Fluorescent enzyme immunoassay

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Fluorescent enzyme immunoassay
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Purpose Detection and quantification of antigens or antibodies
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Fluorescent Enzyme Immunoassay (FEIA) is a type of immunoassay that utilizes fluorescence to detect and quantify the presence of specific antigens or antibodies in a sample. This technique combines the specificity of antibody-antigen interactions with the sensitivity of fluorescent detection, making it a powerful tool in clinical diagnostics and research.

Principle[edit | edit source]

The principle of FEIA is based on the use of an enzyme-linked antibody that binds to the target antigen. The enzyme acts on a non-fluorescent substrate to produce a fluorescent product. The intensity of the fluorescence is directly proportional to the amount of antigen or antibody present in the sample.

Components[edit | edit source]

  • Antigen or Antibody: The target molecule that is being detected.
  • Enzyme-Linked Antibody: An antibody conjugated to an enzyme that catalyzes a reaction to produce a fluorescent signal.
  • Substrate: A non-fluorescent molecule that is converted into a fluorescent product by the enzyme.
  • Fluorometer: An instrument used to measure the intensity of fluorescence.

Procedure[edit | edit source]

The procedure for conducting a fluorescent enzyme immunoassay typically involves the following steps:

Sample Preparation[edit | edit source]

Samples, such as blood, serum, or other biological fluids, are collected and prepared for analysis. The sample may require dilution or other pre-treatment steps to ensure compatibility with the assay.

Coating the Plate[edit | edit source]

A microtiter plate is coated with either the antigen or antibody, depending on whether the assay is designed to detect antibodies or antigens, respectively. The plate is then blocked to prevent non-specific binding.

Addition of Sample[edit | edit source]

The prepared sample is added to the wells of the microtiter plate. If the target antigen or antibody is present, it will bind to the coated surface.

Addition of Enzyme-Linked Antibody[edit | edit source]

An enzyme-linked antibody specific to the target is added to the wells. This antibody binds to the target molecule, forming a complex.

Substrate Addition[edit | edit source]

A substrate that is specific to the enzyme is added. The enzyme catalyzes the conversion of the substrate into a fluorescent product.

Detection[edit | edit source]

The plate is placed in a fluorometer, which measures the intensity of the fluorescence emitted by the product. The fluorescence intensity is proportional to the concentration of the target antigen or antibody in the sample.

Applications[edit | edit source]

FEIA is widely used in various fields, including:

Advantages[edit | edit source]

  • High Sensitivity: Capable of detecting low concentrations of antigens or antibodies.
  • Specificity: High specificity due to the use of antibodies.
  • Quantitative: Provides quantitative results that can be used to determine the concentration of the target molecule.

Limitations[edit | edit source]

  • Complexity: Requires specialized equipment and trained personnel.
  • Cost: Can be more expensive than other types of immunoassays.
  • Interference: Potential for interference from other fluorescent substances in the sample.

See Also[edit | edit source]

External Links[edit | edit source]



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