Screen Quest™ Fluo-8 No Wash Calcium Assay Kit
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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 |
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 |
UNSPSC | 12352200 |
Overview | SDSProtocol |
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 microplate reader
Excitation | 490 nm |
Emission | 525 nm |
Cutoff | 510 nm |
Recommended plate | Black wall/Clear bottom |
Instrument specification(s) | Bottom read mode/Programmable liquid handling |
Other instruments
ArrayScan, FDSS, FLIPR, FlexStation, IN Cell Analyzer, NOVOStar, ViewLuxComponents
Example protocol
AT A GLANCE
Protocol summary
- Prepare cells in growth medium with 1-5% FBS
- Add Fluo-8 NW dye-loading solution (100 µL/well for 96-well plate or 25 µL/well for 384-well plate)
- Incubate at room temperature for 1 hour
- Monitor fluorescence intensity at Ex/Em = 490/525 nm
Important notes
Thaw all the kit components at room temperature before starting the experiment.
PREPARATION OF STOCK SOLUTION
1. Fluo-8 NW stock solution:
Add 20 µL (for Cat. # 36314) or 200 µL (for Cat. # 36315 and # 36316) of DMSO into the vial of Fluo-8 NW (Component A), and mix them well. Note: 20 µL of Fluo-8 NW stock solution is enough for one plate. Un-used Fluo-8 NW stock solution can be aliquoted and stored at < -20 oC for more than one month if the tubes are sealed tightly. Protect from light and avoid repeated freeze-thaw cycles.
2. Assay Buffer (1X):
a) For Cat. # 36314 (1 plate kit) and # 36315 (10 plates kit), make 1X assay buffer by adding 9 mL of HHBS (Component C) into 10X Pluronic® F127 Plus (1 mL, Component B), and mix them well.
b) For Cat. # 36316 (100 plates kit), make 1X assay buffer by adding the whole bottle of 10 X Pluronic® F127 Plus, (10 mL, Component B) into 90 mL of HHBS buffer (not included in the kit), and mix them well. Note: 10 mL of 1X assay buffer is enough for one plate. Aliquot and store un-used 1X assay buffer at < -20 oC. Protect from light and avoid repeated freeze-thaw cycles
PREPARATION OF WORKING SOLUTION
Fluo-8 NW dye-loading solution:
Add 20 µL of Fluo-8 NW stock solution into 10 mL of 1X assay buffer, and mix them well. Note: This working solution is stable for at least 2 hours at room temperature.
For guidelines on cell sample preparation, please visit
https://www.aatbio.com/resources/guides/cell-sample-preparation.html
SAMPLE EXPERIMENTAL PROTOCOL
- Add 100 µL/well (96-well plate) or 25 µL/well (384-well plate) of Fluo-8 NW dye-loading solution into the cell plate. [We offer 2 separate medium removal calcium assay kits (Cat.# 36308 and 36309) for those who prefer to keep the medium removal step].
- Incubate the dye-loading plate in a cell incubator for 30 minutes, and then incubate the plate at room temperature for another 30 minutes. Note: If the assay requires 37 oC, perform the experiment immediately without further room temperature incubation. Note: If the cells can function well at room temperature for longer time, incubate the cell plate at room temperature for 1-2 hours (It is recommended that the incubation time be no longer than 2 hours).
- Prepare the compound plate with HHBS or your desired buffer.
- Run the calcium flux assay by monitoring the fluorescence intensity at Ex/Em = 490/525 nm. Note: It is important to run the signal test before the experiment. Different instruments have their own intensity range. Adjust the signal test intensity to the level of 10% to 15% of the maximum instrument intensity counts. For example, the maximum fluorescence intensity count for FLIPR-384 is 65,000, so the instrument settings should be adjusted to have the signal test intensity around 7,000 to 10,000.
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 |
Screen Quest™ Fluo-4 No Wash Calcium Assay Kit | 495 | 528 | 82000 | 0.161 |
Images
(A) Optimal view of the L-CTS placed in a 10cm-sized UpCell dish. Scale bar = 1 mm. (B) (C) Calcium transient of L-CTSs. (B) Representative Fluo-8 image of L-CTS (see also S1 video) and region of interests (ROIs). Magnification ×100. (C) Chronologic intensity change of Fluo-8. Note that the peak timings of 4 different ROIs are almost same chronologically. Source: Human iPS cell-derived cardiac tissue sheets for functional restoration of infarcted porcine hearts by Masanosuke Ishigami et al., PLOS, Aug 2018.
Citations
Authors: Hsu, Julia Chu-Ning and Tseng, Hsu-Wen and Chen, Chia-Hui and Lee, Tzong-Shyuan
Journal: Experimental Animals (2024): 23--0148
Authors: Tian, Qingqing and Xing, Kunming and Liu, Yongshu and Wang, Qian and Sun, Haonan and Sun, Ying-Nan and Zhang, Shusheng
Journal: STAR Protocols (2023): 102115
Authors: Martino, Pieter and Sunkara, Raghava and Heitman, Nicholas and Rangl, Martina and Brown, Alexia and Saxena, Nivedita and Grisanti, Laura and Kohan, Donald and Yanagisawa, Masashi and Rendl, Michael
Journal: Nature Cell Biology (2023): 1--13
Authors: Froemke, Robert and Ahmed, Ismail and Liu, Jingjing and Gieniec, Krystyna and Bair-Marshall, Chloe and Adewakun, Ayomiposi and Hetzler, Belinda and Arp, Christopher and Khatri, Latika and Vanwalleghem, Gilles and others,
Journal: (2023)
Authors: Zhang, Nan and Sui, Yixuan and Jendrichovsky, Peter and Feng, Hemin and Shi, Heng and Zhang, Xu and Xu, Shenghui and Sun, Wenjian and Zhang, Huatang and Chen, Xi and others,
Journal: (2023)
Authors: Korkus, Eliza and Szustak, Marcin and Madaj, Rafal and Chworos, Arkadiusz and Drzazga, Anna and Kozio{\l}kiewicz, Maria and D{\k{a}}browski, Grzegorz and Czaplicki, Sylwester and Konopka, Iwona and Gendaszewska-Darmach, Edyta
Journal: Food \& Function (2023)
Authors: Pinheiro, In{\^e}s and Calo, Nicolas and Paolini-Bertrand, Marianne and Hartley, Oliver
Journal: (2023)
Authors: Korkus, Eliza and Szustak, Marcin and D{\k{a}}browski, Grzegorz and Czaplicki, Sylwester and Kad{\l}ubowski, S{\l}awomir and Kozio{\l}kiewicz, Maria and Konopka, Iwona and Gendaszewska-Darmach, Edyta
Journal: NFS Journal (2023)
Authors: Gao, Rong and Wang, Heting and Li, Ting and Wang, Jin and Ren, Zhitao and Cai, Nan and Ai, Heying and Li, Shasha and Lu, Yan and Zhu, Yanhua and others,
Journal: Pharmacological Research (2022): 106585
Authors: Yart, Lucile and Frieden, Maud and Konig, St{\'e}phane and Cohen, Marie and Martinez de Tejada, Bego{\~n}a
Journal: Journal of cellular physiology (2022)
Application notes
Evaluation of FLIPR Calcium Assays for Screening GPCR and Calcium Channel Targets
A Novel NO Wash Probeniceid-Free Calcium Assay for Functional Analysis of GPCR and Calcium Channel Targets
Fluorescent Dye AM Esters
Novel Red Fluorescent Calcium Probes for Functional Analysis of GPCRs and Calcium Channel Targets
FAQ
How do I make an AM ester stock solution?
How do you measure Ca2+ response with calcium indicators like Calbryte 520, potassium salt?
Can I intracellularly measure mitochondria calcium flux and changes in mitochondria membrane potential at the same time?
Do you offer any products for measuring intracellular calcium concentration or movement by flow cytometry?