Cal-520®, AM
Price | |
Catalog Number | |
Availability | In stock |
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) | 320 |
Molecular weight | 1102.95 |
Solvent | DMSO |
Excitation (nm) | 492 |
Emission (nm) | 515 |
Quantum yield | 0.751 |
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 1102.95 | Dissociation constant (Kd, nM) 320 | Excitation (nm) 492 | Emission (nm) 515 | Quantum yield 0.751 |
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 Cal-520® AM in high-quality, anhydrous DMSO.
Note: When reconstituted in DMSO, Cal-520® AM is a clear, colorless solution.
PREPARATION OF WORKING SOLUTION
On the day of the experiment, either dissolve Cal-520® AM in DMSO or thaw an aliquot of the indicator stock solution to room temperature.
Prepare a 2 to 20 µM Cal-520® 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-520® 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-520® 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-520® 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 1 to 2 hours.
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 Ex/Em = 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 | 90.666 µL | 453.33 µL | 906.659 µL | 4.533 mL | 9.067 mL |
5 mM | 18.133 µL | 90.666 µL | 181.332 µL | 906.659 µL | 1.813 mL |
10 mM | 9.067 µL | 45.333 µL | 90.666 µL | 453.33 µL | 906.659 µL |
Molarity calculator
Mass (Calculate) | Molecular weight | Volume (Calculate) | Concentration (Calculate) | Moles | ||||
/ | = | x | = |
Spectrum
Spectral properties
Excitation (nm) | 492 |
Emission (nm) | 515 |
Quantum yield | 0.751 |
Product Family
Name | Excitation (nm) | Emission (nm) | Quantum yield |
Cal-520® maleimide | 492 | 515 | 0.751 |
Cal-520FF™, AM | 492 | 515 | 0.751 |
Cal-520N™, AM | 492 | 515 | 0.751 |
Cal-520® amine | 492 | 515 | 0.751 |
Cal-520® azide | 492 | 515 | 0.751 |
Cal-520® alkyne | 492 | 515 | 0.751 |
Cal-590™ AM | 574 | 588 | 0.621 |
Cal-630™ AM | 609 | 626 | 0.371 |
Calbryte™ 520 AM | 493 | 515 | 0.751 |
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Images
Citations
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Journal: Cell Reports (2024)
Authors: Liu, Yang and Chen, Xinyu and Ma, Yunlong and Song, Chenyun and Ma, Jixin and Chen, Cheng and Su, Jianzhong and Ma, Lixiang and Saiyin, Hexige
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Authors: Buckley, Charlotte and Lee, Matthew D and Zhang, Xun and Wilson, Calum and McCarron, John G
Journal: British Journal of Pharmacology (2024)
Authors: Chen, Zhong and Lu, Xiaowen and Watsky, Mitchell A
Journal: Plos one (2024): e0301495
Authors: Wu, Yukun and Xu, Zhehao and Liang, Shanshan and Wang, Lukang and Wang, Meng and Jia, Hongbo and Chen, Xiaowei and Zhao, Zhikai and Liao, Xiang
Journal: Biomedical Optics Express (2024): 2910--2925
Authors: Kazaoka, Akira and Fujimori, Sota and Yamada, Yushiro and Shirayanagi, Tomohiro and Gao, Yuying and Kuwahara, Saki and Sakamoto, Naoki and Susukida, Takeshi and Aoki, Shigeki and Ito, Kousei
Journal: PNAS Nexus (2024): pgae140
Authors: Daida, Atsuro and Kurotani, Tohru and Yamaguchi, Kazuhiko and Takahashi, Yuji and Ichinohe, Noritaka
Journal: (2024)
Authors: Yang, Mingyu
Journal: (2024)
Authors: Kudo, Wataru and Hashitani, Hikaru
Journal: Pfl{\"u}gers Archiv-European Journal of Physiology (2024): 1--12
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