NADP/NADPH Ratio Assay Kit (Red Fluorescence)
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Catalog Number:
CA-N426
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Detailed Description
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CA-N426
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Name
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Elite™ NADP/NADPH Ratio Assay Kit (Red Fluorescence)
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Description
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This Elite™ Fluorimetric NADP/NADPH Ratio Assay Kit 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.
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Application
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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 = 540/590 nm or an absorbance microplate reader at ~576 nm.
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Size
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250 assays in 96-well plates
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Ex/Em
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540/590 nm
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Detection
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Fluorescence microplate reader or absorbance microplate reader
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For Downloading
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===> DATA SHEET ===> MSDS
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Selected Citation:
LncRNA MACC1-AS1/MACC1 enhances the progression of glioma via regulating metabolic plasticity. Cell Cycle 19, 2286–2297 (2020)
Frequently Asked Questions
1. Can we use this ratio assay kit to also detect NADP or NADPH separately?
Answer: Yes. You could measure the NADOH amount with Component D (NADPH Extraction Solution), and measure the NADP amount with NADP Extraction Solution. But according to our experience, the NADP concentrations in cells are a lot higher than the NADPH concentrations. We suggest to measure the Total NADP+NADPH amount and the NADP amount, then to calculate the NADPH amount by subtraction, which gives more accurate results.
2. Why PBS is used to dilute the Standard? Since the samples are in the lysis buffer, do you think I should use the lysis buffer to dilute them?
Answer: PBS is commonly used. People usually use the lysis buffer to lyse the cells, then use PBS to dilute the samples. The point is that you should keep the same concentration of the lysis buffer for the samples and the Standards.
3. I did a comparison, and it showed that the PBS based Standard curve looked better than the lysis buffer based Standard curve. What do you think of this?
Answer: The bubbles in the lysis buffer may cause the inconsistency of the readings.
4. When I do duplicates of Standards, the reading varies a lot in the low range (0.2 uM or lower). Why does that happen?
Answer: It is normal. There are many factors that would cause inconsistent readings, i.e. bubbles in the wells, the volumes.... So try doing more replicates.
5. Sometimes for the 2 sets of Standards I set up, they look good (R2=0.99) individually but the curve is messed up (R2=0.95) when merged. So which curve should I use to quantitate my data?
Answer: You'll need to use the curve set up on the same plate with your samples.
6. We don't have fluorescence microplate reader, so can we use absorbance microplate reader?
Answer: Yes. But the absorbance readings have much lower sensitivity compared to the fluorescence readings. Try using the ratio of 570 to 610 to increase the sensitivity.
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