Bimolecular Fluorescence Complementation (BiFC) is a powerful molecular imaging method used to visualize protein-protein interactions (PPIs) in living cells or organisms. BiFC is based on the reassociation of hemi-fragments of a monomeric fluorescent protein upon spatial proximity. It is compatible with conventional light microscopy, providing a resolution that is constrained by the diffraction of light to around 250 nm. PAmCherry1, a fluorescent protein compatible for BiFC and Photoactivated Localization Microscopy (PALM), allows the analysis of PPIs with nanometer spatial resolution and single molecule sensitivity. To date, this so-called BiFC-PALM approach has only been described in human cell culture. Here, we present a protocol for performing BiFC-PALM in Drosophila larval salivary glands, using the interaction between the Hox protein Ultrabithorax (Ubx) and the generic Hox cofactor Extradenticle (Exd) as a model system.
Keywords: BiFC; Drosophila; Exd; Hox; PALM; PAmCherry1; Super resolution imaging (SR); Transcription factors (TFs).
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