Question

Which of the following method requires availability of antibody

• A. Yeast Two Hybrid Assay
• B. Pull Down Assay
• C. Split Ubiquitin Assay
• D. BiFC

Hint:

To isolate antibody-independent complexes, researchers often tag target proteins with epitopes (such as GST or His-tag) that bind to glutathione or nickel-charged columns.

Answer 1

The Correct Option is B

Step 1: Understanding the Concept:
Protein-protein interactions can be analyzed using *in vivo* genetic reporter systems or *in vitro* physical affinity-purification techniques.

Step 2: Detailed Explanation:
- Pull-Down Assay Co-Immunoprecipitation (Co-IP): An *in vitro* physical technique.
It utilizes specific antibodies immobilized on beads to capture and precipitate a target protein along with its interacting partners from a complex cell lysate.
- Yeast Two-Hybrid (Y2H) Assay: An *in vivo* genetic assay in yeast that relies on reconstituting a functional transcription factor to drive reporter gene expression; it does not require antibodies.
- Split Ubiquitin Assay: Used to detect interactions between membrane-bound proteins; it relies on reporter release by ubiquitin-specific proteases.
- BiFC (Bimolecular Fluorescence Complementation): Relies on the association of two non-fluorescent fragments of a fluorescent protein, which emit light directly upon interaction.

Step 3: Final Answer:
The method that requires antibodies is the pull-down/Co-IP assay.