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Product Details
| Cat # +Size | K612-100 |
|---|---|
| Size | 100 assays |
| Detection Method | Absorbance (570 nm) or Fluorescence (Ex/Em 535/587 nm) |
| Species Reactivity | Mammalian |
| Applications | The kit has a detection limit 2 µM free fatty acid in variety samples. |
| Features & Benefits | • Simple procedure; takes ~40 minutes • Fast and convenient • Kit contains all necessary reagents for rapid, sensitive and accurate measurement of Adipolysis in cell culture samples |
| Kit Components | • Fatty Acid Assay Buffer • Fatty Acid Probe (in DMSO) • ACS Reagent • Enzyme Mix • Enhancer • Palmitic Acid Standard (1nmol/µl) |
| Storage Conditions | -20°C |
| Shipping Conditions | Gel Pack |
| USAGE | For Research Use Only! Not For Use in Humans. |
Details
Fatty Acids play very important roles in normal metabolism and many disease developments. They are precursors to a number of bioactive classes of compounds such as prostaglandins, leucotrienes and others, and have been implicated in diverse functions such as autism, immune system and inflammation response. BioVision’s Free Fatty Acid Quantification Kit provides a convenient, sensitive enzyme-based method for detecting the long-chain free fatty acids in various biological samples, such as serum, plasma and other body fluids, food, growth media, etc. In the assay, Fatty Acids are converted to their CoA derivatives, which are subsequently oxidized with the concomitant generation of color or fluorescence. C-8 (octanoate) and longer fatty acids can then be easily quantified by either colorimetric (spectrophotometry at λ = 570 nm) or fluorometric (at Ex/Em = 535/587 nm) methods with detection limit 2 µM free fatty acid in variety samples.
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What is the detection limit of K612?
The detection limit for fluorometric assay is 10X more than the colorimetric assay. The detection limit for colorimetric assay isa. The lower limit of detection is calculated by taking 2 nmol (lowest conc of Standard) divided by 50 µl (max sample volume)=2 nmol/50µl= 0.0.04 nmol/µl
b. The highest limit of detection is calculated by taking 10 nmol (highest conc of Standard) divided by 1 µl (min sample volume)=10 nmol/1 µl= 10 nmol/µl
Can I stop and store samples mid-way through the sample preparation?
Yes, you can store dried lipids in 200 µl of Fatty Acid Assay Buffer at -80°C. If the lipids precipitate after freeze/thaw, re-dissolve the lipids by sonication.
When comparing Free Fatty Acid (FFA) concentrations in human plasma and human serum, why is serum about twice the concentration? Do you have any information?
Majority of the FFA is bound to serum albumin, which is a major transporter of FFA. Hence there is more FFA in serum as compared to plasma samples.
Which anticoagulants interfere with this assay and which can be used?
Blood needs to be collected using an anticoagulant such as EDTA, sodium citrate, sodium fluoride, or ammonium oxalate. Heparinized plasma is not the best choice as high amounts of heparin could interfere with the assay.
What is the concentration of fatty acid in human serum?
Normal Human Serum has 80-250 ug/ml Fatty acid.
Why is there a large background even after using the correct Excitation/Emission settings?
It seems that the problem is the fluorescence reading overlap between excitation an emission readings for the particular instrument. If a fluorometer has window of sensitivity +/-40nm, the excitationa nd emission overlap and then oit is needed to resort to a longer wavelength for emission which is above 600 nm (Some times up to 620 nm).
Will the phenol red in the media affect the assay readout?
Very low amounts of media are used for each sample. This will generate a very low background at the best. Please use only media as a background control and subtract this reading from all sample readings to accommodate for the phenol red.
What is the length of fatty acid chains that can be quantified using this kit? A palmitic acid std is used, but are oleic and stearic acid chains able to be detected as well?
C-8 (octanoate) and longer fatty acids can be quantified. There is no difference upon measurement of chains C-16 and above.
What is the detection limit for this kit ?
The detection limit is ~2 µM fatty acid.
Can the lipid extraction solution from K610 be used for cells/tissues in this kit?
Homogenization in 1% Triton X-100 in pure chloroform is much more effective than the Lipid extraction solution from K610 and hence is not recommended. Moreover the chloroform is dried in this assay to get lipids which is not possible with the lipid extraction solution.
How long should samples be air-dried ?
Typically up to 1 hour is enough to air- dry at 50C to remove chloroform. The goal to remove as much chloroform at this step as possible.
Are trial sizes of this kit available?
Unfortunately, we do not have trial sizes of this kit available. However,for customers based in the US or Canada, a 10% off list price introductory discount can be offered on the list price. For all other customers based out of this area our regional BioVision distributor should be contacted
Can protein content be used as an internal control for this assay?
Since the tissues/cells are homogenized in chloroform/Triton a protein assay is not recommended.
Can frozen samples be used 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.
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.
Can we use rat fecal samples with this kit? Can you provide a protocol?
Fecal samples are not tested with this kit and therefore, we do not have a protocol for sample processing. The customer can try to extract FFA from fecal samples using their own protocol and test it with the kit.
Takahito Otani, Osteocalcin triggers Fas/FasL-mediated necroptosis in adipocytes via activation of p300. Cell Death Dis., Dec 2018; 30546087.
Marcel Nel, et al., (2018) Modes of cell death induced by tetrahydroisoquinoline-based analogs in MDA-MB-231 breast and A549 lung cancer cell lines, Drug Design, Development and Therapy, Jun. 2018, 29983544
Kei Miyakawa, et al., (2018) Development of a cell-based assay to identify hepatitis B virus entry inhibitors targeting the sodium taurocholate cotransporting polypeptide., Oncotarget, May 2018, 29805766
Wenliang Zhang, et al., (2018) Adipose-specific lipin1 overexpression in mice protects against alcohol-induced liver injury, Scientific Reports, Feb. 2018, 29323242
Zhang W., et al., (2018) , Adipose-specific lipin1 overexpression in mice protects against alcohol-induced liver injury, Sci. Rep., 2018, 8:408
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