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AB219932

Nitric Oxide Assay Kit (Fluorometric - Orange)

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(2 Publications)

Nitric Oxide Assay Kit (Fluorometric - Orange) (ab219932) is a sensitive fluorometric assay to monitor intracellular nitric oxide (NO) levels in live cells.
2 Images
Fluorescence Microscopy - Nitric Oxide Assay Kit (Fluorometric - Orange) (AB219932)
  • Fluorescence Microscopy

Supplier Data

Fluorescence Microscopy - Nitric Oxide Assay Kit (Fluorometric - Orange) (AB219932)

Endogenous nitric oxide (NO) production in RAW 264.7 macrophage cells. Raw 264.7 cells were seeded overnight (105 cells/well/100 μL) in a black wall/clear bottom 96-well plate. Left : cells were treated with 20 μg/mL of lipopolysaccharide (LPS) and 1 mM L-Arginine (L-Arg) and co-incubated with 0.5X NO stain working solution at at 37°C for 16 hours. Right : control RAW 264.7 cells were left untreated in cell culture medium and co-incubted with 0.5X NO stain working solution at 37°C for 16 hours. The solution in each well was removed and Assay Buffer II was added before fluorescence measurement. The fluorescence signal was measured using fluorescence microscope with a TRITC filter.

Functional Studies - Nitric Oxide Assay Kit (Fluorometric - Orange) (AB219932)
  • FuncS

Supplier Data

Functional Studies - Nitric Oxide Assay Kit (Fluorometric - Orange) (AB219932)

Exogenous nitric oxide (NO) production upon DEA/NONOate treatment (NO donor). CHO-K1 (blue) and HeLa (red) cells were incubated with NO stain working solution at 37°C for 30 minutes, and then removed to stop the staining. Cells were further treated with or without 1mM DEA/NONOate in HBSS buffer (with 10 mM HEPES (pH=6.2)) at 37°C for 30 minutes. The solution in each well was removed and Assay Buffer II was added before measuring fluorescence. The fluorescence signal was monitored at Ex/Em = 540/590 nm (cut off = 570 nm) with bottom read mode using a FlexStation microplate reader (Molecular Devices).

Key facts

Detection method

Fluorescent

Sample types

Suspension cells, Adherent cells

Assay type

Cell-based

Assay Platform

Microplate reader, Fluor. microscope, Flow cyt.

Product details

Nitric Oxide Assay Kit (Fluorometric - Orange) (ab219932) is a sensitive fluorometric assay to monitor intracellular nitric oxide (NO) levels in live cells. The assay uses an orange dye that can react with NO to generate a bright orange fluorescent product that can be easily detected at Ex/Em = 540/590 nm, using the same filter set as Cy3® or TRITC.

The assay can be detected by fluorescence microscopy, microplate fluorometry or high-content imaging. It can be easily adapted to use in 384-well microplate format.

Nitric oxide (NO) is an important biological regulator involved in numbers of physiological and pathological processes. Altered NO production is implicated in various immunological, cardiovascular, neurodegenerative and inflammatory diseases. As a free radical, NO is rapidly oxidized and there is relatively low concentrations of NO existing in vivo. It has been challenging to detect and understand the role of NO in biological systems.

What's included?

{ "values": { "200Test": { "sellingSize": "200 Test", "publicAssetCode":"ab219932-200Test", "assetComponentDetails": [ { "size":"1 x 20 mL", "name":"Assay Buffer I", "number":"AB219932-CMP02", "productcode":"" }, { "size":"1 x 50 µL", "name":"500X NO Orange Dye", "number":"AB219932-CMP01", "productcode":"" }, { "size":"1 x 20 mL", "name":"Assay Buffer II", "number":"AB219932-CMP03", "productcode":"" } ] } } }

Properties and storage information

Shipped at conditions
Blue Ice
Appropriate short-term storage conditions
-20°C
Appropriate long-term storage conditions
-20°C
Storage information
-20°C

Supplementary information

This supplementary information is collated from multiple sources and compiled automatically.

Nitric oxide (NO) is a small diatomic molecule with a mass of 30.01 Da. It plays a significant mechanical role as a signaling molecule in numerous systems. Nitric oxide is synthesized by nitric oxide synthases (NOS) and is expressed in endothelial cells neurons and various immune cells. Its production occurs in response to mechanical shear stress calcium influx or immunological stimulation. Nitric oxide exhibits high diffusibility enabling it to cross cellular membranes easily which is key for its function as a signaling messenger.
Biological function summary

Nitric oxide mediates vasodilation neurotransmission and immune response modulation. It is important for maintaining vascular homeostasis where it relaxes smooth muscle tissue promoting blood flow. In the central nervous system nitric oxide acts as a neurotransmitter and neuromodulator influencing synaptic plasticity. Within the immune system it functions to regulate leukocyte adhesion and can mediate microbial destruction. Nitric oxide often works within complexes associating with proteins like guanylate cyclase enhancing its signaling efficacy through nitrosylation.

Pathways

Nitric oxide integrates into key biological processes such as the nitric oxide signaling pathway and the endothelial nitric oxide pathway. It activates soluble guanylate cyclase increasing cyclic GMP levels leading to smooth muscle relaxation and vasodilation. In the endothelial pathway nitric oxide links with other proteins like endothelium-derived relaxing factor and interacts with oxidative stress pathways showing its role in cellular defense mechanisms. These pathways manage critical physiological responses and connect nitric oxide to other cardiovascular regulators.

Nitric oxide dysregulation associates with conditions like cardiovascular disease and neurodegenerative disorders. A deficiency or excess production can contribute to hypertension or atherosclerosis given its role in vascular health. Neurodegenerative diseases such as Alzheimer's disease also show links to nitric oxide largely due to its involvement in neuroinflammation and synaptic dysfunctions. It interacts with proteins like amyloid-beta precursor protein and tau in these contexts highlighting its importance in pathology and therapeutic targets.

Product protocols

Publications (2)

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Communications biology 5:379 PubMed35440683

2022

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Nature communications 13:646 PubMed35115501

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