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Nitric Oxide Assay Kit (Fluorometric) (ab65327)

Key features and details

  • Assay type: Quantitative
  • Detection method: Fluorescent
  • Platform: Microplate reader
  • Assay time: 3 hr
  • Sample type: Cell Lysate, Other biological fluids, Plasma, Serum, Tissue Extracts, Urine

Overview

  • Product name

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

  • Detection method

    Fluorescent

  • Sample type

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

  • Assay type

    Quantitative

  • Assay time

    3h 00m

  • Product overview

    Nitric Oxide Assay Kit (Fluorometric) ab65327 provides an accurate, convenient measure of total nitrate/nitrite concentration in a simple two-step process.


    In the nitric oxide assay protocol:
    - the first step converts nitrate to nitrite by nitrate reductase
    - in the second step, nitrite reacts with the fluorescent probe DAN (2, 3 diaminonaphthalene)


    The fluorescence can be measured at Ex/Em = 360/450 nm. NaOH enhances the fluorescent yield. The fluorescent intensity is proportional to the total nitric oxide production. The kit has been tested with culture media, plasma, and tissue homogenates.


    Nitric oxide assay protocol summary:
    - add samples and standards to wells
    - add enzyme cofactor, to measure nitrite only add assay buffer or to measure total nitrate + nitrite add nitrate reductase 
    - incubate for 1-4 hr at room temp to convert nitrate to nitrite
    - add enhancer and incubate for 30 min at room temp
    - add DAN probe and incubate for 10 min at room temp
    - add NaOH and incubate for 10 min at room temp
    - analyze with microplate reader

  • Notes

    If you are interested in a colorimetric detection kit, please check Nitric Oxide Assay Kit (Colorimetric) (ab65328).

    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. Since NO is rapidly converted to nitrite (NO2-) and nitrate (NO3-), the total concentration of nitrite and nitrate is used as a quantitative measure of NO production.

  • Platform

    Microplate reader

Properties

  • Storage instructions

    Store at -20°C. Please refer to protocols.
  • Components Identifier 200 tests
    Assay Buffer WM 1 x 40ml
    DAN Probe Amber Red 1 x 1ml
    Enhancer (Lyophilized) Purple 1 vial
    Enzyme Cofactor (Lyophilized) Blue 1 vial
    Microtiter plate 2 units
    Nitrate Reductase (Lyophilized) Green 1 vial
    Nitrate Standard (Lyophilized) Yellow 1 vial
    Nitrite Standard (Lyophilized) Orange 1 vial
    Plate Cover 2 units
    Sodium Hydroxide Clear 1 x 1ml

  • 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 (Fluorometric) (ab65327)
    Functional Studies - Nitric Oxide Assay Kit (Fluorometric) (ab65327) Komalavilas, Padmini et al., PloS one vol. 12,11 e0188069., Fig 7, doi:10.1371/journal.pone.0188069
    Effect of arginase inhibitor and L-arginine on eATP-induced reduction of nitric oxide production in HSV endothelial cells.
    HSVEC were either untreated (Ctrl), treated with ATP (2 mM), L-NAME, (100 µM), ATP with L-arginine (L-Arg, 2 mM), or ATP with NOHA (10 µM), for 2 hours. The cells were then stimulated with carbachol (CCH, 1 µM) for 10 minutes and the nitric oxide generated was measured as nitrite using the NO assay kit (ab65327) and relative percent of NO generated was calculated. NO generated with CCH was set as 100%, n = 6 passages, ∗p
  • Functional Studies - Nitric Oxide Fluorimetric Assay Kit (ab65327)
    Functional Studies - Nitric Oxide Fluorimetric Assay Kit (ab65327)

    Nitric oxide measured in cell lysates showing quantity (micromol) per 106 cells tested

  • Functional Studies - Nitric Oxide Fluorometric Assay Kit (ab65327)
    Functional Studies - Nitric Oxide Fluorometric Assay Kit (ab65327)

    Nitric oxide measured in biologicals (average of 1:6 and 1:12 dilutions) showing concentration (micromolar)

  • Functional Studies - Nitric Oxide Fluorometric Assay Kit (ab65327)
    Functional Studies - Nitric Oxide Fluorometric Assay Kit (ab65327)
    Nitrite, nitrate assay in the presence and absence of nitrate reductase. Assays were performed according to the kit protocol with 1 hour conversion of nitrate to nitrite at Step 5.
  • Functional Studies - Nitric Oxide Fluorimetric Assay Kit (ab65327)
    Functional Studies - Nitric Oxide Fluorimetric Assay Kit (ab65327)

    Nitric oxide measured in cell lysates showing quantity (micromol) per 106 cells tested

  • Functional Studies - Nitric Oxide Fluorometric Assay Kit (ab65327)
    Functional Studies - Nitric Oxide Fluorometric Assay Kit (ab65327)

    Nitric oxide measured in biologicals (average of 1:6 and 1:12 dilutions) showing concentration (micromolar)

  • Functional Studies - Nitric Oxide Fluorometric Assay Kit (ab65327)
    Functional Studies - Nitric Oxide Fluorometric Assay Kit (ab65327)
    Nitrite, nitrate assay in the presence and absence of nitrate reductase. Assays were performed according to the kit protocol with 1 hour conversion of nitrate to nitrite at Step 5.

    • Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
    For licensing inquiries, please contact partnerships@abcam.com

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