NOS

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Description

Key Benefits

  • Cell permeable.
  • No wash homogenous assay.
  • Adaptable to High throughput assay platforms .
  • Real time detection of NO activity.
  • Applications – Flow Cytometry, Fluorescence Plate Reader, Fluorescence Microscopy

Additional information

Kit Size

0.125mg, 0.25mg

ASSAY PRINCIPLE

Diaminofluorescein –2 Diacetate (DAF-2DA) is a non-fluorescent cell permeable reagent that can measure free Nitric Oxide (NO) and nitric oxide synthase (NOS) activity in living cells under physiological conditions. Once inside the cell the diacetate groups on the DFA-2DA reagent arehydrolyzed by cytosolic esterases thus releasing FAD-2 and sequestering the reagent inside the cell. Production of nitric oxide converts the non-fluorescent dye, DAF-2, to its fluorescent triaole derivative, DAF-2T.

DAF-2T can be observed by: excitation 488nm and measuring emission at 515nm.

nosdet1-f1

Fig. 1. DAF-2DA, the non-fluorescent probe, enters the cells where cytosolic esterases hydrolysis the diacetate groups, releasing the non-fluorescent DAF-2 dye. DAF-2 then reacts with NO and O2 to produce DAF-2T, its triazole fluorescent derivative.

nosdet2-f2

Fig. 2. Phase Contrast and fluorescent microscopic images of rat aorta-derived vascular smooth muscular cells loaded with DAF-2DA.

Readable Documents

Title Name
Protocol NOS Protocol.pdf
Datasheet NOS Datasheet.pdf
msds msds NOS.pdf

Kit contents and storage

Part# Reagent Temperature
Part# 4006 DAF-2DA, 1 Vial (NOS200-1) 2-8°C
Part# 4006 DAF-2DA, 2 Vials (NOS200-2) 2-8°C

 

References

Direct evidence of NO production in rat hippocampus and cortex using a new fluorescent indicator: DAF-2 DA: H. Kojima, et al.; Neuroreport 9, 3345 (1998) Abstract
Direct evidence of nitric oxide production from bovine aortic endothelial cells using new fluorescence indicators: diaminofluoresceins: N. Nakatsubo, etal.; FEBS Lett. 427, 263 (1998) Abstract
Nitric Oxide Ameliorates Hydrophobic Bile Acid-induced Apoptosis in Isolated Rat Hepatocytes by Non-mitochondrial Pathways: E. Gumpricht, et al.; J. Biol.Chem. 277, 25823 (2002) Full Text; http://www.jbc.org/cgi/content/full/277/28/25823#SEC1
Development of a fluorescent indicator for nitric oxide based on the fluoroscein chromophore: H. Kojima, et al.; Chem. Pharm. Bull. 46, 373 (1998)Abstract
Detection and imaging of nitric oxide with novel fluorescent indicators: diaminofluoresceins: H. Kojima, et al.; Anal. Chem. 70, 2446 (1998) Abstract

 

Citations

Title File Link Author(s) Journal Year; Edition:Pages
Red wine consumption improves in vitro migration of endothelial progenitor cells in young, healthy individuals Am J Clin Nutr 2010 92: 1 July 2010; 161-169

 

Product Specifications

  • Formula: C24H18N2O7
  • MW: 446.4
  • Appearance: Liquid
  • Real-time detection of NOS activity.
  • Manufactured: by Daiichi Pure Chemicals Co. Ltd. Japan