Detection of protein-protein interactions using protein-fragment complementation assays (PCA)

Emma Barnard, Neil V. McFerran, John Nelson, David J. Timson

    Research output: Contribution to journalArticlepeer-review


    Protein-protein interactions play a central role in many cellular processes. Their characterisation is necessary in order to analyse these processes and for the functional identification of unknown proteins. Existing detection methods such as the yeast two-hybrid (Y2H) and tandem affinity purification (TAP) method provide a means to answer rapidly questions regarding protein-protein interactions, but have limitations which restrict their use to certain interaction networks; furthermore they provide little information regarding interaction localisation at the subcellular level. The development of protein-fragment complementation assays (PCA) employing a fluorescent reporter such as a member of the green fluorescent protein (GFP) family has led to a new method of interaction detection termed Bimolecular Fluorescent Complementation (BiFC). These assays have become important tools for understanding protein interactions and the development of whole genome interaction maps. BiFC assays have the advantages of very low background signal coupled with rapid detection of protein-protein interactions in vivo while also providing information regarding interaction compartmentalisation. Modified forms of the assay such as the use of combinations of spectral variants of GFP have allowed simultaneous visualisation of multiple competing interactions in vivo. Advantages and disadvantages of the method are discussed in the context of other fluorescence-based interaction monitoring techniques.

    Original languageEnglish
    Pages (from-to)17-27
    Number of pages11
    JournalCurrent Proteomics
    Issue number1
    Publication statusPublished - 1 Apr 2007


    • Bimolecular fluorescent complementation
    • EGFP
    • Green fluorescent protein
    • Protein complementation assay
    • Protein-protein interactions
    • Two hybrid screen


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