线粒体谷胱甘肽 （GSH） 探针
Mito-RealThiol (MitoRT) Mitochondrial Glutathione (GSH) Probe
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产品编号： EBY003
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相关CAS号：N/A
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分子式：N/A
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分子量：692.59
| 订货号 | 规格 | 价格 | 库存 | 数量 | 购买 | |
|---|---|---|---|---|---|---|
| EBY003 | 50nmol | 4500.00元 | 2 |
|
Mito RealThiol (MitoRT) provides ratiometric fluorescent readout for quantification of mitochondrial GSH in live mammalian cells.
Highlights:
- MitoRT is cell permeable and preferentially accumulated in mitochondria
- Conveniently compare mitochondrial GSH levels in live cells using confocal microscopy, and FACS
- Signals from blue and green channels allow for deduction of mitochondrial GSH concentration, given an appropriate calibration curve
Subcellular detection and localization of GSH is important in understanding the modulation of redox status, the effect of drugs, and the mechanisms of detoxification. Furthermore, differences in GSH levels in response to oxidative stress in subpopulations of cells have been reported, underscoring the importance of detection methods amenable to flow cytometry and automated fluorescence detection.
| Product Type: | Small Molecule |
| Name: | Mito-RealThiol (MitoRT) |
| Chemical Formula: | C38H35BrN3O3P |
| Molecular Weight: | 692.59 |
| Format: | Red to orange solid (Lyophilized powder in tubes) |
| Purity: | >97%, HPLC |
| Tested Applications: | 1 µM for cell staining |
| Solubility: | DI Water (DO NOT dissolve in DMSO) |
| Spectral Information: | Ex/Em: MitoRT: 488/565; MitoRT-GSH: 405/488 |
| Platform: | Confocal microscopy/FACS |
| Compatible Cells: | Mammalian cells e.g. HeLa, HT1080 |
| Detection Method: | Fluorescence |
| Comments: | - Dilute stock with imaging buffer. - A laser-based light source is preferred for best results. |
| Storage: | -80C |
| Shipped: | Ambient Temperature |
产品使用Protocol：Mito-RealThiol (MitoRT) Mitochondrial Glutathione (GSH) Detection Probe User Manual
产品说明书：Mito-RealThiol (MitoRT) Mitochondrial Glutathione (GSH) Detection Probe Datasheet
参考文献
Jianwei Chen, Xiqian Jiang, Chengwei Zhang, Kevin R. MacKenzie, Fabio Stossi, Timothy Palzkill, Meng C. Wang, and Jin Wang*, "Reversible Reaction-Based Fluorescent Probe for Real-Time Imaging of Glutathione Dynamics in Mitochondria", ACS Sensors, 2017, ASAP.
Jiang X, Chen J, Wang MC, Wang J. Glutathione Quantification in Live Cells with Real-Time Imaging and Flow Cytometry. STAR Protoc. 2020 Nov 14;1(3):100170. View Article
实验问题：
1.细胞膜信号过多：（a）.检查MitoRT浓度，必要时降低染色浓度;（b）.加入台盼蓝作为表面荧光淬灭剂，1-5%台盼蓝即可测试。
2.看不到信号：（a）.检查显微镜设置，特别是绿色通道波长，（b）如果使用共聚焦显微镜，请仔细检查焦点，因为染色仅在非常有限的Z范围内可见;检查染色时间和温度，较低的温度（室温）和较短的染色时间（<15分钟）将减慢探针在线粒体中的积累，导致线粒体特异性信号减少，并且较长的染色时间（>60分钟）通常由于探针的代谢而导致信号降低，特别是在癌细胞中。
3.对照细胞中的比例不稳定：（a）.确保在整个实验过程中所有设置保持不变，尤其是两个通道中的PMT设置;（b）.尝试限制染色时间（15-30分钟），并确保每个样品的染色时间相似（差异5分钟内），以尽量减少细胞代谢，从而从细胞中清除MitoRT;（c）.检查成像缓冲液，确保缓冲液中没有兴奋剂（如高剂量生长激素或葡萄糖等），并且没有可能改变细胞氧化还原状态或导致探针分解的DMSO;使用ABC转运蛋白抑制剂，如丙磺舒（1mM）。
4.来自两个通道的图像有明显的偏移：检查来自两个通道的信号是同时记录还是顺序记录，确保两个信号同时记录，否则线粒体的运动会影响结果。
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