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Nitric Oxide Assay Kit (Colorimetric) (ab65328)

Reviews (2)Q&A (27)References (99)
Functional Studies - Nitric Oxide Assay Kit (Colorimetric) (ab65328)
  • Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)
  • Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)
  • Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)
  • Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)

Key features and details

  • Assay type: Quantitative
  • Detection method: Colorimetric
  • Platform: Microplate reader
  • Assay time: 1 hr 30 min
  • Sample type: Cell Lysate, Other biological fluids, Plasma, Serum, Tissue Extracts, Urine
  • Sensitivity: 1 nmol/well

Overview

  • Product name

    Nitric Oxide Assay Kit (Colorimetric)
    See all Nitric oxide kits

  • Detection method

    Colorimetric

  • Sample type

    Urine, Serum, Plasma, Other biological fluids, Tissue Extracts, Cell Lysate

  • Assay type

    Quantitative

  • Sensitivity

    1 nmol/well

  • Range

    1 nmol/well - 10 nmol/well

  • Assay time

    1h 30m

  • Species reactivity

    Reacts with: Mammals, Other species

  • Product overview

    Nitric Oxide Assay Kit (Colorimetric) ab65328 provides an accurate, convenient measure of total nitrate/nitrite in a simple two-step process.


    In the nitric oxide assay protocol:
    - the first step converts nitrate to nitrite by using nitrate reductase
    - the second step uses Griess Reagent to convert nitrite to a deep purple azo compound.
    The amount of the azochromophore accurately reflects nitric oxide amount in samples.


    Nitric oxide assay protocol summary:
    - add samples and standards to wells
    - add nitrate reductase and enzyme cofactor and incubate for 1 hr at room temp to convert nitrate to nitrite
    - add enhancer and incubate for 10 min at room temp
    - add Griess Reagent R1 and Griess Reagent R2
    - analyze with microplate reader

  • Notes

    This product is manufactured by BioVision, an Abcam company and was previously called K262 Nitric Oxide Colorimetric Assay Kit. K262-200 is the same size as the 2 96 tests size of ab65328.

    If you are interested in a fluorometric detection kit, please check Nitric Oxide Assay Kit (Fluorometric) (ab65327).

    For an assay kit to detect nitrite alone see Griess Reagent Kit ab234044.

    Nitric oxide (NO) plays an important role in neurotransmission, vascular regulation, immune response and apoptosis. NO is rapidly oxidized to nitrite and nitrate which are used to quantitate NO production.

  • Platform

    Microplate reader

Properties

  • Storage instructions

    Store at -20°C. Please refer to protocols.
  • Components 2 x 96 tests
    Assay Buffer XVII 1 x 30ml
    Enhancer II 1 vial
    Enzyme Cofactor 1 vial
    Griess Reagent I   1 x 10ml
    Griess Reagent II 1 x 10ml
    96-Well Clear Plate 2 units
    Nitrate Reductase I 1 vial
    Nitrate Standard 1 vial
    Nitrite Standard 1 vial
    TEMPPLATE® SEALING FILM 2 units

  • Research areas

  • Relevance

    Nitric oxide (NO) is a key vertebrate biological messenger, playing an important role in neurotransmission, vascular regulation, immune responses and apoptosis. NO , also known as "endothelium-derived relaxing factor" or "EDRF", is synthesized from L-arginine, oxygen and NADPH by various NO synthases. Most of the NO in the cell is oxidized to nitrite (NO2-) and nitrate (NO3-), and therefore the concentrations of these anions are generally as a quantitative measure of NO production.

Images

  • Functional Studies - Nitric Oxide Assay Kit (Colorimetric) (ab65328)
    Functional Studies - Nitric Oxide Assay Kit (Colorimetric) (ab65328)Nam, Eunryel et al., Scientific reports vol. 8,1 7312., Fig 4 , doi:10.1038/s41598-018-25786-x
    Blood NOx concentration measured with ab65328. Blood NOx concentration after application of the NE with NO. The level of blood NOx concentration increased 3-fold after the application of the NE with NO compared with control group.
  • Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)
    Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)

    Nitrite and nitrate measured in cell lysates showing quantity (micromol) per 106 cells

  • Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)
    Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)

    Nitrite and nitrate measured in biologicals showing concentration (micromolar)

  • Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)
    Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)

    Standard curve (colorimetric) : mean of duplicates (+/-SD) with background readings subtracted

  • Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)
    Functional Studies - Nitric Oxide Colorimetric Assay Kit (ab65328)

    Standard curve (Colorimetric) : mean of duplicates (+/-SD) with background readings subtracted

References (99)

Publishing research using ab65328? Please let us know so that we can cite the reference in this datasheet.

ab65328 has been referenced in 99 publications.

  • Feng Y  et al. Pyruvate kinase M2 (PKM2) improve symptoms of post-ischemic stroke depression by activating VEGF to mediate the MAPK/ERK pathway. Brain Behav 12:e2450 (2022). PubMed: 34898024
  • Lin YK  et al. Apolipoprotein (a)/Lipoprotein(a)-Induced Oxidative-Inflammatory a7-nAChR/p38 MAPK/IL-6/RhoA-GTP Signaling Axis and M1 Macrophage Polarization Modulate Inflammation-Associated Development of Coronary Artery Spasm. Oxid Med Cell Longev 2022:9964689 (2022). PubMed: 35096275
  • Abd-Elhakim YM  et al. Time-dependent expression of high-mobility group box-1 and toll-like receptors proteins as potential determinants of skin wound age in rats: Forensic implication. Int J Legal Med 136:1781-1789 (2022). PubMed: 35132471
  • Lau M  et al. Enhanced Antioxidant Effects of the Anti-Inflammatory Compound Probucol When Released from Mesoporous Silica Particles. Pharmaceutics 14:N/A (2022). PubMed: 35335878
  • Mangmool S  et al. Anti-Inflammatory Effects of Mitrephora sirikitiae Leaf Extract and Isolated Lignans in RAW 264.7 Cells. Molecules 27:N/A (2022). PubMed: 35630789
View all Publications for this product

Customer reviews and Q&As

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1-2 of 2 Abreviews

Rat Plasma

 Good Below average 2/5 (Ease of Use)
Abreviews
abreview image
Overall the test worked well for us. All of the required information was included, but the way that the steps were structured was awkward at times. This made the protocol difficult to follow.
Specifically, the section describing how to deproteinize your sample (step 11.4.4 in protocol booklet). An online protocol is also available from abcam, and was a little more clear (https://www.abcam.com/protocols/deproteinization-protocol). It would be nice if these protocols were updated to be more clear and uniform across abcam.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

Submitted Feb 27 2018

Use of nitric oxide colormetric assay on avian plasma samples

 Average Good 4/5 (Ease of Use)
Abreviews
I used this kit to measure nitric oxide (nitrate and nitrite combined) in the plasma of adult male red-winged blackbirds (Agelaius phoeniceus) housed in an outdoor aviary. Preliminary data suggests that the assay requires a minimum of 15uL of plasma per well (ie, 30uL plasma to run in duplicate) to produce repeatable results. Thus far, we have found birds to have levels of NOx between 14-250 uM. It appears that birds with an active infection could have elevated NOx.
Preliminary data also suggests that error can be easily introduced when ready-ing the samples for the assay by deproteinating plasma with PCA. I suspect that this is particularly problematic for samples with absorbance reading less than 0.2, but we need more evidence to be certain.
I did notice that the instructions on the web site were not the same as in the booklet sent. In the booklet I received, the instructions said to use the plate reader after adding the second Griess reagent with no wait time listed. On the online book an incubation time was listed (10 min). There were also inconsistencies in the the list of reagents provided and what was actually provided (We got plates, which is great!). The plates were labeled with a store in -20C, even though they're just plastic plates that isn't necessary.
Technical support was very helpful in answering questions quickly whenever I called - was really happy with the support.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

Submitted Feb 01 2016

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