Labelled Antibodies in Western Blotting: Detecting Specific Proteins
Labelled Antibodies are used to detect
a) Detect the presence of a particular DNA molecule in Southern blotting
b) Detect the presence of a particular RNA molecule in Southern blotting
c) Detect the presence of a particular protein molecule in Southern blotting
d) Detect the presence of a particular protein molecule in Western blotting
Correct Answer: (d) Detect the presence of a particular protein molecule in Western blotting
Labelled Antibodies in Western Blotting : Labelled antibodies are essential tools in Western blotting, a widely used molecular biology technique for detecting specific proteins in a complex sample. Western blotting is highly specific and sensitive, making it a crucial method for protein identification, disease diagnostics, and biomedical research.
Why Other Options Are Incorrect
- (a) Detect the presence of a particular DNA molecule in Southern blotting – Southern blotting detects DNA sequences using a DNA or RNA probe, not antibodies.
- (b) Detect the presence of a particular RNA molecule in Southern blotting – RNA is detected using Northern blotting with a complementary RNA or DNA probe, not antibodies.
- (c) Detect the presence of a particular protein molecule in Southern blotting – Southern blotting is not used for protein detection.
Role of Labelled Antibodies in Western Blotting
Western blotting involves the use of labelled antibodies to detect and visualize specific proteins. The process includes the following steps:
- Protein Separation – Proteins are separated based on size using SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis).
- Transfer to Membrane – Separated proteins are transferred onto a nitrocellulose or PVDF membrane.
- Blocking – The membrane is blocked with a non-specific protein like bovine serum albumin (BSA) to prevent non-specific binding.
- Primary Antibody Binding – A primary antibody specific to the target protein is applied to the membrane.
- Secondary Antibody Binding – A secondary antibody labeled with an enzyme (e.g., horseradish peroxidase) or a fluorescent tag binds to the primary antibody.
- Detection – The signal from the labelled secondary antibody is detected using:
- Chemiluminescence – Produces light when the enzyme reacts with a substrate.
- Fluorescence – Produces a fluorescent signal.
- Colorimetric Detection – Produces a colored product.
Types of Labelled Antibodies
- Enzyme-linked antibodies – Common enzymes used include horseradish peroxidase (HRP) and alkaline phosphatase (AP).
- Fluorescent-labelled antibodies – Used in advanced imaging techniques for higher sensitivity.
Applications of Western Blotting
- Medical Diagnostics – Used to confirm diseases like HIV and Lyme disease.
- Protein Expression Studies – Helps analyze protein levels and modifications.
- Cancer Research – Used to identify and quantify biomarkers.
Conclusion
Labelled antibodies are vital in Western blotting for the detection of specific proteins with high accuracy and sensitivity. Their ability to target and bind specific antigens allows researchers to study protein expression, modification, and interaction, making Western blotting a cornerstone of molecular biology and clinical diagnostics.
81 Comments
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