-Linked Antibody Addition: An enzyme-linked antibody, specific to the target substance, is added. This antibody binds to the target substance, forming a complex.
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Washing: The well is washed to remove any unbound antibodies.
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Substrate Addition: A substrate is added that the enzyme can act upon, producing a detectable signal (usually a color change).
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Detection: The intensity of the signal is measured using a spectrophotometer. The intensity is directly proportional to the amount of the target substance in the sample.
Types of ELISA
There are several variations of the ELISA method, each with its own specific application:
- Direct ELISA: The antigen is directly coated onto the plate, and a labeled antibody binds to it.
- Indirect ELISA: The antigen is coated onto the plate, an unlabeled primary antibody binds to the antigen, and a labeled secondary antibody (that binds to the primary antibody) is added.
- Sandwich ELISA: A capture antibody is coated onto the plate, the antigen binds to the capture antibody, and a detection antibody (either direct or indirect) binds to the antigen.
- Competitive ELISA: An antigen is pre-incubated with the sample. The mixture is then added to a plate coated with the same antigen. If the sample contains the target antigen, it will compete with the coated antigen for binding to the antibody, resulting in a weaker signal.
Applications of ELISA
ELISA is a versatile technique with a wide range of applications in:
- Diagnostics: Detecting infectious diseases (e.g., HIV, Lyme disease, Zika virus), diagnosing autoimmune diseases, and identifying allergies.
- Research: Quantifying proteins, antibodies, and other molecules in biological samples.
- Quality Control: Ensuring the quality of food, pharmaceuticals, and other products.
- Pregnancy testing: Detecting the presence of hCG hormone.
Advantages of ELISA
- High Sensitivity: Able to detect even small amounts of the target substance.
- High Specificity: Able to distinguish between different substances.
- Relatively Inexpensive: Compared to some other analytical techniques.
- Easy to Perform: Can be automated for high-throughput screening.
In conclusion, the ELISA method is a powerful and widely used technique in various fields due to its sensitivity, specificity, and versatility in detecting and quantifying specific substances in complex samples.
