Cal-500™ AM
Price | |
Catalog Number | |
Unit Size | |
Quantity |
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) | 303 |
Molecular weight | 1019.92 |
Solvent | DMSO |
Excitation (nm) | 388 |
Emission (nm) | 482 |
Quantum yield | 0.481 |
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 1019.92 | Dissociation constant (Kd, nM) 303 | Excitation (nm) 388 | Emission (nm) 482 | Quantum yield 0.481 |
Platform
Fluorescence microscope
Excitation | DAPI |
Emission | DAPI |
Recommended plate | Black wall/clear bottom |
Fluorescence microplate reader
Excitation | 400 |
Emission | 500 |
Cutoff | 470 |
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 Cal-500™ AM in anhydrous DMSO.
Note: When reconstituted in DMSO, Cal-500™ AM is a clear, colorless solution.
PREPARATION OF WORKING SOLUTION
On the day of the experiment, either dissolve Cal-500™ AM in DMSO or thaw an aliquot of the indicator stock solution to room temperature.
Prepare a 2 to 20 µM Cal-500™ 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, Cal-500™ 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 Cal-500™ 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 Cal-500™ 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 DAPI filter set or a fluorescence plate reader containing a programmable liquid handling system such as an FDSS, FLIPR, or FlexStation, at Ex/Em = 400/500 nm cutoff 470 nm.
Calculators
Common stock solution preparation
0.1 mg | 0.5 mg | 1 mg | 5 mg | 10 mg | |
1 mM | 98.047 µL | 490.235 µL | 980.469 µL | 4.902 mL | 9.805 mL |
5 mM | 19.609 µL | 98.047 µL | 196.094 µL | 980.469 µL | 1.961 mL |
10 mM | 9.805 µL | 49.023 µL | 98.047 µL | 490.235 µL | 980.469 µL |
Molarity calculator
Mass (Calculate) | Molecular weight | Volume (Calculate) | Concentration (Calculate) | Moles | ||||
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Spectrum
Spectral properties
Excitation (nm) | 388 |
Emission (nm) | 482 |
Quantum yield | 0.481 |
Product Family
Name | Excitation (nm) | Emission (nm) | Quantum yield |
Cal-590™ AM | 574 | 588 | 0.621 |
Cal-630™ AM | 609 | 626 | 0.371 |
Cal-520®, AM | 492 | 515 | 0.751 |
Cal-520FF™, AM | 492 | 515 | 0.751 |
Cal-520N™, AM | 492 | 515 | 0.751 |
Images
Citations
Authors: Wu, Yanjiao and Xu, Xiaoli and Ma, Lunkun and Yi, Qian and Sun, Weichao and Tang, Liling
Journal: The International Journal of Biochemistry & Cell Biology (2017)
Authors: Lu, Jiang and Yao, Xue-qin and Luo, Xin and Wang, Yu and Chung, Sookja Kim and Tang, He-xin and Cheung, Chi Wai and Wang, Xian-yu and Meng, Chen and Li, Qing and others, undefined
Journal: Neural Regeneration Research (2017): 945
Authors: Yang, Gang and Xiao, Zhenghua and Ren, Xiaomei and Long, Haiyan and Ma, Kunlong and Qian, Hong and Guo, Yingqiang
Journal: Scientific Reports (2017): 41781
Authors: Liu, Jia and Du, Qing and Zhu, He and Li, Yu and Liu, Maodong and Yu, Shoushui and Wang, Shilei
Journal: Int J Clin Exp Med (2017): 6861--6868
Authors: Sun, Xia and Lin, Yi and Huang, Qiansheng and Shi, Junpeng and Qiu, Ling and Kang, Mei and Chen, Yajie and Fang, Chao and Ye, Ting and Dong, Sijun
Journal: Journal of cellular and molecular medicine (2015): 581--594
Authors: Zhao, Lantao and Li, Shuhong and Wang, Shilei and Yu, Ning and Liu, Jia
Journal: Biochemical and biophysical research communications (2015): 537--542
Authors: Liang, Nan and Wang, Peng and Wang, Shilei and Li, Shuhong and Li, Yu and Wang, Jinying and Wang, Min
Journal: Journal of Neural Transmission (2014): 593--600
Authors: Li, Qing and Lu, Jiang and Wang, Xianyu and others, undefined
Journal: Neural regeneration research (2014): 2002
Authors: Peng, Xu-Dong and Zhao, Gui-Qiu and Lin, Jing and Jiang, Nan and Xu, Qiang and Zhu, Cheng-Cheng and Qu, Jain-Qiu and Cong, Lin and Li, Hui
Journal: International journal of ophthalmology (2014): 441--447
Authors: Hagiyama, M and Inoue, T and Furuno, T and Iino, T and Itami, S and Nakanishi, M and Asada, H and Hosokawa, Y and Ito, A
Journal: British Journal of Dermatology (2013): 771--778
References
Authors: Bailey S, Macardle PJ.
Journal: J Immunol Methods (2006): 220
Authors: Orlicky J, Sulova Z, Dovinova I, Fiala R, Zahradnikova A, Jr., Breier A.
Journal: Gen Physiol Biophys (2004): 357
Authors: Patel H, Porter RH, Palmer AM, Croucher MJ.
Journal: Br J Pharmacol (2003): 671
Authors: Loughrey CM, MacEachern KE, Cooper J, Smith GL.
Journal: Cell Calcium (2003): 1
Authors: Zhang Z, Kitching P.
Journal: J Virol Methods (2000): 187
Authors: Rockwell PL, Storey BT.
Journal: Mol Reprod Dev (2000): 335
Authors: Cantz T, Nies AT, Brom M, Hofmann AF, Keppler D.
Journal: Am J Physiol Gastrointest Liver Physiol (2000): G522
Authors: Walczysko P, Wagner E, Albrechtova JT.
Journal: Cell Calcium (2000): 23
Authors: Wohlfart B., undefined
Journal: Acta Physiol Scand (2000): 1
Authors: Rockwell PL, Storey BT.
Journal: Mol Reprod Dev (1999): 418
Application notes
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A Novel NO Wash Probeniceid-Free Calcium Assay for Functional Analysis of GPCR and Calcium Channel Targets
Abbreviation of Common Chemical Compounds Related to Peptides
FAQ
Are there any alternatives to BrdU (Bromodeoxyuridine)?
Are there any alternatives to Cy5?
Are there any alternatives to indocyanine green (ICG)?
Are there any calcium indicators that don't require probenecid (PBC)?