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#BiFC

2 public questions tagged with this topic.

Protein-Protein Interaction

BiFC (Bimolecular Fluorescence Complementation) detects:

Bimolecular Fluorescence Complementation is a genetic technique to visualize protein-protein association in living cells. Candidate proteins are fused to non-fluorescent N- and C-terminal fragments of a fluorescent protein such as YFP or Venus. Individually fragments are non-fluorescent. When fused proteins interact, fragments are brought into proximity to refold into functional fluorophore, emitting fluorescence detectable by microscopy or flow cytometry. Irreversible complementation reports localization of interaction. It does not measure RNA cleavage, phosphorylation per se, or mismatch repair but specifically protein-protein interaction.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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Protein-Protein Interaction

Which interaction is label-free and measured in real-time?

Ideal interaction analysis provides association rate, dissociation rate, and equilibrium dissociation without labeling proteins, avoiding artefacts from fluorophores or radioisotopes and enabling kinetic quantification. Surface Plasmon Resonance achieves this by optical detection of refractive index changes upon analyte-ligand binding on gold sensor chip. It operates continuously, generating sensorgrams in real-time with high sensitivity. EMSA requires labeled nucleic acid, FRET needs fluorescent fusions, BiFC relies on split fluorescent protein complementation. SPR therefore provides unbiased, label-free quantification of protein-protein binding specificity and affinity in real-time.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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