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FAD Colorimetric/Fluorometric Assay Kit

based on 3 citations in multiple journalsFAD Colorimetric/Fluorometric Assay Kit34.1 4
Sensitive Assay, HTS
Catalog #: K357
$410.00

Product Details

Cat # +Size K357-100
Size 100 assays
Detection Method Absorbance (570 nm) or Fluorescence (Ex/Em 535/587 nm)
Species Reactivity Mammalian
Applications Applications- FAD functions as the cofactor of an oxidase which catalyze the formation of a product that reacts with OxiRed probe generating color and fluorescence.The lower limit of detection is less than 1 nM FAD.
Features & Benefits • Simple procedure
• Fast and convenient
• The assay is sensitive, stable and high-throughput adaptable.
Kit Components • FAD Assay Buffer
• OxiRed Probe
• Dimethylsulfoxide (DMSO; Dried)
• FAD Enzyme Mix (lyophilized)
• FAD Standard (1 nmol)
• Stop Solution
Storage Conditions -20°C
Shipping Conditions Gel Pack
USAGE For Research Use Only! Not For Use in Humans.

Details

Flavin Adenine Dinucleotide (FAD) is a redox cofactor which plays an important role in metabolism. FAD exists in different redox states and cycles between FAD, FADH and FADH2. The primary sources of reduced FAD in eukaryotic metabolism are the citric acid cycle and the beta oxidation reaction pathways. In BioVision’s FAD Assay Kit, FAD functions as the cofactor of an oxidase which catalyze the formation of a product that reacts with OxiRed probe generating color and fluorescence. FAD can be detected by either colorimetric (λ=570 nm) or fluorometric (Ex/Em=535/587 nm) methods. The kit provides a rapid, simple, ultra-sensitive, and reliable test suitable for high throughput assay of FAD. The lower limit of detection is less than 1 nM FAD.


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We bought the FAD Colorimetric/Fluorometric Assay Kit from your company.I have a few question to consult. "Reading the samples and standards every 5 min. How long should we last this operation? How many times do we need to read ?
Take readings every 5 min until the signal reaches the highest point on the standard curve.
Adding 10 µl of Stop Solution What should we do after that? Should we wait for 24 hours? Should we read the samples within the 24 hours? If we shoud wait for 24 hours what should we do in such hours? Could you tell me more details?
You can take a reading right after you add the stop solution. You can take the same readings any time after the stop solution is added upto the 24 hrs time point.
Can this kit be used with samples like bacteria, plants, drosophila, yeast etc?
We have optimized the kit with mammalian samples. However, theoretically these kits should work with samples from multiple species/sources. Since the optimal conditions depend on the sample type, the protocol has to be be adapted to fit the samples for efficient results. Please refer to this kit's citations to see what kind of samples have been used with this kit other than mammalian samples.
Can we use frozen samples with this assay?
Fresh samples are always preferred over frozen samples. However, frozen samples can also be used, provided, they were frozen right after isolation, were not freeze thawed multiple time (for which we recommend aliquoting the samples before freezing) and have been frozen for relatively short periods.
Can we use a different wavelength than recommended for the final analysis?
It is always recommended to use the exact recommended wavelength for the most efficient results. However, most plate readers have flexibility in their band width of detection in increments of +/- 10 nm. Depending on this flexibility range, you can deviate from the recommended wavelengths within limits.
What is the exact volume of sample required for this assay?
There is no specific volume we can recommend for the amount any sample to be used since it is completely sample concentration and quality based. You have to do a pilot expt with multiple sample volumes to determine the optimal volume which gives a reading within the linear range of the standard curve. Please refer to the citations for this product to see what other clients have used with similar sample types.
Do you have trial sizes of this kit?
Unfortunately, we do not have trial sizes of this kit available. However, if you are based in the US or Canada, we will give you a 10% off list price introductory discount on its purchase price. If you are based out of this area please contact your regional BioVision distributor.
What is the shelf life of this kit?
This kit is good for 12 months from the date of shipment in the unopened form when stored at the appropriate temperature and appropriate conditions. After opening and reconstitution, some of the components in this kit are good for 2 months at -20°C. Please refer to the datasheet for storage information and shelf life of each of the components.
Why are my standard curve values lower than those shown on the datasheet?
There are multiple factors which influence the signals like the incubation times, room temperature, handling etc. In general, to increase the value of the standards, you can increase the incubation time. As long as the standard curve is linear, it should be fine to use, since all of your samples will also be measured under the same conditions on this curve.
How do I normalize my samples against protein concentration
You can use a protein quantitation assay on the supernatants you get from cell/tissue lysates or with any other liquid sample in the assay buffer.
Can we purchase individual components of this kit?
Yes, you can purchase any of the kit's components without the whole kit. Please refer to the component Cat #s mentioned on the datasheet for ordering.
Can we use an alternate buffer for sample preparation (cell lysis, sample dilutions etc)?
Our assay buffers are optimized for the reactions they are designed for. They not only contain some detergents for efficient lysis of your cells/tissue, but also contain some proprietary components required for the further reactions. Therefore, we highly recommend using the buffers provided in the kit for the best results.
Should I make a standard curve for every expt I do, or is one curve/kit enough?
Yes, I would strongly recommend you to do the standards every time you do the expt. There is always a chance that something was done differently that day and we do not want any conditions to differ between standards and samples.
Does the FAD Assay kit measures total FAD pool (FAD+FADH+FADH2) or just the oxidized form (FAD)?
The kit measures the oxidized form of FAD.
Reynolds, Merrick S. et al. (2016) β-Cell deletion of Nr4a1 and Nr4a3 nuclear receptors impedes mitochondrial respiration and insulin secretion. Am J Physiol Endocrinol Metab. 2016 Jul 1;311(1):E186-201.
Wang et al., Identification of riboflavin: revealing different metabolic characteristics between Escherichia coli BL21(DE3) and MG1655. FEMS Microbiol Lett, May 2015; 362: fnv071.
Ying Ni  et al., Germline SDHx variants modify breast and thyroid cancer risks in Cowden and Cowden-like syndrome via FAD/NAD-dependant destabilization of p53, Hum. Mol. Genet., Jan 2012; 21: 300 - 310
Ni. Y. et. al. Germline SDHx variants modify breast and thyroid cancer risks in Cowden and Cowden-like syndrome via FAD/NAD-dependant destabilization of p53. Hum. Mol. Genet., Oct 2011; 10.1093/hmg/ddr459.
For more citations of this product click here