概述

  • 产品名称
    Free Fatty Acid Assay试剂盒- Quantification
    参阅全部 Fatty acid 试剂盒
  • 样品类型
    Cell culture supernatant, Urine, Serum, Plasma, Other biological fluids, Tissue Extracts
  • 检测类型
    Quantitative
  • 灵敏度
    > 2 µM
  • 检测时间
    0h 40m
  • 产品概述

    Free Fatty Acid Quantification Assay Kit ab65341 provides a convenient, sensitive enzyme-based method for detecting long-chain free fatty acids (FA) in various mammalian and other samples, such as serum, plasma and other body fluids, food, growth media, etc.


    In this assay, FA are converted to their CoA derivatives (coenzyme A), which are subsequently oxidized, leading to formation of color/ fluorescence. Fatty acids can then be easily quantified by either colorimetric (spectrophotometry at λ= 570 nm) or fluorometric (at Ex/Em= 535/587 nm)



    Visit our FAQs page for tips and troubleshooting.


    Review our Metabolism Assay Guide to learn about assays for metabolites, metabolic enzymes, mitochondrial function, and oxidative stress, and also about how to assay metabolic function in live cells using your plate reader.

  • 说明

    This assay detects formation of C-8 (octanoate) and longer fatty acids.

性能

图片

  • Plasma FFA levels was measured using ab65341 in male and female wild-type (WT) or AT2KO mice either on normal diet (ND) or high fat diet (HFD).

  • Colorimetric standard curve: mean of duplicates (+/-SD) with background readings subtracted.

  • Free Fatty Acid measured in biologicals showing concentration (µM)

  • Flourometric standard curve: mean of duplicates (+/-SD) with background readings subtracted.

实验方案

文献

This product has been referenced in:
  • Harm S  et al. Removal of stabilizers from human serum albumin by adsorbents and dialysis used in blood purification. PLoS One 13:e0191741 (2018). Human . Read more (PubMed: 29364955) »
  • Shimazu T  et al. Role of METTL20 in regulating ß-oxidation and heat production in mice under fasting or ketogenic conditions. Sci Rep 8:1179 (2018). Read more (PubMed: 29352221) »
See all 40 Publications for this product

客户评价及客户问答

21-30 of 34 Abreviews or Q&A

Answer


I'm afraid the exact details of the mechanism are proprietary, however we do have this description as noted in the Protocol Booklet:
"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
= 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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Answer

Repeating in triplicate certainly reduces the margin of error in any reading, however you should be able to produce a linear curve using the protocol found in the kit booklet.

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Answer

The kit should be able to detect fatty acids of chain lengths between C8 and above with equal efficiency and you should therefore not need to perform a different standard for the caprylic acid and palmitic.

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Answer

I can now confirm that no cross reactivity would be expected with the Free Fatty Acid Quantification Kit (ab65341) with lechitins, phosphoatidyl choline or other triglycerides. This kit is specific for free fatty acids and would require an enzyme to convert the triglycerides to free fatty acids to be detected by the kit.

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Answer


Yes, these kits are suitable for all mammalians.

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Question
Answer

If your sample reading was 2.5 nmol of fatty acid, and you used 6uL of serum per well, the calculations would be as follows: (2.5 nmol / 6uL) * (256 ng / nmol) = 106.67 ng / uL = 106.67 * (10-9 g / 10-6 L) = (106.67 / 10) (10-3 g / 10-1 L) = 10.667 mg/dL

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Answer

The standard wells contain 0, 2, 4, 6, 8, 10 nmol per well. The molecular weight of the standard, palmitic acid, is 256 g/mol, or 256 ng/nmol. so each standard well contains, in micrograms: 0, 0.51, 1.02, 1.54, 2.05, 2.56 After subtracting the optical density value at 570 nm of the blank (0) well from all other wells, plot the optical densities of your samples against the optical densities of the standards to obtain the amount of fatty acid per well of your samples, in micrograms. To obtain the ug/ul concentration of fatty acid in the samples, divide the micrograms per well by the number of microliters of sample that were added per well.

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Answer

Here is what the lab is suggesting: Serum samples can be tested directly in the following manner: For testing liquid samples, different volume of samples can be directly added to each well in a 96-well plate, then bring up the volume to 50 µl/well with Assay Buffer. For unknown samples, we suggest using different doses to ensure the readings are within the standard curve range. If the kit is to be used for extended times we would recommend aliquoting and storing the enzyme mix, ACS reagent, probe and enhancer just to be on the safest side.

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Answer

I have contacted the lab and advice that after pelleting the cells by trypsinization you follow these steps: 106 cells or 10 mg tissue samples can be extracted by homogenization with 200 µl of chloroform-Triton X-100 (1% Triton X-100 in pure chloroform) in a microhomogenizer. Then spin the extract  5-10 minutes at top speed in a microcentrifuge. Collect organic phase (lower phase), air dry at 50°C to remove chloroform. Vacuum dry 30 min to remove trace chloroform. Dissolve the dried lipids (in Triton X-100) in 200 µl of Fatty Acid Assay Buffer by vortexing extensively for 5 mins (Note: The solution may be slightly turbid or opalescent, but this does not affect the assay). The extraction procedure can be proportionally scaled up if larger amount of sample is desired. Use 1- 50 µl of the extracted sample per assay.

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21-30 of 34 Abreviews or Q&A

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

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