Overview

  • Product name
    Acetyl CoA Assay Kit
  • Sample type
    Cell Lysate, Tissue Lysate
  • Assay type
    Quantitative
  • Sensitivity
    > 0.01 nmol/well
  • Range
    0.01 nmol/well - 1 nmol/well
  • Product overview

    Abcam's PicoProbe Acetyl CoA Assay Kit is a highly sensitive assay for determining Acetyl CoA level in a variety of biological samples. In the assay, free CoA is quenched then Acetyl CoA is converted to CoA. The CoA is reacted to form NADH which interacts with PicoProbe to generate fluorescence (Ex=535/Em=587 nm). The assay can detect 10 to 1000 pmol of Acetyl CoA (with detection limit ~0.4 µM) in a variety of samples.
    Visit our FAQs page for tips and troubleshooting.

  • Notes

    Acetyl CoA is a central molecule of metabolism. It carries acetate, used in the build-up and breakdown of larger molecules. Acetyl CoA is key in synthetic pathways leading to sesquiterpenes, precursors to cholesterol and other sterols, flavenoids and other polyketides, polyenes and long-chain fatty acids. It is the source of the acetyl group used in histone acetylation. The acetyl group is also incorporated into a variety of other molecules such as acetylcholine, melatonin, heme and TCA cycle intermediates.

Properties

Images

  • Examples of standard curves generated following the ab87546 kit protocol.
  • Induction of acetyl-CoA in mitochondria by palmitate. Mitochondria were isolated from cells after palmitate treatment (300 μM) at 1, 4 and 16 h, and then used in the concentration assay of acetyl-CoA. 

    The acetyl-CoA content was determined immediately after isolation of the mitochondria using ab87546. Briefly, acetyl-CoA standard curve was made in the range of 0–100 pM and the correlation coefficient was 0.990 or higher. Protein was removed in the sample using the perchloric acid protocol and the supernatant was neutralized with 3 M KHCO3. The CoASH Quencher and Quencher remover were added into the sample to correct the background generated by free CoASH and succ-CoA. The sample was then diluted with the reaction mix, and the fluorescence signal was measured at Ex/Em = 535/589 nm with Spectra max Gemini XPS (Molecular Devices, Sunnyvale, CA). The relative acetyl-CoA concentration was normalized with the mitochondrial protein.

Protocols

References

This product has been referenced in:
  • Jung TW  et al. ß-aminoisobutyric acid attenuates LPS-induced inflammation and insulin resistance in adipocytes through AMPK-mediated pathway. J Biomed Sci 25:27 (2018). Functional Studies . Read more (PubMed: 29592806) »
  • Ravera S  et al. Concentration-dependent metabolic effects of metformin in healthy and Fanconi anemia lymphoblast cells. J Cell Physiol 233:1736-1751 (2018). Read more (PubMed: 28681917) »

See all 15 Publications for this product

Customer reviews and Q&As

1.We recommend using ~2-5 X 10 ^ 6 cells for each manipulation.
2. Deproteinization is recommended for cells in culture. I have attached the protocol that the lab recommends for this step.

In principle, eliminating the quenching and conversion step will allow you to measure CoA alone. However, the kit was not produced nor validated for this purpose, so we cannot guarantee that it will work, nor do we have a CoA standard that has been val...

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J’ai contacté le laboratoire concernant le kit ab87546 :

1.Nous recommandons d’utiliser ˜2-5 X 10^6 cellules pour chaque manip.

2. Il est recommandé de faire une déprotéinisation pour des cellules en culture. J’ai attaché ...

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1. Yes it is very crucial to do the sample prep exactly as recommended as Acetyl CoA is very sensitive to degradation and you risk the chance of inefficient results if you make changes to the protocol.
2. The deproteinization approach, though not a...

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Thank you for contacting us.

I have had the opportunity to discuss the specificity of this kit with the lab as you have requested. I have found that in this particular assay the probe is specific to just the NADH.

I hope this ...

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Thank you for your interest in our acetyl CoA assay kit, ab87546.

The assay measures relative amounts of acetyl CoA in samples indirectly, compared to a standard included with the kit, by converting the acetyl CoA to CoA, and then using the ...

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High background values in the fission yeast

Poor Average 3/5 (Ease of Use)
Abreviews
Protocol:
Two methods of sample preparation were tested.
1. Cell lysis by bead beating in acetyl-CoA assay buffer, deproteinization by 10 kDa spin column
2. Cell lysis by bead beating in TCA which also serves to denature proteins, neutralization by Na2CO3 to pH ~7
Extracts were prepared from fresh yeast biomass of S. pombe cells grown to exponential phase and processed as quickly as possible due to acetyl-CoA instability. Variable sample volumes in the assay - 5, 10 or 50 ul (samples diluted to 50 ul by assay buffer) were used. The standards and samples were measured in the 0-100 pmol range.

Figure legend:
A. Standard curve in the 0-100 pmol range. B. Fluorescence values of samples prepared from two yeast strains (A and B) using the 10kDa spin column. The acquired values of background samples (without conversion enzyme) are higher than those of samples. C. Fluorescence values of samples prepared from two yeast strains (A and B) using TCA precipitation. The acquired values of background samples is lower than those of samples only when 10 ul extract were used. Moreover, high volume (50 ul) of sample seems inhibitory for the assay.
Username

Jarmila Tvarůžková

Verified customer

Submitted Feb 08 2018

The sample can be stored at -80 degree centigrade once prepared. However, we recommend to use the sample immediately after preparation for maximum efficiency.

I can confirm that kit PicoProbe Acetyl CoA Assay Kit (ab87546) can be used with frozen mouse tissues. Please see the protocol booklet for more information regarding sample preparation.

1. Mitochonrial lysate can be assayed by ab87546.

2. For the amount sample to be added into each sample well, we recommended to perform a pilot experiment with a range of dilutions of the sample to determine the optimal concentration/dilutio...

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1-10 of 19 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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