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NADP+/NADPH Colorimetric Assay Kit (WST-8)

  • Cat.No.:E-BC-K803-M

  • Detection instrument: Microplate reader (450 nm)

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  • 96T
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Protocols for metabolism assays

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Detection principle

Detect total content of NADP+ and NADPH: Glucose 6-phosphate (G6P) is oxidized to 6-phosphate gluconolactone (6-PG) by glucose-6-phosphate dehydrogenase (G6PDH), and NADP+ is reduced to NADPH during this reaction. NADPH, under the action of 1-mPMS, transfer electrons to WST-8 to produce the yellow product, which has a characteristic absorption peak at 450 nm. Therefore, the total content of NADP+ and NADPH can be quantified by measure the OD value at 450 nm. Detect NADPH: After treating sample, heat at 60℃ water bath for 30 min. the NADP+ of the sample is decomposed and only NADPH remains. NADPH reduces WST-8 to form Formazan, and the amount of NADPH is determined by measure the OD value at 450 nm. Detect NADP+ and NADP+/NADPH: The content of NADP+ and the ratio of NADP+/NADPH in the sample can be obtained according to the total content of NADP+ and NADPH obtained of the first two steps as well as the separate content of NADPH.

Performance characteristics

Synonyms NADP/NADPH
Sample type animal tissu, cells
Sensitivity 0.02 μmol/L
Detection range 0.02-5.0 μmol/L
Detection method Colorimetric method
Assay type Quantitative
Assay time 70 min
Precision Average inter-assay CV: 5.500%Average intra-assay CV: 2.100%
Other instruments required Test tube, Micropipettor, 37℃ water bath, 10 KD filters tube.
Other reagents required Ultrapure water
Storage -20℃
Valid period 12 months

Images

The ratio of NADP+/NADPH was significantly changed in different groups. (P<0.05)

T P Cimmino et al investigate the function of formyl-peptide receptor 2 signaling in anabolic pathways. Ratio of NADP+/NADPH in cell was determined using NADP+/NADPH colorimetric assay kit (E-BC-K803-M).

Dilution of sample

It is recommended to take 2~3 samples with expected large difference to do pre-experiment before formal experiment and dilute the sample according to the result of the pre-experiment and the detection range (0.02-5.0 μmol/L).

The recommended dilution factor for different samples is as follows (for reference only):

Sample type

Dilution factor

Jurkat cells

1

Mark cells

1

HCT116 cells

1

293T cells

1

10% Mouse kidney tissue homogenate

1

Hela cells

1

Note: The diluent is reagent 1.

Citations

  1. Antioxidants (2022) IF: 7.675
    Formyl-Peptide Receptor 2 Signaling Redirects Glucose and Glutamine into Anabolic Pathways in Metabolic Reprogramming of Lung Cancer Cells

    DOI: 10.3390/antiox11091692

    PMID: 36139766

    Sample: CaLu-6 cell
  2. ENVIRONMENTAL TOXICOLOGY (2023) IF: 4.109
    GPX4 utilization by selenium is required to alleviate cadmium-induced ferroptosis and pyroptosis in sheep kidney

    DOI: 10.1002/tox.23740

    Sample: Kidney
  3. Journal of Diabetes Investigation (2021) IF: 4.232
    Diet-induced prediabetes: Effects on the activity of the renin-angiotensin-aldosterone system (RAAS) in selected organs

    DOI: 10.1111/jdi.13690

    Sample: skeletal muscle,heart,pancreas
  4. BIOTECHNOLOGY AND BIOENGINEERING (2022) IF: 4.395
    Physiological metabolic topology analysis of Halomonas elongata DSM 2581T in response to sodium chloride stress

    DOI: 10.1002/bit.28222

    PMID: 36062959

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