Cell Meter™ Fluorimetric Intracellular Nitric Oxide (NO) Activity Assay Kit *Red Fluorescence Optimized for Flow Cytometry*
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Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
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International | See distributors |
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Custom size | Inquire |
Shipping | Standard overnight for United States, inquire for international |
Excitation (nm) | 586 |
Emission (nm) | 607 |
H-phrase | H303, H313, H333 |
Hazard symbol | XN |
Intended use | Research Use Only (RUO) |
R-phrase | R20, R21, R22 |
UNSPSC | 12352200 |
Overview | ![]() ![]() |
Excitation (nm) 586 | Emission (nm) 607 |
Platform
Flow cytometer
Excitation | 488 nm laser |
Emission | 610/20 nm filter |
Instrument specification(s) | Texas Red channel |
Components
Example protocol
AT A GLANCE
- Prepare cells (0.5 - 1×106 cells/mL)
- Add 1 µL 500X Nitrixyte™ Red
- Incubate cells with test compounds and Nitrixyte™ Red at 37 ºC for desired period of time
Analyze cells with a flow cytometer using 610/20 nm filter
Important
Thaw all the components at room temperature before starting the experiment.
CELL PREPARATION
For guidelines on cell sample preparation, please visit https://www.aatbio.com/resources/guides/cell-sample-preparation.html
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.
Add 200 uL of ddH2O into the vial of NONOate Positive Control (Component B) to make 50 mM NONOacte Positive Control treatment stock solution.
PREPARATION OF WORKING SOLUTION
Dilute 50 mM NONOate Positive Control treatment stock solution with Assay Buffer (Component C) to make 1-2 mM NONOate positive control working solution.
SAMPLE EXPERIMENTAL PROTOCOL
For each sample, prepare cells in 0.5 mL warm medium or buffer of your choice at a density of 5×105 to 1×106 cells/mL. Note: Each cell line should be evaluated on an individual basis to determine the optimal cell density for NO induction.
Add 1 µL of 500X Nitrixyte™ Red (Component A) into 0.5 mL cell suspension. Note: For adherent cells, gently lift the cells with 0.5 mM EDTA to keep the cells intact, and wash the cells once with serum-containing media prior to incubation with Nitrixyte™ Red.
Incubate cells with test compounds and Nitrixyte™ Red at 37 ºC for a desired period of time to generate endogenous or exogenous NO. Note: The appropriate incubation time depends on the individual cell type and test compound used. Optimize the incubation time for each experiment. Note: We have used Raw 264.7 cells incubated with Nitrixyte™ Red working solution, 20 µg/mL of lipopolysaccharide (LPS) and 1 mM L-Arginine (L-Arg) in cell culture medium at 37 ºC for 16 hours.
- Spin down cells that have pre-incubated with Nitrixyte™ Red for 30 minutes. Resuspend cells with 1 mM DEA NONOate positive control working solution, and incubate at 37 ºC for another 30 minutes. See Figure 1 for details.
Monitor the fluorescence intensity using the 610/20 nm filter in a flow cytometer. Gate on the cells of interest, excluding debris.
Product Family
Name | Excitation (nm) | Emission (nm) |
Cell Meter™ Fluorimetric Intracellular Nitric Oxide (NO) Activity Assay Kit *Orange Fluorescence Optimized for Flow Cytometry* | 552 | 575 |
Images
![Detection of exogenous nitric oxide (NO) in Jurkat cells upon DEA NONOate treatment (NO donor) using Cell Meter™ Fluorimetric Intracellular Nitric Oxide Assay Kit (Cat#16356). Cells were incubated with Nitrixyte™ Red at 37 °C for 30 minutes. Cells were further treated with (Red line) or without (Blue line) 1 mM DEA NONOate at 37 °C for another 30 minutes. The fluorescence signal was monitored at FL4 channel using a flow cytometer (BD FACSCalibur).](/_next/image?url=https%3A%2F%2Fimages.aatbio.com%2Fproducts%2Ffigures-and-data%2Fcell-meter-fluorimetric-intracellular-nitric-oxide-no-activity-assay-kit-red-fluorescence-optimized-for-flow-cytometry%2Ffigure-for-cell-meter-fluorimetric-intracellular-nitric-oxide-no-activity-assay-kit-red-fluorescence-optimized-for-flow-cytometry_pKaVe.png&w=3840&q=75)
Citations
Authors: Mostek-Majewska, Agnieszka and Majewska, Anna and Janta, Anna and Ciereszko, Andrzej
Journal: Cell Communication and Signaling (2023): 1--23
Authors: Schweikl, Helmut and Birke, Margaritha and Gallorini, Marialucia and Petzel, Christine and Bolay, Carola and Waha, Claudia and Hiller, Karl-Anton and Buchalla, Wolfgang
Journal: Dental Materials (2020)
Authors: Ghimire, Kedar and Zaric, Jelena and Alday-Parejo, Begona and Seebach, Jochen and Bousquenaud, Mélanie and Stalin, Jimmy and Bieler, Grégory and Schnittler, Hans-Joachim and Rüegg, Curzio
Journal: Cells (2019): 388
Authors: Schweikl, Helmut and Gallorini, Marialucia and P{\"o}schl, Gerd and Urmann, Vera and Petzel, Christine and Bolay, Carola and Hiller, Karl-Anton and Cataldi, Amelia and Buchalla, Wolfgang
Journal: Dental Materials (2018): 1661--1678
Authors: Luo, Zhen and Zhao, Qin and Liu, Jixiang and Liao, Jinfang and Peng, Ruogu and Xi, Yunting and Diwu, Zhenjun
Journal: Analytical biochemistry (2017): 44--48
Authors: Adamiak, Mateusz and Abdelbaset-Ismail, Ahmed and Moore, Joseph B and Zhao, J and Abdel-Latif, Ahmed and Wysoczynski, Marcin and Ratajczak, Mariusz Z
Journal: Stem Cell Reviews and Reports (2016): 1--12
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Application notes
Abbreviation of Common Chemical Compounds Related to Peptides
Attenuation of lysyl oxidase and collagen gene expression in keratoconus patient corneal epithelium corresponds to disease severity
Copper Transport Protein Antioxidant-1 Promotes Inflammatory Neovascularization via Chaperone and Transcription Factor Function
Dopamine-Mediated Oxidation of Methionine 127 in α-Synuclein