Cardiovascular and Oxidative Stress Assay Kit

NOS Colorimetric Assay Kit

$405.00

The NOS Colorimetric Assay Kit is for the quantitative determination of Nitric Oxide Synthase in biological fluids by a microplate assay.

SKU: NSC31-K01 Categories: , ,

Nitric Oxide Synthase Colorimetric Assay Kit

The NOS Colorimetric Assay Kit is For Research Use Only

Size: 1 x 96 wells
Sensitivity: 1 pmol/µL or ~0.5 µM
Dynamic Range: 0.5 – 100 µM
Incubation Time: 5 minutes
Sample Type: Biological Fluids
Sample Size: 5 µl
Alternative Name: Nitric Oxide Synthase

Product Developed and Manufactured in the USA

Assay Background

The Nitric Oxide Synthase Colorimetric Assay kit employs immunoaffinity purified Nitrate Reductase (NaR) enabling the measure of total nitric oxide (NO) produced in in vitro experimental systems.  Nitric oxide can be spectrophotometrically assayed by measuring the accumulation of its stable degradation products, nitrate and nitrite.  The ratio of these two products in biological fluids, tissue culture media, etc. may vary substantially.  Hence, for accurate assessment of the total nitric oxide generated, one must monitor both nitrate and nitrite.  An excellent solution to this problem is the enzymatic conversion of nitrate to nitrite by the enzyme nitrate reductase, followed by quantitation of nitrite using Griess Reagent.  In addition to providing all necessary components in a microtiter format, this kit employs affinity purified nitrate reductase and NADH, thereby circumventing some of the potential problems reported for NO measurement using NADPH dependent nitrate reductases.

Related Products

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Additional Information

Assay Principle

In aqueous solution, nitric oxide rapidly degrades to nitrate and nitrite. Spectrophotometric quantitation of nitrite using Griess Reagent is straightforward, but does not measure nitrate. This Nitric Oxide Synthase Colorimetric Assay kit employs the NADH-dependent enzyme nitrate reductase for conversion of nitrate to nitrite prior to the quantitation of nitrite using Griess reagent — thus providing for accurate determination of total NO production.

This Nitric Oxide Synthase Colorimetric Assay kit can be used to accurately measure as little as 1 pmol/mL (~1mM) NO produced in aqueous solutions.  Very little sample is required (5 to 85 mL depending on the [NO] in the sample. The completed reaction is read at 540 nm.

Assay Procedure

  1. Add 85 µL of Standards and 5-85 µL of Samples (depending on [NO]) to the corresponding wells on the microplate in duplicate.
  2. Add sufficient Buffer to each sample well to bring the final volume to 85 µL. (i.e. 80 µL Buffer for 5 µL of Sample.)
  3. Add 10 µL of the reconstituted Nitrate Reductase to each well.
  4. Add 10 µL of NADH working solution to each well and shake the plate for 20 minutes at RT.
  5. Add 50 µL of Color Reagent #1 to each well and shake briefly
  6. Add 50 µL of Color Reagent #2 to each well and shake for 5 minutes at RT.
  7. Read the plate at 540 nm in the plate reader.

Typical Standard Curve

Package Inserts

Package Inserts

Please note: All documents above are for reference use only and should not be used in place of the documents included with this physical product. If digital copies are needed, please contact us.

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