Fura-8FF™, AM

Fluorescence excitation spectra of Fura-8™ in the presence of 0 to 39 µM free Ca2+.

Ordering information

Price
Catalog Number
Unit Size
Quantity

Add to cart

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)	6000
Molecular weight	973.86
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	12352200

Overview SDS Protocol

Molecular weight

973.86

Dissociation constant (K_d, nM)

6000

Excitation (nm)

354

Emission (nm)

524

The cell-permeant Fura-8FF AM is an analog of Fura-8 AM with much lower calcium binding affinity, Kd ~10 µM. Fura-8FF has its emission shifted into longer visible wavelength that is compatible with the common filter sets. Fura-8FF™ AM is more sensitive to calcium than Fura-2FF AM with higher signal/background ratio than that of Fura-2FF AM. e., calculating the excitation intensity ratios at 354 nm and 415 nm by monitoring emission intensity at 530 nm.

Platform

Fluorescence microscope

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

Fluorescence microplate reader

Excitation	355, 415
Emission	530
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-8FF™ AM Stock Solution

Prepare a 2 to 5 mM stock solution of Fura-8FF™ AM in high-quality, anhydrous DMSO.

PREPARATION OF WORKING SOLUTION

Fura-8FF™ AM Working Solution

On the day of the experiment, either dissolve Fura-8FF™ 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-8FF™ 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-8FF™ 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-8FF™ 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 = 355/530 nm cutoff 475 nm and Ex/Em₂ = 415/530 nm cutoff 475 nm.

Calculators

Common stock solution preparation

Table 1. Volume of DMSO needed to reconstitute specific mass of Fura-8FF™, 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	102.684 µL	513.421 µL	1.027 mL	5.134 mL	10.268 mL
5 mM	20.537 µL	102.684 µL	205.368 µL	1.027 mL	2.054 mL
10 mM	10.268 µL	51.342 µL	102.684 µL	513.421 µL	1.027 mL

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-8™, 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
Fluo-8FF™, AM	495	516
Fura-10™, AM	354	524

Images

Figure 1. Fluorescence excitation spectra of Fura-8™ in the presence of 0 to 39 µM free Ca2+.

Citations

View all 2 citations: Citation Explorer

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

Lasting inhibition of receptor-mediated calcium oscillations in pancreatic acini by neutrophil respiratory burst--A novel mechanism for secretory blockade in acute pancreatitis?
Authors: Liang, Hui Yuan and Song, Zhi Min and Cui, Zong Jie
Journal: Biochemical and biophysical research communications (2013): 361--367

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