Immunoglobulin d

Immunoblot assay is a widely used biochemical technique in molecular biology, immunogenetics and other molecular biology disciplines to detect proteins in a sample. The technique uses gel electrophoresis to separate native or denatured proteins by the length of the polypeptide (denaturing conditions) or by the 3-D structure of the protein (native/ non-denaturing conditions). The proteins are then transferred to a membrane (typically nitrocellulose or PVDF), where they are stained with antibodies specific to the target protein.

History

The immunoblot assay was first developed in 1979 by Harry Towbin and colleagues as a method to detect and analyze proteins. It was originally called "immunoblotting" because of the use of antibodies to detect the proteins. The technique has since been adapted for a wide range of applications, including diagnosis of diseases and identification of antibodies in research and clinical laboratories.

Procedure

The procedure for an immunoblot assay involves several steps:

1. Sample Preparation: The sample containing the protein of interest is prepared for the assay. This may involve purification or concentration of the protein.
2. Gel Electrophoresis: The proteins in the sample are separated by size using gel electrophoresis.
3. Transfer: The separated proteins are transferred from the gel to a membrane, typically made of nitrocellulose or PVDF.
4. Blocking: The membrane is treated to prevent any nonspecific binding of antibodies to the membrane.
5. Antibody Incubation: The membrane is incubated with a specific antibody that binds to the protein of interest.
6. Detection: The presence of the antibody on the membrane is detected, usually by using a secondary antibody that binds to the first antibody and has an enzyme attached that can produce a color change.

Applications

Immunoblot assays are used in a variety of applications, including:

1. Disease Diagnosis: Immunoblot assays can be used to detect specific proteins in a patient's blood sample, which can help diagnose diseases such as HIV, Lyme disease, and certain types of cancer.
2. Research: In research laboratories, immunoblot assays are used to study protein function and expression. They can also be used to identify and characterize antibodies.
3. Quality Control: In the pharmaceutical and biotechnology industries, immunoblot assays are used for quality control to ensure that products contain the correct proteins and that these proteins are correctly folded.

See Also

• Western blot
• Southern blot
• Northern blot
• Dot blot