Fluorescence imaging detection of nanodomain redox signaling events at organellar contacts
Summary: This protocol describes how to visualize, detect, and analyze redox signals (oxidative bursts) at the ER-mitochondrial interface. It uses drug-inducible crosslinking to target the genetically encoded glutathione redox sensor Grx1roGFP2 to organellar contact sites to measure local redox chan...
| Main Authors: | , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Elsevier
2022-03-01
|
| Series: | STAR Protocols |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S266616672100825X |
| _version_ | 1818774446785167360 |
|---|---|
| author | David M. Booth Péter Várnai Suresh K. Joseph György Hajnóczky |
| author_facet | David M. Booth Péter Várnai Suresh K. Joseph György Hajnóczky |
| author_sort | David M. Booth |
| collection | DOAJ |
| description | Summary: This protocol describes how to visualize, detect, and analyze redox signals (oxidative bursts) at the ER-mitochondrial interface. It uses drug-inducible crosslinking to target the genetically encoded glutathione redox sensor Grx1roGFP2 to organellar contact sites to measure local redox changes associated with transient depolarizations of the mitochondrial membrane potential (flickers). The strategy allows imaging of the oxidized to reduced glutathione ratio (GSSG:GSH) in subcellular regions below the diffraction limit with good temporal resolution and minimum phototoxicity. Moreover, the strategy also applies to diverse parameters including pH, H2O2, and Ca2+.For complete details on the use and execution of this profile, please refer to Booth et al. (2016) and Booth et al. (2021). |
| first_indexed | 2024-12-18T10:41:17Z |
| format | Article |
| id | doaj.art-1134d342c13544f2a7b3b44d93ce631f |
| institution | Directory Open Access Journal |
| issn | 2666-1667 |
| language | English |
| last_indexed | 2024-12-18T10:41:17Z |
| publishDate | 2022-03-01 |
| publisher | Elsevier |
| record_format | Article |
| series | STAR Protocols |
| spelling | doaj.art-1134d342c13544f2a7b3b44d93ce631f2022-12-21T21:10:38ZengElsevierSTAR Protocols2666-16672022-03-0131101119Fluorescence imaging detection of nanodomain redox signaling events at organellar contactsDavid M. Booth0Péter Várnai1Suresh K. Joseph2György Hajnóczky3MitoCare Center, Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Jefferson Alumni Hall 1020 Locust Street, Philadelphia, PA 19107, USA; Corresponding authorDepartment of Physiology, Semmelweis University, Faculty of Medicine, 1444 Budapest, HungaryMitoCare Center, Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Jefferson Alumni Hall 1020 Locust Street, Philadelphia, PA 19107, USAMitoCare Center, Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Jefferson Alumni Hall 1020 Locust Street, Philadelphia, PA 19107, USA; Corresponding authorSummary: This protocol describes how to visualize, detect, and analyze redox signals (oxidative bursts) at the ER-mitochondrial interface. It uses drug-inducible crosslinking to target the genetically encoded glutathione redox sensor Grx1roGFP2 to organellar contact sites to measure local redox changes associated with transient depolarizations of the mitochondrial membrane potential (flickers). The strategy allows imaging of the oxidized to reduced glutathione ratio (GSSG:GSH) in subcellular regions below the diffraction limit with good temporal resolution and minimum phototoxicity. Moreover, the strategy also applies to diverse parameters including pH, H2O2, and Ca2+.For complete details on the use and execution of this profile, please refer to Booth et al. (2016) and Booth et al. (2021).http://www.sciencedirect.com/science/article/pii/S266616672100825XCell BiologyMicroscopyMolecular/Chemical Probes |
| spellingShingle | David M. Booth Péter Várnai Suresh K. Joseph György Hajnóczky Fluorescence imaging detection of nanodomain redox signaling events at organellar contacts STAR Protocols Cell Biology Microscopy Molecular/Chemical Probes |
| title | Fluorescence imaging detection of nanodomain redox signaling events at organellar contacts |
| title_full | Fluorescence imaging detection of nanodomain redox signaling events at organellar contacts |
| title_fullStr | Fluorescence imaging detection of nanodomain redox signaling events at organellar contacts |
| title_full_unstemmed | Fluorescence imaging detection of nanodomain redox signaling events at organellar contacts |
| title_short | Fluorescence imaging detection of nanodomain redox signaling events at organellar contacts |
| title_sort | fluorescence imaging detection of nanodomain redox signaling events at organellar contacts |
| topic | Cell Biology Microscopy Molecular/Chemical Probes |
| url | http://www.sciencedirect.com/science/article/pii/S266616672100825X |
| work_keys_str_mv | AT davidmbooth fluorescenceimagingdetectionofnanodomainredoxsignalingeventsatorganellarcontacts AT petervarnai fluorescenceimagingdetectionofnanodomainredoxsignalingeventsatorganellarcontacts AT sureshkjoseph fluorescenceimagingdetectionofnanodomainredoxsignalingeventsatorganellarcontacts AT gyorgyhajnoczky fluorescenceimagingdetectionofnanodomainredoxsignalingeventsatorganellarcontacts |