NADP/NADPH Assay Kit (Fluorometric) (ab176724)

Overview

  • Product name
    NADP/NADPH Assay Kit (Fluorometric)
    See all NADP/NADPH kits
  • Sample type
    Cell culture extracts, Tissue Extracts
  • Assay type
    Quantitative
  • Species reactivity
    Reacts with: Human
    Predicted to work with: Mammals
  • Product overview

    Abcam's NADP/NADPH Assay Kit (Fluorometric) (ab176724) provides a convenient method for sensitive detection of NADP, NADPH and their ratio. The enzymes in the system specifically recognize NADP/NADPH in an enzyme recycling reaction that significantly increases detection sensitivity. In addition, this assay has very low background since it is run in the red visible range that considerably reduces the sample interference. The assay can be performed in a convenient 96-well or 384-well microtiter-plate format. Its signal can be easily read by either a fluorescence microplate reader at Ex/Em = 530-570/590-600 nm (maximum Ex/Em = 540/590 nm) or an absorbance microplate reader at ~576 nm. This also provides NADP, NADPH extraction buffer, and cell lysis buffer.

  • Notes

    Nicotinamide adenine dinucleotide (NAD+) and nicotinamide adenine dinucleotide phosphate (NADP+) are two important cofactors found in cells. NADH is the reduced form of NAD+, and NAD+ is the oxidized form of NADH. NAD forms NADP with the addition of a phosphate group to the 2' position of the adenyl nucleotide through an ester linkage. NADP is used in anabolic biological reactions, such as fatty acid and nucleic acid synthesis, which require NADPH as a reducing agent. In chloroplasts, NADP is an oxidizing agent important in the preliminary reactions of photosynthesis. The NADPH produced by photosynthesis is used as reducing power for the biosynthetic reactions in the Calvin cycle of photosynthesis.

Properties

  • Storage instructions
    Store at -20°C. Please refer to protocols.
  • Components 250 tests
    NADP Extraction Solution 1 x 10ml
    NADP/NADPH Control Solution 1 x 10ml
    NADP/NADPH Lysis Buffer 1 x 10ml
    NADP/NADPH Recycling Enzyme Mixture 2 vials
    NADPH Extraction Solution 1 x 10ml
    NADPH Sensor Buffer 1 x 20ml
    NADPH Standard 1 vial
  • Research areas
  • Relevance
    NADP (Nicotinamide adenine dinucleotide phosphate) is a coenzyme composed of ribosylnicotinamide 5-phosphate (NMN) coupled by pyrophosphate linkage to the 5-phosphate adenosine 2,5-biphosphate. It serves as an electron carrier in a number of reactions, being alternately oxidised (NADP+) and reduced (NADPH). The oxidative phase of the pentose phosphate pathway is the major source of NADPH in cells, producing approxiamtely 60% of the NADPH required. NADPH provides the reducing equivalents for biosynthetic reactions and the oxidation-reduction involved in protecting against the toxicity of ROS, allowing the regeneration of GSH. NADPH is also used for anabolic pathways, such as lipid synthesis, cholesterol synthesis and fatty acid chain elongation.

Images

  • Quantity of NADP, NADPH and total (NADP+NADPH) in U937 cells (duplicates; +/- SD). 0.5 x 107 cells were lysed in 1 mL of lysis buffer.

  • Quantity of NADP, NADPH and total (NADP+NADPH) in RAW 264.7 cells (duplicates; +/- SD). 0.5 x 107 cells were lysed in 1 mL lysis buffer.

  • Standard curves with background signal subtracted (duplicates; +/- SD).

Protocols

References

ab176724 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

NADPH measurement for mouse cell lines

Average Good 4/5 (Ease of Use)
Abreviews
The kit seems to work with mouse cell lines: min6b1 cells. But surprisingly Nadph quantity decreases with lower concentration of protein, and nadp is going in the opposite direction.
The protocole of the kit was not that clear ( it should be good to make an update of it) but the technical support is always ready to answer.
Username

Abcam user community

Verified customer

Submitted Jan 17 2017

The lysis buffer is an addition for the kit and it is not essential that you use this lysis buffer to prepare your samples. Most of our customers who are using 96 well plate based on 25 uL/well find that 10 ml of lysis buffer is sufficient.

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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