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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 proteinProtein 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.

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Using Bimolecular Fluorescence Complementation (BiFC) with Photoactivated Localization Microscopy (PALM) to Analyze Hox/Cofactor Interactions at the Super Resolution Scale in Drosophila Salivary Glands

  • 实验方案

  • 发表日期：

    2025年

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  • Solène Vanderperre,
  • Marilyne Duffraisse,
  • Christophe Place,
  • Samir Merabet

摘要

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 proteinProtein 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.