Fura-2, AM UltraPure Grade CAS 108964-32-5

Fluorescence excitation spectra of Fura-2 in solutions conctaining 0 to 39uM free Ca2+.

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Physical properties

Dissociation constant (K_d, nM)	145
Molecular weight	1001.86
Solvent	DMSO

Spectral properties

Excitation (nm)	336
Emission (nm)	505

Storage, safety and handling

Certificate of Origin	Download PDF
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

Alternative formats

Fura-2, AM *CAS 108964-32-5*

Overview SDS Protocol

CAS

108964-32-5

Molecular weight

1001.86

Dissociation constant (K_d, nM)

145

Excitation (nm)

336

Emission (nm)

505

Among the ratiometric calcium indicators, Fura-2 and Indo-1 are most commonly used. Fura-2 is excitation-ratioable while Indo-1 is emission-ratioable. Fura-2 is preferred for ratio-imaging microscopy, in which it is more practical to change excitation wavelengths than emission wavelengths. Upon binding Ca2+, Fura-2 exhibits an absorption shift that can be observed by scanning the excitation spectrum between 300 and 400 nm, while monitoring the emission at ~510 nm. Fura-2, AM is a cell-permeable calcium indicator that is emission-ratiometric and UV light'excitable. This AM ester form can be loaded into live cells noninvasively.

Platform

Fluorescence microscope

Excitation	Fura 2 filter set
Emission	Fura 2 filter set
Recommended plate	Black wall/clear bottom

Fluorescence microplate reader

Excitation	340, 380
Emission	510
Cutoff	475
Recommended plate	Black wall/clear bottom
Instrument specification(s)	Bottom read mode/Programmable liquid handling

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.

Fura-2 AM *UltraPure Grade* Stock Solution

Prepare a 2 to 5 mM stock solution of Fura-2 AM in high-quality, anhydrous DMSO.

PREPARATION OF WORKING SOLUTION

Fura-2 AM *UltraPure Grade* Working Solution

On the day of the experiment, either dissolve Fura-2 AM in DMSO or thaw an aliquot of the indicator stock solution to room temperature. Prepare a dye working solution of 2 to 20 µM in a buffer of your choice (e.g., Hanks and Hepes buffer) with 0.04% Pluronic® F-127. For most cell lines, Fura-2 AM at a final concentration of 4-5 μM is recommended. The exact concentration of indicators required for cell loading must be determined empirically.
Note The nonionic detergent Pluronic® F-127 is sometimes used to increase the aqueous solubility of Fura-2 AM. A variety of Pluronic® F-127 solutions can be purchased from AAT Bioquest.
Note If your cells contain organic anion-transporters, probenecid (1-2 mM) may be added to the dye working solution (final in well concentration will be 0.5-1 mM) to reduce leakage of the de-esterified indicators. A variety of ReadiUse™ probenecid products, including water-soluble, sodium salt, and stabilized solution, can be purchased from AAT Bioquest.

SAMPLE EXPERIMENTAL PROTOCOL

Following is our recommended protocol for loading AM esters into live cells. This protocol only provides a guideline and should be modified according to your specific needs.

Prepare cells in growth medium overnight.
On the next day, add 1X Fura-2 AM working solution into your cell plate.
Note If your compound(s) interfere with the serum, replace the growth medium with fresh HHBS buffer before dye-loading.
Incubate the dye-loaded plate in a cell incubator at 37 °C for 30 to 60 minutes.
Note Incubating the dye for longer than 1 hour can improve signal intensities in certain cell lines.
Replace the dye working solution with HHBS or buffer of your choice (containing an anion transporter inhibitor, such as 1 mM probenecid, if applicable) to remove any excess probes.
Add the stimulant as desired and simultaneously measure fluorescence using either a fluorescence microscope equipped with a Fura 2 filter set or a fluorescence plate reader containing a programmable liquid handling system such as a FlexStation, at Ex/Em₁ = 340/510 nm cutoff 475 nm and Ex/Em₂ = 380/510 nm cutoff 475 nm.

Calculators

Common stock solution preparation

Table 1. Volume of DMSO needed to reconstitute specific mass of Fura-2, AM *UltraPure Grade* *CAS 108964-32-5* to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.

	0.1 mg	0.5 mg	1 mg	5 mg	10 mg
1 mM	99.814 µL	499.072 µL	998.143 µL	4.991 mL	9.981 mL
5 mM	19.963 µL	99.814 µL	199.629 µL	998.143 µL	1.996 mL
10 mM	9.981 µL	49.907 µL	99.814 µL	499.072 µL	998.143 µL

Molarity calculator

Enter any two values (mass, volume, concentration) to calculate the third.

Mass (Calculate)		Molecular weight		Volume (Calculate)		Concentration (Calculate)		Moles
	/		=		x		=

Spectrum

Open in Advanced Spectrum Viewer

Spectral properties

Excitation (nm)	336
Emission (nm)	505

Product Family

Name	Excitation (nm)	Emission (nm)
Fura-8FF™, AM	354	524
Fura-FF, AM [Fura-2FF, AM] CAS 348079-12-9	336	505
Fura Red, AM CAS 149732-62-7	435	639
Fura-8™, AM	354	524
Rhod-2, AM CAS#: 145037-81-6	553	577
Rhod-2, AM UltraPure Grade CAS#: 145037-81-6	553	577
Fura-10™, AM	354	524

Images

Figure 1. Fluorescence excitation spectra of Fura-2 in solutions conctaining 0 to 39uM free Ca2+.

Citations

View all 13 citations: Citation Explorer

Inhibitory responses to retinohypothalamic tract stimulation in the circadian clock of the diurnal rodent Rhabdomys pumilio
Authors: Schoonderwoerd, Robin A and de Torres Guti{\'e}rrez, Pablo and Blommers, Ruben and van Beurden, Anouk W and Coenen, Tineke CJJ and Klett, Nathan J and Michel, Stephan H and Meijer, Johanna H
Journal: The FASEB Journal (2022): e22415

IKCa channels control breast cancer metabolism including AMPK-driven autophagy
Authors: Gross, Dominic and Bischof, Helmut and Maier, Selina and Sporbeck, Katharina and Birkenfeld, Andreas L and Malli, Roland and Ruth, Peter and Proikas-Cezanne, Tassula and Lukowski, Robert
Journal: Cell death \& disease (2022): 1--14

Transmembrane Domain 3 Is a Transplantable Pharmacophore in the Photodynamic Activation of Cholecystokinin 1 Receptor
Authors: Li, Yuan and Cui, Zong Jie
Journal: ACS Pharmacology \& Translational Science (2022)

Catalpol attenuates renal injury by regulating oxidative stress and inflammation response
Authors: Liu, Zhihui and Wang, Yu and Zhou, Chong and Xu, Qingyang and Gao, Hongxin and Huo, Mohan and Jiang, Xiaowen and Yu, Wenhui
Journal: (2022)

NanoLuc Bioluminescence-Driven Photodynamic Activation of Cholecystokinin 1 Receptor with Genetically-Encoded Protein Photosensitizer MiniSOG
Authors: Li, Yuan and Cui, Zong Jie
Journal: International Journal of Molecular Sciences (2020): 3763

Photodynamic Activation of Cholecystokinin 1 Receptor with Different Genetically Encoded Protein Photosensitizers and from Varied Subcellular Sites
Authors: Li, Yuan and Cui, Zong Jie
Journal: Biomolecules (2020): 1423

Aspirin eugenol ester attenuates oxidative injury of vascular endothelial cells by regulating NOS and Nrf2 signaling pathways
Authors: Huang, Mei-Zhou and Yang, Ya-Jun and Liu, Xi-Wang and Qin, Zhe and Li, Jian-Yong
Journal: British Journal of Pharmacology (2019)

Effect of PEGylated Magnetic PLGA-PEI Nanoparticles on Primary Hippocampal Neurons: Reduced Nano-neurotoxicity and Enhanced Transfection Efficiency with Magnetofection
Authors: Cui, Yanna and Li, Xiao and Zeljic, Kristina and Shan, Shifang and Qiu, Zilong and Wang, Zheng
Journal: ACS Applied Materials & Interfaces (2019)

Periprosthetic hypoxia as consequence of TRPM7 mediated cobalt influx in osteoblasts
Authors: Römmelt, Constantin and Munsch, Thomas and Drynda, Andreas and Lessmann, Volkmar and Lohmann, Christoph H and Bertr, undefined and , Jessica
Journal: Journal of Biomedical Materials Research Part B: Applied Biomaterials (2018)

An essential role of NAD (P) H oxidase 2 in UVA-induced calcium oscillations in mast cells
Authors: Li, Zhi Ying and Jiang, Wen Yi and Cui, Zong Jie
Journal: Photochemical \& Photobiological Sciences (2015): 414--428

References

View all 119 references: Citation Explorer

Load of calcium probe Fura -2/AM in Escherichia coli cells
Authors: Shao M, Wang HM, Liu ZH, Shen P, Cai RX.
Journal: Wei Sheng Wu Xue Bao (2005): 805

An Excel-based model of Ca2+ diffusion and fura 2 measurements in a spherical cell
Authors: McHugh JM, Kenyon JL.
Journal: Am J Physiol Cell Physiol (2004): C342

Problems caused by high concentration of ATP on activation of the P2X7 receptor in bone marrow cells loaded with the Ca2+ fluorophore fura-2
Authors: Paredes-Gamero EJ, Franca JP, Moraes AA, Aguilar MO, Oshiro ME, Ferreira AT.
Journal: J Fluoresc (2004): 711

Photonic crystal fibre enables short-wavelength two-photon laser scanning fluorescence microscopy with fura-2
Authors: McConnell G, Riis E.
Journal: Phys Med Biol (2004): 4757

Abnormal spectra alteration observed in Triton calibration method for measuring [Ca2+]i with fluorescence indicator, fura-2
Authors: Xu T, Yang W, Huo XL, Song T.
Journal: J Biochem Biophys Methods (2004): 219

Two-photon microscopy of fura-2-loaded cardiac myocytes with an all-solid-state tunable and visible femtosecond laser source
Authors: McConnell G, Smith GL, Girkin JM, Gurney AM, Ferguson AI.
Journal: Opt Lett (2003): 1742

AMPA-induced Ca(2+) influx in cultured rat cortical nonpyramidal neurones: pharmacological characterization using fura-2 microfluorimetry
Authors: Fischer W, Franke H, Scheibler P, Allgaier C, Illes P.
Journal: Eur J Pharmacol (2002): 53

Selective measurement of endothelial or smooth muscle [Ca(2+)](i) in pressurized/perfused cerebral arteries with fura-2
Authors: Marrelli SP., undefined
Journal: J Neurosci Methods (2000): 145

Excitation wavelengths for fura 2 provide a linear relationship between [Ca(2+)] and fluorescence ratio
Authors: Palmer BM, Moore RL.
Journal: Am J Physiol Cell Physiol (2000): C1278

Tyrosine kinase inhibitors and Ca2+ signaling: direct interactions with fura-2
Authors: Berts A, Minneman KP.
Journal: Eur J Pharmacol (2000): 35