Protonex™ Red 670 AM *Cell-Permeable*
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Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
sales@aatbio.com | |
International | See distributors |
Bulk request | Inquire |
Custom size | Inquire |
Shipping | Standard overnight for United States, inquire for international |
Molecular weight | 521.05 |
Solvent | DMSO |
Excitation (nm) | 643 |
Emission (nm) | 660 |
H-phrase | H303, H313, H333 |
Hazard symbol | XN |
Intended use | Research Use Only (RUO) |
R-phrase | R20, R21, R22 |
Storage | Freeze (< -15 °C); Minimize light exposure |
UNSPSC | 12352200 |
Overview | ![]() ![]() |
Molecular weight 521.05 | Excitation (nm) 643 | Emission (nm) 660 |
Platform
Fluorescence microscope
Excitation | Cy5 Filter Set |
Emission | Cy5 Filter Set |
Recommended plate | Black wall/clear bottom |
Fluorescence microplate reader
Excitation | 640 nm |
Emission | 680 nm |
Cutoff | 665 nm |
Recommended plate | Black wall/clear bottom |
Example protocol
PREPARATION OF STOCK SOLUTIONS
Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles
Prepare a 10 to 20 mM stock solution of Protonex™ Red 670 AM in high-quality anhydrous DMSO.
Note: An unused Protonex™ Red 670 AM stock solution should be divided into single-use aliquots and stored at ≤ -20 º C. Protect from light and avoid repeated freeze-thaw cycles.
PREPARATION OF WORKING SOLUTION
On the day of the experiment, either dissolve Protonex™ Red 670 AM dye in DMSO or thaw an aliquot of the indicator stock solution to room temperature.
Prepare a Protonex™ Red 670 AM dye working solution of 5 to 20 μM in a buffer of your choice (e.g., Hanks and Hepes buffer).
Note: The nonionic detergent Pluronic® F-127 can be used to increase the aqueous solubility of AM esters. The final Pluronic® F-127 concentration should be approximately 0.02% in the staining buffer. A variety of Pluronic® F-127 products can be purchased from AAT Bioquest (here). Avoid long-term storage of AM esters in the presence of Pluronic® F-127.
Note: If your cells contain organic anion-transporters, probenecid (1-4 mM) may be added to the dye working solution (final in well concentration will be 0.5-2 mM) to reduce leakage of the de-esterified indicators. A variety of ReadiUse™ probenecid products, including water-soluble, sodium salt, and stabilized solutions, can be purchased from AAT Bioquest (here).
SAMPLE EXPERIMENTAL PROTOCOL
The following is a recommended protocol for loading Protonex™ Red 670 AM dye into live mammalian cells. This protocol only provides a guideline and should be modified according to your specific needs.
Prepare viable cells as desired (e.g., 100 µL/well/96-well plate or 25 µL/well/384-well plate).
On the next day, add the Protonex™ Red 670 AM dye working solution into the cell plate in equal volumes, such as 100 µL/well/96-well plate or 25 µL/well/384-well plate.
Incubate the dye-loaded plate in a cell incubator at 37 °C for 30 to 60 minutes.
Replace the dye working solution with HHBS or buffer of your choice to remove any excess dye.
Prepare the compound plates using HHBS or a buffer of your choice.
Perform the pH assay as desired while simultaneously monitoring fluorescence. This can be done with either a fluorescence microscope that has a Cy5 filter set or a fluorescence plate reader at Ex/Em = 640/680 nm (cutoff 665 nm).
Calculators
Common stock solution preparation
0.1 mg | 0.5 mg | 1 mg | 5 mg | 10 mg | |
1 mM | 191.92 µL | 959.601 µL | 1.919 mL | 9.596 mL | 19.192 mL |
5 mM | 38.384 µL | 191.92 µL | 383.84 µL | 1.919 mL | 3.838 mL |
10 mM | 19.192 µL | 95.96 µL | 191.92 µL | 959.601 µL | 1.919 mL |
Molarity calculator
Mass (Calculate) | Molecular weight | Volume (Calculate) | Concentration (Calculate) | Moles | ||||
/ | = | x | = |
Product Family
Name | Excitation (nm) | Emission (nm) |
Protonex™ Red 670 acid | 643 | 660 |
Protonex™ Red 780 AM *Cell-Permeable* | 748 | 769 |
Protonex™ Lyso-Red 670 | 643 | 660 |
Images
![Response of HeLa cells labeled with Protonex™ Red 670 AM. HeLa cells were incubated with 5 µM of Protonex™ Red 670 AM for 30 minutes at 37°C. Incubation of Protonex™ Red 670 AM solution with HeLa cells showed a homogenous uptake of Protonex™ Red 670 AM and stained cell cytosol. The Spexyte™ Intracellular pH Calibration Buffer Kit (Cat No. 21235) was used to clamp the intracellular pH with extracellular buffers at pH 4 to 10. (A) Images were acquired using a fluorescence microscope with a Cy5 filter set, and (B) fluorescence was measured using a ClarioStar fluorescence microplate reader (Ex/Em = 640/680 nm, cutoff = 665 nm).](/_next/image?url=https%3A%2F%2Fimages.aatbio.com%2Fproducts%2Ffigures-and-data%2Fprotonex-red-670-am-cell-permeable%2Ffigure-for-protonex-red-670-am-cell-permeable_joBQA.png&w=3840&q=75)
References
Authors: Wang, Meng and Tong, Kun and Chen, Zhe and Wen, Zhengde
Journal: The Journal of surgical research (2023): 245-255
Authors: Sarker, Rafiquel and Tse, Chung Ming and Lin, Ruxian and McNamara, George and Singh, Varsha and Donowitz, Mark
Journal: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology (2022): 39-49
Authors: Miao, Yumeng and Zheng, Yun and Geng, Yanzhi and Yang, Ling and Cao, Na and Dai, Yue and Wei, Zhifeng
Journal: Theranostics (2021): 4531-4548
Authors: He, Qifu and Wu, Shenghui and Huang, Ming and Wang, Ying and Zhang, Kang and Kang, Jian and Zhang, Yong and Quan, Fusheng
Journal: Frontiers in cell and developmental biology (2021): 747722
Authors: Ishikawa, Mayumi and Toyomura, Junko and Yagi, Takashi and Kuboki, Koji and Morita, Toshisuke and Sugihara, Hitoshi and Hirose, Takahisa and Minami, Shiro and Yoshino, Gen
Journal: Growth hormone & IGF research : official journal of the Growth Hormone Research Society and the International IGF Research Socie (2020): 101334
Authors: Bonnet, Udo and Wiemann, Martin
Journal: CNS & neurological disorders drug targets (2020): 264-275
Authors: Yemelyanov, Vladislav V and Chirkova, Tamara V and Shishova, Maria F and Lindberg, Sylvia M
Journal: Plants (Basel, Switzerland) (2020)
Authors: Xu, Zhi-Shen and Li, Feng-Jun and Hide, Geoff and Lun, Zhao-Rong and Lai, De-Hua
Journal: Parasites & vectors (2020): 214
Authors: Voronina, Polina P and Adamovich, Ksenia V and Adamovich, Tatyana V and Dubouskaya, Tatsiana G and Hrynevich, Sviatlana V and Waseem, Tatsiana V and Fedorovich, Sergei V
Journal: Journal of molecular neuroscience : MN (2020): 112-119
Authors: Hayes, Elaine and Murphy, Mark P and Pohl, Kerstin and Browne, Niall and McQuillan, Karen and Saw, Le Er and Foley, Clare and Gargoum, Fatma and McElvaney, Oliver J and Hawkins, Padraig and Gunaratnam, Cedric and McElvaney, Noel G and Reeves, Emer P
Journal: Frontiers in immunology (2020): 600033
Application notes
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FAQ
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