A co-culture-based multiparametric imaging technique to dissect local h2o2 signals with targeted hyper7

Seçilmiş, Melike and Altun, Hamza Yusuf and Pilic, Johannes and Erdoğan, Yusuf Ceyhun and Çokluk, Zeynep and Ata, Büşra Nur and Sevimli, Gülşah and Ghaffari Zaki, Asal and Yigit, Esra Nur and Öztürk, Gürkan and Malli, Roland and Eroğlu, Emrah (2021) A co-culture-based multiparametric imaging technique to dissect local h2o2 signals with targeted hyper7. Biosensors, 11 (9). ISSN 2079-6374

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Multispectral live-cell imaging is an informative approach that permits detecting biological processes simultaneously in the spatial and temporal domain by exploiting spectrally distinct biosen-sors. However, the combination of fluorescent biosensors with distinct spectral properties such as different sensitivities, and dynamic ranges can undermine accurate co-imaging of the same analyte in different subcellular locales. We advanced a single-color multiparametric imaging method, which allows simultaneous detection of hydrogen peroxide (H2 O2 ) in multiple cell locales (nucleus, cytosol, mitochondria) using the H2 O2 biosensor HyPer7. Co-culturing of endothelial cells stably expressing differentially targeted HyPer7 biosensors paved the way for co-imaging compartmentalized H2 O2 signals simultaneously in neighboring cells in a single experimental setup. We termed this approach COMPARE IT, which is an acronym for co-culture-based multiparametric imaging technique. Em-ploying this approach, we detected lower H2 O2 levels in mitochondria of endothelial cells compared to the cell nucleus and cytosol under basal conditions. Upon administering exogenous H2 O2, the cytosolic and nuclear-targeted probes displayed similarly slow and moderate HyPer7 responses, whereas the mitochondria-targeted HyPer7 signal plateaued faster and reached higher amplitudes. Our results indicate striking differences in mitochondrial H2 O2 accumulation of endothelial cells. Here, we present the method’s potential as a practicable and informative multiparametric live-cell imaging technique.
Item Type: Article
Uncontrolled Keywords: Co-imaging; Endothelial cell line; Hydrogen peroxide; HyPer7; Mitochondria; Multiparametric imaging; Multiplex live-cell imaging; Stable cell lines; Targeted biosensors
Divisions: Faculty of Engineering and Natural Sciences > Academic programs > Biological Sciences & Bio Eng.
Faculty of Engineering and Natural Sciences
Sabancı University Nanotechnology Research and Application Center
Depositing User: Emrah Eroğlu
Date Deposited: 28 Aug 2022 13:07
Last Modified: 28 Aug 2022 13:07
URI: https://research.sabanciuniv.edu/id/eprint/43810

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