Cal-590™ 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) | 561 |
Molecular weight | 1266.81 |
Solvent | DMSO |
Extinction coefficient (cm -1 M -1) | 78000 |
Excitation (nm) | 574 |
Emission (nm) | 588 |
Quantum yield | 0.621 |
Certificate of Origin | Download PDF |
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 1266.81 | Dissociation constant (Kd, nM) 561 | Extinction coefficient (cm -1 M -1) 78000 | Excitation (nm) 574 | Emission (nm) 588 | Quantum yield 0.621 |
Platform
Fluorescence microscope
Excitation | TRITC/Cy3 |
Emission | TRITC/Cy3 |
Recommended plate | Black wall/clear bottom |
Fluorescence microplate reader
Excitation | 540 |
Emission | 590 |
Cutoff | 570 |
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-590™ AM in anhydrous DMSO.
Note: When reconstituted in DMSO, Cal-590™ AM is a clear, colorless solution.
PREPARATION OF WORKING SOLUTION
On the day of the experiment, either dissolve Cal-590™ AM in DMSO or thaw an aliquot of the indicator stock solution to room temperature.
Prepare a 2 to 20 µM Cal-590™ 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-590™ 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-590™ 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-590™ 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 TRITC/Cy3 filter set or a fluorescence plate reader containing a programmable liquid handling system such as an FDSS, FLIPR, or FlexStation, at Ex/Em = 540/590 nm cutoff 570 nm.
Calculators
Common stock solution preparation
0.1 mg | 0.5 mg | 1 mg | 5 mg | 10 mg | |
1 mM | 78.938 µL | 394.692 µL | 789.384 µL | 3.947 mL | 7.894 mL |
5 mM | 15.788 µL | 78.938 µL | 157.877 µL | 789.384 µL | 1.579 mL |
10 mM | 7.894 µL | 39.469 µL | 78.938 µL | 394.692 µL | 789.384 µL |
Molarity calculator
Mass (Calculate) | Molecular weight | Volume (Calculate) | Concentration (Calculate) | Moles | ||||
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Spectrum
Spectral properties
Extinction coefficient (cm -1 M -1) | 78000 |
Excitation (nm) | 574 |
Emission (nm) | 588 |
Quantum yield | 0.621 |
Product Family
Name | Excitation (nm) | Emission (nm) | Quantum yield |
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 |
Calbryte™ 590 AM | 581 | 593 | - |
Cal-500™ AM | 388 | 482 | 0.481 |
Images
Citations
Authors: Oropeza-Almaz{\'a}n, Yuriana and Blatter, Lothar A
Journal: Biomolecules (2024): 144
Authors: Shcherban, Igor V and Fedotova, Victoria S and Matukhno, Aleksey E and Shepelev, Igor E and Shcherban, Oxana G and Lysenko, Larisa V
Journal: Journal of Neuroscience Methods (2024): 110097
Authors: Ataka, Mitsutoshi and Otomo, Kohei and Enoki, Ryosuke and Ishii, Hirokazu and Tsutsumi, Motosuke and Kozawa, Yuichi and Sato, Shunichi and Nemoto, Tomomi
Journal: Biomedical Optics Express (2024): 1089--1101
Authors: Granzotto, Alberto and McQuade, Amanda and Chadarevian, Jean Paul and Davtyan, Hayk and Sensi, Stefano L and Parker, Ian and Blurton-Jones, Mathew and Smith, Ian
Journal: bioRxiv (2024): 2024--01
Authors: Matukhno, Aleksey E and Petrushan, Mikhail V and Kiroy, Valery N and Arsenyev, Fedor V and Lysenko, Larisa V
Journal: Journal of Computational Neuroscience (2023): 1--12
Authors: Wang, Anna YM and Kulkarni, Manoj M and McLaughlin, Amanda J and Gayet, Jacqueline and Smith, Benjamin E and Hauptschein, Max and McHugh, Cyrus F and Yao, Yvette Y and Puthussery, Teresa
Journal: Nature (2023): 1--6
Authors: Khaddaj-Mallata, Rayan and Aldiba, Natija and Bernarda, Maxime and Paquettea, Anne-Sophie and Ferreirab, Aymeric and Lecordiera, Sarah and Saghatelyanb, Armen and Flamanda, Louis and ElAlia, Ayman
Journal: Analyse d'{\'e}v{\`e}nements neurobiologiques h{\'e}t{\'e}rog{\`e}nes {\`a} l'aide d'outils computationnels. (2023): 200
Authors: Yu, Vanessa and Yong, Fiona and Marta, Angellica and Khadayate, Sanjay and Osakwe, Adrien and Bhattacharya, Supriyo and Varghese, Sneha S and Chabosseau, Pauline and Tabibi, Sayed M and Chen, Keran and others,
Journal: bioRxiv (2023)
Authors: Qian, Tongrui and Wang, Huan and Wang, Peng and Geng, Lan and Mei, Long and Osakada, Takuya and Wang, Lei and Tang, Yan and Kania, Alan and Grinevich, Valery and others,
Journal: Nature Biotechnology (2023): 1--14
Authors: Jorfi, Mehdi and Park, Joseph and Hall, Clare K and Lin, Chih-Chung Jerry and Chen, Meng and von Maydell, Djuna and Kruskop, Jane M and Kang, Byunghoon and Choi, Younjung and Prokopenko, Dmitry and others,
Journal: Nature Neuroscience (2023): 1--16
Application notes
A Meta-Analysis of Common Calcium Indicators
Novel improved Ca2+ indicator dyes on the market-a comparative study of novel Ca2+ indicators with fluo-4
Cal-520 ® , Cal-590 ™, and Cal-630™ Calcium Detection Reagents
Calibration Protocol for Fluorescent Calcium Indicators