Overview

  • Product name
    Luminescent ATP Detection Assay Kit
    See all ATP kits
  • Detection method
    Luminescent
  • Assay type
    Quantitative
  • Assay time
    0h 30m
  • Assay duration
    Multiple steps standard assay
  • Product overview

    ATP bioluminescence assays measure the level of ATP within the cell by:
    - lysing the cell sample
    - adding luciferase enzyme and luciferin
    - measuring emitted light using a tube or microplate-based luminometer.


    Luminescent ATP Detection Assay Kit (ab113849) irreversibly inactivates ATP degrading enzymes (ATPases) during the lysis step, ensuring that the luminescent signal obtained truly corresponds to the endogenous levels of ATP.


    Total levels of cellular ATP can be used to assess cell viability, cell proliferation and cytotoxicity of a wide range of compounds and biological response modifiers.


    Review our cell health assays guide to learn more about our other cell viability, cytotoxicity and cell proliferation assay kits.

  • Platform
    Reagents

Properties

Images

  • Example of ATP standard curve using an opaque white plate
    Example of ATP standard curve using an opaque white plate

    The ATP standard curve was prepared as described in the protocol. Background-subtracted data values (mean +/- SD) are graphed.

  • Functional Studies - Luminescent ATP Detection Assay Kit (ab113849)
    Functional Studies - Luminescent ATP Detection Assay Kit (ab113849)
    ab113849 ATP detection kit cytotoxicity data. 25000 HepG2 cells were seeded into each well, allowed to adhere and treated for 4 hours with 25µM rotenone and vehicle control (DMSO) in glucose based complete media. After treatment, cells were lysed, exposed to the ATP substrate solution and signal was measured on a luminescent counter. Mean and standard deviation is plotted for 3 replicates from each condition. Rotenone induces cytotoxicity in HepG2 cells.
  • Simultaneous quantification of mitochondrial respiration and glycolytic flux
    Simultaneous quantification of mitochondrial respiration and glycolytic flux

    Cellular Energy Flux for HepG2 cells (seeded at 65,000 per well), treated with a combination of drug compounds modulating the ETC (Antimycin A [1 µM] and FCCP [2.5 µM]), shown as a percentage relative to untreated control cells. Comparative measurements were taken with Extracellular Oxygen Consumption Assay (ab197243) (white column) and Glycolysis Assay [Extracellular acidification] (ab197244) (black column) show the shift between mitochondrial respiration and glycolysis and the cellular control of energy (ATP; measured 1h post-treatment using Luminescent ATP Detection Assay kit (ab113849) (striped column)).

  • Extracellular detection of ATP (ab113849)
    Extracellular detection of ATP (ab113849)This image is courtesy of an Abreview submitted by Heiko Lemcke.

    Analysis of the release of ATP by connexin hemichannels in stem cells using ATP luminescence kit (ab113849).

    Cells were cultured in HBSS to induce hemichannel opening. Calcium and GAP-inhibitor were used to trigger hemichannel closure.
    After two hours the supernatant was collected and ATP was measured according to the protocol (detergent was also applied).

    Calcium treatment and inhibition by GAP decreased ATP concentration, compared to HBSS control. Graph shows data of three independent experiments.

Protocols

References

This product has been referenced in:
  • Colombo F  et al. Cytokines Stimulate the Release of Microvesicles from Myeloid Cells Independently from the P2X7 Receptor/Acid Sphingomyelinase Pathway. Front Immunol 9:204 (2018). Read more (PubMed: 29467770) »
  • Džinic T & Dencher NA Oxygen Concentration and Oxidative Stress Modulate the Influence of Alzheimer's Disease Aß1-42 Peptide on Human Cells. Oxid Med Cell Longev 2018:7567959 (2018). Read more (PubMed: 29576854) »

See all 21 Publications for this product

Publishing research using ab113849? Please let us know so that we can cite the reference in this datasheet.

Customer reviews and Q&As

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32 reviews or Q&A

10
Votes: Highest First

Extracellular detection of ATP (ab113849)

 Excellent Excellent 5/5 (Ease of Use)
Abreviews
We have analyzed the release of ATP by connexin hemichannels in stem cells using the ATP luminescence kit (ab113849).

Cells were cultured in HBSS to induce hemichannel opening. Calcium and GAP-inhibitor were used to trigger hemichannel closure.
After two hours the supernatant was collected and ATP was measured according to the protocol (we also applied the detergent).

As expected calcium treatment and inhibition by GAP decreased ATP concentration, compared to HBSS control. Graph shows data of three independent experiments.

In summary, we can recommend the ATP luminescence kit (ab113849) for measuring extracellular ATP concentrations.
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Heiko Lemcke

Verified customer

Submitted Jul 31 2017

Measurement of extracellular ATP levels

 Good Excellent 5/5 (Ease of Use)
Abreviews
I enrolled to an Abcam trial to check the suitability of this ATP luminescence kit (ab113849) to detect extracellular ATP levels in cancer cells.

I personally recommend this kit to measure extracellular ATP levels for its ease of use and for the kindly attention and help I received from the Abcam scientific support specialists. When measuring extracellular ATP levels, it is important to avoid light exposure as much as possible and measure the supernatant without FBS and without phenol-red.

Brief description of the protocol:
I seeded 50.000 murine breast-to-brain cancer cells in duplicates or triplicates in 96-well-plate. Once they are attached I starve them in FBS-free and phenol red-free media.

24 hours after starving I collected the supernatant (100 ul) and transfer it to a white opaque plate. There I followed the protocol proposed in the booklet with only one difference: I do not use detergent.

Using the same conditions (cell line, passage, starving time and machine) I repeated 3 times with the same cancer cell line and the results I got are 12,89 nM ATP, 12,30 nM ATP and 7,3 nM ATP.

Regarding the conditions I use, I repeated this procedure several times and I chose this condition (24-hours starvation in phenol red-free and serum-free media) because if I starve them for less hours the luminescence units I get from my samples are super low. If I starve them with HBBS overnight the cells look ugly the following day. Moreover I also tried to add detergent in the media but then the RLU I got were more variable in the same biological replicate. There is also higher variability of RLU if I use media with FBS. I did not try to use media with phenol red because I know it interferes with the luminescence readout. To sum up, I got less variability when I starve them for 24 hours with DMEM without FBS and without phenol red.
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Lisa Sevenich

Verified customer

Submitted Jul 07 2017

Inquiry: Hello, I've used the Luminescent ATP Detection Assay Kit to determine the concentration of ATP in E. coli cells. The kit worked perfectly. My question is, will the lysis protocol denature ATP binding proteins such that they release ATP allowin...

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We haven’t addressed this question experimentally with the reagents.  The answer is probably.  The lysis reagent is denaturing so bound ATP may be made available to the luciferase. This would be difficult to be definitive and different ATP binding prot...

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Is the Luminescent ATP Detection Assay Kit (ab113849) suitable for use with plasma samples, or is it purely for use with cell samples?


In theory it should work, but we don’t know for sure as we have not tested it. Therefore, if you are willing to test and publish the data in the form of review on our website then I would like to invite you to our Abtrial programme.

Inquiry: Could I have any suggestions about the right use of luminometer? In particular the time of reading and the range of wavelength. Are there other parameters to set up? And the bottom of the plate has to be clear or opaque for this assay? Thanks

Thank you for your enquiry.

Luminescence does not have multiple wavelengths. In Luminescence mode, the instrument simply measures the emission of visible light from the well. What is key in the measurement is the PMT (photomultiplier time). Th...

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Is it essential to leave my cells to "rest" or adhere before measuring the signal? I am sorting my cells first and think I might not have enough time

Thank you for contacting us.



The cells do not need to rest or adhere for this assay, you can measure right away.



Can you please provide the customer with the list of the detergents and their concentrations which can harm the luciferase reaction or any other stage of ATP reaction? Thank you very much for your help.

Any anionic detergent will inhibit the luciferase reaction. What is interesting is that non-ionic detergents have the opposite effect and can in fact enhance the luminescent signal. The following detergents used between 0.5 and 1.5% can enhance the sig...

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Our customer intends to use ATP luminescent kit (ab113849) in order to quantify ATP concentration in lysates containing 4% SDS and 50 mM DTT. Please advise if such concentrations of DTT and SDS may influence the ATP kit components and thus the test co...

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SDS is a very strong inhibitor of the luciferase reaction. The lowest possible concentration will likely be below 0.05%. We suggest the customer running a standard ATP curve with and w/o SDS to determine empirically the effect.

Theoretically,...

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Is it possible to use a fluorescent protein quantification kit/reagent with Luminescent ATP detection assay kit (113849)? I'm looking for a method to standarize results obtained with the ab113849 kit eg. to know if the differences in ATP level are not ...

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The mammalian cell lysis solution contains 0.1M of alkaline solution. I don't think it will be compatible with protein quantification. Indeed, changes in ATP may be due to unevenness in seeding or low viability. Using multiple replicates per each dat...

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I'd like to know about possibility of standarization of the results obtained by use of ab113849 kit?

I contacted the laboratory to have more information about the standardization of the results obtained with the ab113849 but unfortunately the lab does not have this kind of information. I am very sorry for this inconvenience.


However, fro...

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1-10 of 32 Abreviews or Q&A

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