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CAS:348079-12-9
Storage:Powder:-20℃，2 years;Insolvent(Mother Liquid):-20℃，6 months;-80℃，1 year
Purity:≥95%
Appearance:Solid
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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.? The cell-permeant Fura-2FF AM is an analog of Fura-2 AM with much lower calcium binding affinity.? This AM ester form can be loaded into live cells noninvasively. ? ?
Fluorescence microscope（for reference only）
Excitation:Fura 2 filter set
Emission:Fura 2 filter set
Recommended plate:Black wall/clear bottom
Fluorescence microplate reader（for reference only）
Excitation:340, 380
Emission:510
Cutoff:475
Recommended plate:Black wall/clear bottom
Instrument specification(s)：Bottom read mode/Programmable liquid handling
Example protocol（For reference only）
Fura-FF AM Stock Solution
Prepare a 2 to 5 mM stock solution of Fura-FF AM in high-quality, anhydrous DMSO.
If it is necessary to prepare a stock solution, it is recommended to store in aliquots to avoid product failure caused by repeated freezing and thawing.
Fura-FF AM Working Solution
On the day of the experiment, either dissolve Fura-FF 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-FF 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-FF AM.
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.
SAMPLE EXPERIMENTAL PROTOCOL（For reference only）
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-FF 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 ℃ 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.
