Fluo-8FF™, AM
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Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
sales@aatbio.com | |
International | See distributors |
Bulk request | Inquire |
Custom size | Inquire |
Shipping | Standard overnight for United States, inquire for international |
Dissociation constant (Kd, nM) | 10000 |
Molecular weight | 1082.91 |
Solvent | DMSO |
Correction Factor (260 nm) | 1.076 |
Correction Factor (280 nm) | 0.769 |
Extinction coefficient (cm -1 M -1) | 23430 |
Excitation (nm) | 495 |
Emission (nm) | 516 |
Quantum yield | 0.161 |
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 | SDSProtocol |
Molecular weight 1082.91 | Dissociation constant (Kd, nM) 10000 | Correction Factor (260 nm) 1.076 | Correction Factor (280 nm) 0.769 | Extinction coefficient (cm -1 M -1) 23430 | Excitation (nm) 495 | Emission (nm) 516 | Quantum yield 0.161 |
Platform
Fluorescence microscope
Excitation | FITC |
Emission | FITC |
Recommended plate | Black wall/clear bottom |
Fluorescence microplate reader
Excitation | 490 |
Emission | 525 |
Cutoff | 515 |
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
Prepare a 2 to 5 mM stock solution of Fluo-8FF™ AM in high-quality, anhydrous DMSO.
PREPARATION OF WORKING SOLUTION
On the day of the experiment, either dissolve Fluo-8FF™ AM in DMSO or thaw an aliquot of the indicator stock solution to room temperature.
Prepare a 2 to 20 µM Fluo-8FF™ AM working solution in a buffer of your choice (e.g., Hanks and Hepes buffer) with 0.04% Pluronic® F-127. For most cell lines, Fluo-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 Fluo-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 solutions, 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 Fluo-8FF™ AM working solution to 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 2 hours 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 FITC filter set or a fluorescence plate reader containing a programmable liquid handling system such as an FDSS, FLIPR, or FlexStation, at 490/525 nm cutoff 515 nm.
Calculators
Common stock solution preparation
0.1 mg | 0.5 mg | 1 mg | 5 mg | 10 mg | |
1 mM | 92.344 µL | 461.719 µL | 923.438 µL | 4.617 mL | 9.234 mL |
5 mM | 18.469 µL | 92.344 µL | 184.688 µL | 923.438 µL | 1.847 mL |
10 mM | 9.234 µL | 46.172 µL | 92.344 µL | 461.719 µL | 923.438 µL |
Molarity calculator
Mass (Calculate) | Molecular weight | Volume (Calculate) | Concentration (Calculate) | Moles | ||||
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Spectrum
Spectral properties
Correction Factor (260 nm) | 1.076 |
Correction Factor (280 nm) | 0.769 |
Extinction coefficient (cm -1 M -1) | 23430 |
Excitation (nm) | 495 |
Emission (nm) | 516 |
Quantum yield | 0.161 |
Product Family
Name | Excitation (nm) | Emission (nm) | Extinction coefficient (cm -1 M -1) | Quantum yield |
Fluo-8®, AM | 495 | 516 | 23430 | 0.161 |
Fluo-8H™, AM | 495 | 516 | 23430 | 0.161 |
Fluo-8L™, AM | 495 | 516 | 23430 | 0.161 |
Fluo-4 AM *Ultrapure Grade* *CAS 273221-67-3* | 495 | 528 | 82000 | 0.161 |
Fluo-3, AM *CAS 121714-22-5* | 506 | 515 | 86,0001 | 0.151 |
Fluo-3, AM *UltraPure grade* *CAS 121714-22-5* | 506 | 515 | 86,0001 | 0.151 |
Fluo-3, AM *Bulk package* *CAS 121714-22-5* | 506 | 515 | 86,0001 | 0.151 |
Fluo-3FF, AM *UltraPure grade* *Cell permeant* | 506 | 515 | 86,0001 | 0.151 |
Fluo-5F, AM *Cell permeant* | 494 | 516 | - | - |
Show More (2) |
Images
Citations
Authors: Czekalska, Magdalena A and Kaminski, Tomasz S and Makuch, Karol and Garstecki, Piotr
Journal: Sensors and Actuators B: Chemical (2019)
Authors: Ayala, Mónica Rubio and Syrovets, Tatiana and Hafner, Susanne and Zablotskii, Vitalii and Dejneka, Alex and r , undefined and Simmet, Thomas
Journal: Biomaterials (2018)
Authors: Minami, K and Hayashi, T and Sato, K and Nakahara, T
Journal: Biomedical microdevices (2018): 10
Authors: Huang, Chao and Li, Na and Yuan, Shengwu and Ji, Xiaoya and Ma, Mei and Rao, Kaifeng and Wang, Zijian
Journal: Environmental Pollution (2017): 775--786
Authors: Sun, Qiaoling and Choudhary, Saba and Mannion, Ciaran and Kissin, Yair and Zilberberg, Jenny and Lee, Woo Y
Journal: Bone (2017)
Authors: Iwamoto, Satoshi and Koga, Tomoaki and Ohba, Mai and Okuno, Toshiaki and Koike, Masato and Murakami, Akira and Matsuda, Akira and Yokomizo, Takehiko
Journal: Scientific Reports (2017)
Authors: Ishii, Masaaki and Rohrer, Bärbel
Journal: Cell Death Discovery (2017): 16071
Authors: Liu, ZhongJie and Ma, ChangQing and Zhao, Wei and Zhang, QingGuo and Xu, Rui and Zhang, HongFei and Lei, HongYi and Xu, ShiYuan
Journal: Neurochemical Research (2017): 1--14
Authors: Takada, Hiroya and Yonekawa, Jun and Matsumoto, Masami and Furuya, Kishio and Sokabe, Masahiro
Journal: BioMed Research International (2017)
Authors: Schmunk, Galina and Nguyen, Rachel L and Ferguson, David L and Kumar, Kenny and Parker, Ian and Gargus, J Jay
Journal: Scientific Reports (2017): 40740
Application notes
A Meta-Analysis of Common Calcium Indicators
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A New Robust No-Wash FLIPR Calcium Assay Kit for Screening GPCR and Calcium Channel Targets
A Novel NO Wash Probeniceid-Free Calcium Assay for Functional Analysis of GPCR and Calcium Channel Targets
FAQ
Are there any calcium indicators that don't require probenecid (PBC)?
Are there any substitutes for probenecid in calcium assays?
Are there upgraded trypan blue derivatives for cell viability testing?
Can I intracellularly measure mitochondria calcium flux and changes in mitochondria membrane potential at the same time?