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RatioWorks™ PH165, AM

RatioWorks™ PH165 AM is a novel fluorescent probe that can be used to measure intracellular pH in live cells. It shows pH-dependent fluorescence spectra. Its dual excitation and dual emission make it a unique ratiometric pH probe for analyzing live cells. At low pH, RatioWorks™ PH165 is weakly fluorescent in far red spectrum region and highly fluorescent in red spectrum region. As pH increases, it shows higher fluorescence in far red spectrum and lower fluorescence in red spectrum. This probe can quantify cytosolic pH in the range from 4 to 9 with dual excitation/emission at 497/594 nm and 578/654 nm respectively. The pKa of PH165 is ~7.1. The red fluorescence spectrum of RatioWorks™ PH165 makes it suitable for multiplexing tests with green fluorophores such as GFP. RatioWorks™ PH165 AM is cell-permeable, and compatible with various platforms such as fluorescence microscopy and microplate reader.

Spectrum

Product family

NameExcitation (nm)Emission (nm)
RatioWorks™ BCFL, AM *Superior replacement for BCECF*504527

Citations

View all 3 citations: Citation Explorer
Intracellular pH Measurements in Glioblastoma Cells Using the pH-Sensitive Dye BCECF
Authors: Galifianakis, N. V., Placantonakis, D. G., Chesler, M.
Journal: Methods Mol Biol (2018): 103-109
Evaluation of BCECF fluorescence ratio imaging to properly measure gastric intramucosal pH variations in vivo
Authors: Rochon, P., Jourdain, M., Mangalaboyi, J., Fourrier, F., Soulie-Begu, S., Buys, B., Dehlin, G., Lesage, J. C., Chambrin, M. C., Mordon, S.
Journal: J Biomed Opt (2007): 064014
Calibration methods and avoidance of errors in measurement of intracellular pH (pHcyt) using the indicator bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF) in human platelets
Authors: Valant, P. A., Haynes, D. H.
Journal: J Fluoresc (1992): 191-206

References

View all 1 references: Citation Explorer
Estimation of intramitochondrial pca and ph by fura-2 and 2,7 biscarboxyethyl-5(6)-carboxyfluorescein (bcecf) fluorescence
Authors: Davis MH, Altschuld RA, Jung DW, Brierley GP.
Journal: Biochem Biophys Res Commun (1987): 40
Page updated on October 28, 2024

Ordering information

Price
Unit size
Catalog Number21212
Quantity
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Additional ordering information

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Physical properties

Molecular weight

583.53

Solvent

DMSO

Spectral properties

Excitation (nm)

463

Emission (nm)

641

Storage, safety and handling

H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22

Storage

Freeze (< -15 °C); Minimize light exposure
UNSPSC12171501

Platform

Fluorescence microscope

ExcitationCy3-Cy5 filter set
EmissionCy3-Cy5 filter set
Recommended plateBlack wall, clear bottom
Instrument specification(s)Cy3-Cy5 filter set

Fluorescence microplate reader

Excitation497-578 nm
Emission594-654 nm
Cutoff570-630 nm
Recommended plateBlack wall, clear bottom
Instrument specification(s)Bottom read mode
Ratiometric analysis of HeLa cells using RatioWorks&trade; PH165, AM with fluorescence microscopy. HeLa cells were stained with RatioWorks&trade; PH165, AM for 90 minutes at 37&deg;C and shifted to solutions with different pH. Images were acquired using Cy3/TRITC (green, pseudo color) and Cy5 filter sets (red color).
Ratiometric analysis of HeLa cells using RatioWorks&trade; PH165, AM with fluorescence microscopy. HeLa cells were stained with RatioWorks&trade; PH165, AM for 90 minutes at 37&deg;C and shifted to solutions with different pH. Images were acquired using Cy3/TRITC (green, pseudo color) and Cy5 filter sets (red color).
Ratiometric analysis of HeLa cells using RatioWorks&trade; PH165, AM with fluorescence microscopy. HeLa cells were stained with RatioWorks&trade; PH165, AM for 90 minutes at 37&deg;C and shifted to solutions with different pH. Images were acquired using Cy3/TRITC (green, pseudo color) and Cy5 filter sets (red color).
Ratiometric analysis of HeLa cells using RatioWorks&trade; PH165, AM with microplate reader. HeLa cells were stained with RatioWorks&trade; PH165, AM for 30 minutes at 37&deg;C and shifted to solutions with different pH. RFU was measured using Ex/Em= 497/594 nm and 578/654 nm using fluorescence microplate reader and ratio for 594/654 nm was calculated and plotted.