Medizinische Universität Graz - Research portal
Selected Publication:
SHR Neuro Cancer Cardio Lipid Metab Microb
Eroglu, E; Rost, R; Bischof, H; Blass, S; Schreilechner, A; Gottschalk, B; Depaoli, MR; Klec, C; Charoensin, S; Madreiter-Sokolowski, CT; Ramadani, J; Waldeck-Weiermair, M; Graier, WF; Malli, R.
Application of Genetically Encoded Fluorescent Nitric Oxide (NO•) Probes, the geNOps, for Real-time Imaging of NO• Signals in Single Cells.
J Vis Exp. 2017; 5(121):
Doi: 10.3791/55486
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- Leading authors Med Uni Graz
- EROGLU Emrah
- Malli Roland
- Co-authors Med Uni Graz
- Bischof Helmut
- Blass Sandra
- Depaoli Maria Rosa
- Gottschalk Benjamin
- Graier Wolfgang
- Klec Christiane
- Madreiter-Sokolowski Corina
- Ramadani-Muja Jeta
- Rost René
- Waldeck-Weiermair Markus
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- Abstract:
- Nitric Oxide (NO•) is a small radical, which mediates multiple important cellular functions in mammals, bacteria and plants. Despite the existence of a large number of methods for detecting NO• in vivo and in vitro, the real-time monitoring of NO• at the single-cell level is very challenging. The physiological or pathological effects of NO• are determined by the actual concentration and dwell time of this radical. Accordingly, methods that allow the single-cell detection of NO• are highly desirable. Recently, we expanded the pallet of NO• indicators by introducing single fluorescent protein-based genetically encoded nitric oxide (NO•) probes (geNOps) that directly respond to cellular NO• fluctuations and, hence, addresses this need. Here we demonstrate the usage of geNOps to assess intracellular NO• signals in response to two different chemical NO•-liberating molecules. Our results also confirm that freshly prepared 3-(2-hydroxy-1-methyl-2-nitrosohydrazino)-N-methyl-1-propanamine (NOC-7) has a much higher potential to evoke change in intracellular NO• levels as compared with the inorganic NO• donor sodium nitroprusside (SNP). Furthermore, dual-color live-cell imaging using the green geNOps (G-geNOp) and the chemical Ca2+ indicator fura-2 was performed to visualize the tight regulation of Ca2+-dependent NO• formation in single endothelial cells. These representative experiments demonstrate that geNOps are suitable tools to investigate the real-time generation and degradation of single-cell NO• signals in diverse experimental setups.
- Find related publications in this database (using NLM MeSH Indexing)
- Animals -
- Cells, Cultured -
- Endothelial Cells - metabolism
- Fluorescent Dyes - metabolism
- Fura-2 - chemistry
- Genetic Vectors -
- HEK293 Cells -
- Humans -
- Hydrazines - pharmacology
- Microscopy, Fluorescence - methods
- Nitric Oxide - metabolism
- Nitric Oxide Donors - pharmacology
- Nitroprusside - pharmacology
- Parvovirinae - genetics
- Find related publications in this database (Keywords)
- Molecular Biology
- Issue 121
- Ca2+
- eNOS
- Fluorescence Microscopy
- Fura-2
- Genetically Encoded Probes
- Live-Cell Imaging
- Multichannel Imaging
- Nitric Oxide
- NO center dot-Donors
- NOC-7
- Single Cell Analysis
- SNP