Fura-10™, AM

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Additional ordering information

Telephone	1-800-990-8053
Fax	1-800-609-2943
Email	sales@aatbio.com
Quotation	Request
International	See distributors
Shipping	Standard overnight for United States, inquire for international

Physical properties

Dissociation constant (K_d, nM)	260
Molecular weight	1194.14
Solvent	DMSO

Spectral properties

Excitation (nm)	354
Emission (nm)	524

Storage, safety and handling

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	12171501

Overview SDS Protocol

Molecular weight

1194.14

Dissociation constant (K_d, nM)

260

Excitation (nm)

354

Emission (nm)

524

Among ratiometric calcium ion indicators, Fura-2 and Indo-1 are the two most popular ones. However, there are still a few challenges for using these two calcium ion indicators, in particular, for live cells. UV-excitation of Fura 2 caused fast photobleaching. Fura-8™ was introduced a few years ago to shift the excitation closer to visible light. Although Fura-8 demonstrated significant improvement in the ratio of signal/background, it is not well retained in live cells just like Fura-2. Fura-10 have recently been introduced to address this cellular retention issue. Fura 10 demonstrated dramatic improvement in the ratio of signal/background in the absence of probenecid.

Platform

Fluorescence microplate reader

Excitation	354 nm and 415 nm
Emission	524 nm
Cutoff	475 nm
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-10™ AM stock solution

Prepare a 2 to 5 mM Fura-10™ AM stock solution in high-quality, anhydrous DMSO.

PREPARATION OF WORKING SOLUTION

Fura-10™ AM working solution

On the day of the experiment, either dissolve Fura-10™ 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-10™ 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-10™ 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-10™ 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 monitor fluorescence intensity using a fluorescence plate reader containing a programmable liquid handling system such as a FlexStation, at Ex/Em₁ = 354/524 nm cutoff 475 nm and Ex/Em₂ = 415/524 nm cutoff 475 nm.

Calculators

Common stock solution preparation

Table 1. Volume of DMSO needed to reconstitute specific mass of Fura-10™, AM 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	83.742 µL	418.711 µL	837.423 µL	4.187 mL	8.374 mL
5 mM	16.748 µL	83.742 µL	167.485 µL	837.423 µL	1.675 mL
10 mM	8.374 µL	41.871 µL	83.742 µL	418.711 µL	837.423 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)	354
Emission (nm)	524

Product Family

Name	Excitation (nm)	Emission (nm)
Fura-8FF™, AM	354	524
Fura-2, AM CAS 108964-32-5	336	505
Fura-2, AM UltraPure Grade CAS 108964-32-5	336	505
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

Images

Figure 1. ATP-stimulated calcium response of endogenous P2Y receptor in CHO-K1 cells measured with Fura-2 AM, Fura-8™ AM and Fura-10™ AM in the absence of Probenecid. CHO-K1cells were seeded overnight in 50,000 cells per 100 µL per well in a 96-well black wall/clear bottom costar plate. 100 µL of 5 uM Fura-2 AM or Fura-8™ AM or Fura-10™ AM without probenecid was added into the cells, and the cells were incubated at 37 ^oC for 45 minutes and RT for 30 minutes. ATP (50µL/well) was added by FlexStation (Molecular Devices) to achieve the final indicated concentrations.

References

View all 50 references: Citation Explorer

Rumex acetosa modulates platelet function and inhibits thrombus formation in rats.
Authors: Jeong, Dahye and Irfan, Muhammad and Lee, Dong-Ha and Hong, Seung-Bok and Oh, Jae-Wook and Rhee, Man Hee
Journal: BMC complementary medicine and therapies (2020): 98

High-salt intake increases TRPC3 expression and enhances TRPC3-mediated calcium influx and systolic blood pressure in hypertensive patients.
Authors: Hu, Yingru and Xia, Weijie and Li, Yingsha and Wang, Qianran and Lin, Shaoyang and Wang, Bin and Zhou, Cui and Cui, Yuanting and Jiang, Yanli and Pu, Xiaona and Wei, Xiao and Wu, Hao and Zhang, Hengshu and Zhu, Zhiming and Liu, Daoyan and Li, Zhiyong
Journal: Hypertension research : official journal of the Japanese Society of Hypertension (2020)

Reduced store-operated Ca2+ entry impairs mesenteric artery function in response to high external glucose in type 2 diabetic ZDF rats.
Authors: Schach, Christian and Wester, Michael and Leibl, Florian and Redel, Andreas and Gruber, Michael and Maier, Lars S and Endemann, Dierk and Wagner, Stefan
Journal: Clinical and experimental pharmacology & physiology (2020)

Acupotomy Alleviates Energy Crisis at Rat Myofascial Trigger Points.
Authors: Zhang, Yi and Du, Ning-Yu and Chen, Chen and Wang, Tong and Wang, Li-Juan and Shi, Xiao-Lu and Li, Shu-Ming and Guo, Chang-Qing
Journal: Evidence-based complementary and alternative medicine : eCAM (2020): 5129562

Pharmacological and genetic characterisation of the canine P2X4 receptor.
Authors: Sophocleous, Reece A and Berg, Tracey and Finol-Urdaneta, Rocio K and Sluyter, Vanessa and Keshiya, Shikara and Bell, Lachlan and Curtis, Stephen J and Curtis, Belinda L and Seavers, Aine and Bartlett, Rachael and Dowton, Mark and Stokes, Leanne and Ooi, Lezanne and Sluyter, Ronald
Journal: British journal of pharmacology (2020)

Histamine induces intracellular Ca2+ oscillations and nitric oxide release in endothelial cells from brain microvascular circulation.
Authors: Berra-Romani, Roberto and Faris, Pawan and Pellavio, Giorgia and Orgiu, Matteo and Negri, Sharon and Forcaia, Greta and Var-Gaz-Guadarrama, Verónica and Garcia-Carrasco, Mario and Botta, Laura and Sancini, Giulio and Laforenza, Umberto and Moccia, Francesco
Journal: Journal of cellular physiology (2020): 1515-1530

Low concentration of morphine protects against cell death, oxidative stress and calcium accumulation by nicotine in PC12 cells.
Authors: Amini, K and Zhaleh, H and Tahvilian, R and Farnia, V
Journal: Bratislavske lekarske listy (2019): 256-262

NHE8 attenuates Ca2+ influx into NRK cells and the proximal tubule epithelium.
Authors: Wiebe, Shane A and Plain, Allein and Pan, Wanling and O'Neill, Debbie and Braam, Branko and Alexander, R Todd
Journal: American journal of physiology. Renal physiology (2019): F240-F253

A3 receptor agonist, Cl-IBMECA, potentiate glucose-induced insulin secretion from MIN6 insulinoma cells possibly through transient Ca2+ entry.
Authors: Keyvanloo Shahrestanaki, Mohammad and Aghaei, Mahmoud
Journal: Research in pharmaceutical sciences (2019): 107-114

TRPV4 plays an important role in rat prefrontal cortex changes induced by acute hypoxic exercise.
Authors: Huang, Xing and Hu, Yanxin and Zhao, Li and Gu, Boya and Zhu, Rongxin and Li, Yan and Yang, Yun and Han, Tianyu and Yu, Jiabei and Mu, Lianwei and Han, Peng and Li, Cui and Zhang, Weijia and Hu, Yang
Journal: Saudi journal of biological sciences (2019): 1194-1206