Related Experiment Video
Updated: Jun 25, 2025
Conventional BODIPY Conjugates for Live-Cell Super-Resolution Microscopy and Single-Molecule Tracking
Published on: June 8, 2020
Fluorescent fatty acid conjugates for live cell imaging of peroxisomes
Daria Korotkova1, Anya Borisyuk1, Anthony Guihur2
1Global Health Institute, Faculty of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland.
Insights
New fluorescent probes, PeroxiSPY650 and PeroxiSPY555, enable live imaging of peroxisomes and diagnosis of peroxisome biogenesis disorders (PBD). These probes offer bright, specific staining for studying peroxisome function and dysfunction.
Area of Science:
- Cell Biology
- Biochemistry
- Molecular Imaging
Background:
- Peroxisomes are vital eukaryotic organelles involved in crucial metabolic processes like fatty acid oxidation and lipid biosynthesis.
- Dysfunction of peroxisomes causes severe pediatric neurodegenerative diseases, known as Peroxisome Biogenesis Disorders (PBD).
- Live cell imaging is critical for understanding peroxisome dynamics and diagnosing PBD, but current tools are limited.
Purpose of the Study:
- To develop novel fluorescent probes for specific and efficient live cell imaging of peroxisomes.
- To create tools for diagnosing peroxisome dysfunction and PBD-linked abnormalities.
- To overcome limitations in the existing peroxisomal imaging toolkit.
Main Methods:
- Utilized the BODIPY-C12 fluorescent fatty acid probe for initial peroxisome staining.
- Engineered enhanced peroxisome-specific probes, PeroxiSPY650 and PeroxiSPY555, from BODIPY-C12.
- Validated probe performance in live mammalian cells, including PBD cell models and patient-derived lines.
Main Results:
- BODIPY-C12 successfully stained both functional and dysfunctional peroxisomes in live cells.
- PeroxiSPY650 and PeroxiSPY555 demonstrated high peroxisome specificity, bright red/far-red fluorescence, and rapid, non-cytotoxic staining.
- The PeroxiSPY probes effectively enabled the diagnosis of peroxisome abnormalities in PBD models.
Conclusions:
- PeroxiSPY probes represent a significant advancement for live cell, whole organism, and tissue imaging of peroxisomes.
- These novel reagents are ideal for fundamental research on peroxisome biology and clinical diagnostics of PBD.
- The developed probes address critical unmet needs in the field of peroxisomal research and diagnostics.
Abstract:
Peroxisomes are eukaryotic organelles that are essential for multiple metabolic pathways, including fatty acid oxidation, degradation of amino acids, and biosynthesis of ether lipids. Consequently, peroxisome dysfunction leads to pediatric-onset neurodegenerative conditions, including Peroxisome Biogenesis Disorders (PBD). Due to the dynamic, tissue-specific, and context-dependent nature of their biogenesis and function, live cell imaging of peroxisomes is essential for studying peroxisome regulation, as well as for the diagnosis of PBD-linked abnormalities. However, the peroxisomal imaging toolkit is lacking in many respects, with no reporters for substrate import, nor cell-permeable probes that could stain dysfunctional peroxisomes. Here we report that the BODIPY-C12 fluorescent fatty acid probe stains functional and dysfunctional peroxisomes in live mammalian cells. We then go on to improve BODIPY-C12, generating peroxisome-specific reagents, PeroxiSPY650 and PeroxiSPY555. These probes combine high peroxisome specificity, bright fluorescence in the red and far-red spectrum, and fast non-cytotoxic staining, making them ideal tools for live cell, whole organism, or tissue imaging of peroxisomes. Finally, we demonstrate that PeroxiSPY enables diagnosis of peroxisome abnormalities in the PBD CRISPR/Cas9 cell models and patient-derived cell lines.
