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Cal Green™ 1, AM [Equivalent to Calcium Green-1, AM]

Chemical structure for Cal Green™ 1, AM [Equivalent to Calcium Green-1, AM]
Chemical structure for Cal Green™ 1, AM [Equivalent to Calcium Green-1, AM]
Ordering information
Price ()
Catalog Number20501
Unit Size
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Additional ordering information
Telephone1-408-733-1055
Fax1-408-733-1304
Emailsales@aatbio.com
InternationalSee distributors
ShippingStandard overnight for United States, inquire for international
Physical properties
Dissociation constant (Kd, nM)190
Molecular weight1290.96
SolventDMSO
Spectral properties
Excitation (nm)498
Emission (nm)517
Quantum yield0.751
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
StorageFreeze (< -15 °C); Minimize light exposure
UNSPSC12352200
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Show More (87)

OverviewpdfSDSpdfProtocol


CAS
186501-28-0
Molecular weight
1290.96
Dissociation constant (Kd, nM)
190
Excitation (nm)
498
Emission (nm)
517
Quantum yield
0.751
Cal Green™ 1 (AAT Bioquest) is the same molecule to Calcium Green-1 (Invitrogen). It exhibits an increase in fluorescence intensity upon binding calcium ion. The cell-permeant dye, Cal Green 1 AM (Calcium Green-1 AM) is a 488 nm-excitable calcium indicator. Compared to Fluo-3 AM, Cal Green-1 AM is more fluorescent at low calcium concentrations in cells, facilitating the determination of baseline calcium levels and increasing the visibility of resting cells. It has been used in many calcium signaling investigations, including measuring intracellular calcium, following calcium influx and release, and multiphoton excitation imaging of calcium in living tissues. Cells can be loaded with Cal Green-1 AM by adding the dissolved indicator directly to cultured cells in medium. The fluorescence signal from these cells is generally measured using fluorescence microscopy, fluorescence microplate assays, or flow cytometry.

Platform


Fluorescence microscope

ExcitationFITC
EmissionFITC
Recommended plateBlack wall/clear bottom

Fluorescence microplate reader

Excitation490
Emission525
Cutoff515
Recommended plateBlack 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.

Cal Green™ 1 AM Stock Solution
Prepare a 2 to 5 mM stock solution of Cal Green™ 1 AM in high-quality, anhydrous DMSO.

PREPARATION OF WORKING SOLUTION

Cal Green™ 1 AM Working Solution
On the day of the experiment, either dissolve Cal Green™ 1 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, Cal Green™ 1 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 Green™ 1 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 solution, 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.
  1. Prepare cells in growth medium overnight.
  2. On the next day, add 1X Cal Green™ 1 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.
  3. Incubate the dye-loaded plate in a cell incubator at 37 °C for 30 to 60 minutes.
    Note     Incubating the dye for longer than 1 hour can improve signal intensities in certain cell lines.
  4. 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.
  5. 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

Table 1. Volume of DMSO needed to reconstitute specific mass of Cal Green™ 1, AM [Equivalent to Calcium Green-1, AM] to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.

0.1 mg0.5 mg1 mg5 mg10 mg
1 mM77.462 µL387.309 µL774.617 µL3.873 mL7.746 mL
5 mM15.492 µL77.462 µL154.923 µL774.617 µL1.549 mL
10 mM7.746 µL38.731 µL77.462 µL387.309 µL774.617 µL

Molarity calculator

Enter any two values (mass, volume, concentration) to calculate the third.

Mass (Calculate)Molecular weightVolume (Calculate)Concentration (Calculate)Moles
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Spectrum


Open in Advanced Spectrum Viewer
spectrum

Spectral properties

Excitation (nm)498
Emission (nm)517
Quantum yield0.751

Citations


View all 4 citations: Citation Explorer
Network pharmacology-based research uncovers cold resistance and thermogenesis mechanism of Cinnamomum cassia
Authors: Jiang, Xiao-wen and Lu, Hong-yuan and Xu, Zi-Hua and Zhang, Ying-Shi and Zhao, Qing-Chun and others,
Journal: Fitoterapia (2021): 104824
Binding to carboxypeptidase M mediates protective effects of fibrinopeptide Bβ (15-42)
Authors: Sörensen-Zender, Inga and Chen, Rongjun and Song, Rong and David, Sascha and Melk, Anette and Haller, Hermann and Schmitt, Rol and , undefined
Journal: Translational Research (2019)
Tuning the Color Palette of Fluorescent Copper Sensors through Systematic Heteroatom Substitution at Rhodol Cores
Authors: Jia, Shang and Ramos-Torres, Karla M and Kolemen, Safacan and Ackerman, Cheri M and Chang, Christopher J
Journal: (2017)
Tuning the color palette of fluorescent copper sensors through systematic heteroatom substitution at rhodol cores
Authors: Jia, Shang and Ramos-Torres, Karla M and Kolemen, Safacan and Ackerman, Cheri M and Chang, Christopher J
Journal: ACS chemical biology (2017): 1844--1852

References


View all 11 references: Citation Explorer
Microscopic imaging of intracellular calcium in live cells using lifetime-based ratiometric measurements of Oregon Green BAPTA-1
Authors: Lattarulo C, Thyssen D, Kuchibholta KV, Hyman BT, Bacskaiq BJ.
Journal: Methods Mol Biol (2011): 377
Calcium Green FlAsH as a genetically targeted small-molecule calcium indicator
Authors: Tour O, Adams SR, Kerr RA, Meijer RM, Sejnowski TJ, Tsien RW, Tsien RY.
Journal: Nat Chem Biol (2007): 423
An ensemble and single-molecule fluorescence spectroscopy investigation of Calcium Green 1, a calcium-ion sensor
Authors: Lu Y, Paige MF.
Journal: J Fluoresc (2007): 739
Measurement of intracellular calcium levels by the fluorescent Ca2+ indicator Calcium-Green
Authors: Silei V, Fabrizi C, Venturini G, Tagliavini F, Salmona M, Bugiani O, Lauro GM.
Journal: Brain Res Brain Res Protoc (2000): 132
Monitoring of Ca2+ release from intracellular stores in permeabilized rat parotid acinar cells using the fluorescent indicators Mag-fura-2 and calcium green C18
Authors: Tojyo Y, Tanimura A, Matsumoto Y.
Journal: Biochem Biophys Res Commun (1997): 189
Optical imaging of neuronal activity in tissue labeled by retrograde transport of Calcium Green Dextran
Authors: McPherson DR, McClellan AD, O'Donovan MJ.
Journal: Brain Res Brain Res Protoc (1997): 157
Modulation of an Intracellular Calmodulin-Stimulated Ca2+-Pumping ATPase in Cauliflower by Trypsin (The Use of Calcium Green-5N to Measure Ca2+ Transport in Membrane Vesicles)
Authors: Askerlund P., undefined
Journal: Plant Physiol (1996): 913
A method for recording intracellular [Ca2+] transients in cardiac myocytes using calcium green-2
Authors: Spencer CI, Berlin JR.
Journal: Pflugers Arch (1995): 579
Characterization of calcium translocation across the plasma membrane of primary osteoblasts using a lipophilic calcium-sensitive fluorescent dye, calcium green C18
Authors: Lloyd QP, Kuhn MA, Gay CV.
Journal: J Biol Chem (1995): 22445
Calcium green-5N, a novel fluorescent probe for monitoring high intracellular free Ca2+ concentrations associated with glutamate excitotoxicity in cultured rat brain neurons
Authors: Rajdev S, Reynolds IJ.
Journal: Neurosci Lett (1993): 149