For the best experience on the Abcam website please upgrade to a modern browser such as Google Chrome

Hello. We're improving abcam.com and we'd welcome your feedback.

Hello. We're improving abcam.com and we'd welcome your feedback.

Infomation icon

We haven't added this to the BETA yet

New BETA website

New BETA website

Hello. We're improving abcam.com and we'd welcome your feedback.

Take a look at our BETA site and see what we’ve done so far.

Switch on our new BETA site

Now available

Search and browse selected products

  • A selection of primary antibodies

Purchase these through your usual distributor

In the coming months

  • Additional product types
  • Supporting content
  • Sign in to your account
  • Purchase online
United States
Your country/region is currently set to:

If incorrect, please enter your country/region into the box below, to view site information related to your country/region.

Call (888) 77-ABCAM (22226) or contact us
Need help? Contact us

  • My account
  • Sign out
Sign in or Register with us

Welcome

Sign in or

Don't have an account?

Register with us
My basket
Quick order
Abcam homepage

  • Research Products
    By product type
    Primary antibodies
    Secondary antibodies
    ELISA and Matched Antibody Pair Kits
    Cell and tissue imaging tools
    Cellular and biochemical assays
    Proteins and Peptides
    By product type
    Proteomics tools
    Agonists, activators, antagonists and inhibitors
    Cell lines and Lysates
    Multiplex miRNA assays
    Multiplex Assays
    By research area
    Cancer
    Cardiovascular
    Cell Biology
    Epigenetics
    Metabolism
    Developmental Biology
    By research area
    Immunology
    Microbiology
    Neuroscience
    Signal Transduction
    Stem Cells
  • Customized Products & Partnerships
    Customized Products & Partnerships

    Customized products and commercial partnerships to accelerate your diagnostic and therapeutic programs.

    Customized products

    Partner with us

  • Support
    Support hub

    Access advice and support for any research roadblock

    View support hub

    Protocols

    Your experiments laid out step by step

    View protocols

  • Events
    • Conference calendar
    • Cancer
    • Cardiovascular
    • Epigenetics & Nuclear signaling
    • Immunology
    • Neuroscience
    • Stem cells
    • Tradeshows
    • Scientific webinars
    Keep up to date with the latest events

    Full event breakdown with abstracts, speakers, registration and more

    View global event calendar

  • Pathways
    Cell signalling pathways

    View all pathways

    View all interactive pathways

Supporting our customers and employees during the COVID-19 pandemic. Read more

  1. Link

    adpatp-ratio-assay-kit-bioluminescent-ab65313.pdf

  1. Send me a copy of this email
    I agree to the terms and conditions.
Kits/ Lysates/ Other Kits Cell Metabolism Kits Other Metabolism Assay
Share by email

ADP/ATP Ratio Assay Kit (Bioluminescent) (ab65313)

  • Datasheet
  • SDS
  • Protocol Booklet
Reviews (2)Q&A (39)References (73)

Product price, shipping and contact information

Currently unavailable

Sorry, we can't display this right now.

Please contact us to place your order, or try again later.

 

Loading size & price…

 

Shipping and order information

Shipping info

Promotion Information

Abpromise

Guaranteed product quality, expert customer support.

Find out more.

Functional studies - ab65313
  • Functional studies - ab65313
  • Functional studies - ab65313

Key features and details

  • Assay type: Semi-quantitative
  • Detection method: Luminescent
  • Platform: Microplate reader
  • Assay time: 30 min
  • Sample type: Adherent cells, Suspension cells, Tissue
  • Sensitivity: 100 cells/well

You may also be interested in

Assay
Product image
Luminescent ATP Detection Assay Kit (ab113849)
Assay
Product image
Extracellular Oxygen Consumption Assay (ab197243)
Kit
Product image
Deproteinizing Sample Preparation Kit - TCA (ab204708)

View more associated products

Overview

  • Product name

    ADP/ATP Ratio Assay Kit (Bioluminescent)
  • Detection method

    Luminescent
  • Sample type

    Tissue, Adherent cells, Suspension cells
  • Assay type

    Semi-quantitative
  • Sensitivity

    < 100 cells/well
  • Assay time

    0h 30m
  • Product overview

    ADP/ATP Ratio Assay Kit (Bioluminescent) ab65313 is based on the bioluminescent detection of ADP and ATP levels. It can be used for a rapid screening of apoptosis, necrosis, growth arrest, and cell proliferation simultaneously in mammalian cells.


    In the ADP/ATP assay protocol, luciferase catalyzes the conversion of ATP and luciferin to light, which in turn can be measured using a luminometer or Beta Counter. The ADP level is measured by its conversion to ATP that is subsequently detected using the same reaction.


    The assay can be fully automatic for high throughput and is highly sensitive (detects 100 mammalian cells/well).


    ADP/ATP Ratio assay protocol summary:
    - transfer suspension cells, or nucleotide releasing buffer treated adherent cells, to plate
    - add ATP reaction mix and incubate for 2 min
    - analyze with luminescence plate reader to measure ATP
    - after preparing ADP reaction mix and measuring luminescence levels again, add ADP reaction mix to same wells and incubate for 2 min
    - analyze with luminescence plate reader to measure ADP

  • Notes

    This product is manufactured by BioVision, an Abcam company and was previously called K255 ADP/ATP Ratio Bioluminescence Assay Kit, ApoSENSOR. K255-200 is the same size as the 200 test size of ab65313.

    Changes in the ADP/ATP ratio have been used to differentiate the different modes of cell death and viability. Increased levels of ATP and decreased levels of ADP have been recognized in proliferating cells. In contrast, decreased levels of ATP and increased levels of ADP are recognized in apoptotic cells. The decrease in ATP and increase in ADP are much more pronounced in necrosis than apoptosis.

    Related assays

    Review the cell health assay guide to learn about kits to perform a cell viability assay, cytotoxicity assay and cell proliferation assay. 

    Review the 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.

  • Platform

    Microplate reader

Properties

  • Storage instructions

    Store at -20°C. Please refer to protocols.
  • Components Identifier 200 tests
    ADP Converting Enzyme (Lyophilised) Blue Cap 1 vial
    ATP Monitoring Enzyme (Lyophilised) 1 vial
    Enzyme Reconstitution Buffer 1 x 2.15ml
    Nucleotide Releasing Buffer 1 x 50ml
  • Research areas

    • Kits/ Lysates/ Other
    • Kits
    • Cell Metabolism Kits
    • Other Metabolism Assay
    • Kits/ Lysates/ Other
    • Kits
    • Cell Metabolism Kits
    • Intermediary Metabolism Kits
  • Relevance

    The changes in ADP/ATP ratio have been used to differentiate the different modes of cell death and viability. Increased levels of ATP and decreased levels of ADP have been recognized in proliferating cells. In contrast, decreased levels of ATP and increased levels of ADP are recognized in apoptotic cells. The decrease in ATP and increase in ADP are much more pronounced in necrosis than apoptosis.

Associated products

  • Assay kits

    • Luminescent ATP Detection Assay Kit (ab113849)
    • ATP Assay Kit (Colorimetric/Fluorometric) (ab83355)

Images

  • Functional studies - ab65313
    Functional studies - ab65313Image from Amrani A et al., PLoS One 9(9). Fig 5A. doi: 10.1371/journal.pone.0106831 Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

    Quantitation of glutamate levels (A) using ab138883 and intracellular ATP (B) using ab65313 in D. hydrothermalis cells grown under different pressure conditions.

  • Functional studies - ab65313
    Functional studies - ab65313Bhattacharyya S.,PLoS One 8(11), Fig 5d & f. doi: 10.1371/journal.pone.0079167 Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

    ADP/ATP ratio in cystathionine-beta-synthase slienced A2780 (Ovarian cancer cell line) cells and AOAA (aminooxyacetic acid) treated A2780 cells were measured using ADP/ATP ratio assay kit (ab65313).

     

  • Functional studies - ab65313
    Functional studies - ab65313Image from Julien S.G., PLoS Biol 15(2), Fig 8C. doi: 10.1371/journal.pbio.1002597. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

    ADP/ATP ratio were examined in both narciclasine (ncls) and vehicle (veh) treated C2C12 myotubes with or without PA treatment using ADP/ATP ratio assay kit (ab65313). 

Protocols

  • Protocol Booklet

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (73)

Publishing research using ab65313? Please let us know so that we can cite the reference in this datasheet.

ab65313 has been referenced in 73 publications.

  • Wang Y  et al. Improvement of obesity-associated disorders by a small-molecule drug targeting mitochondria of adipose tissue macrophages. Nat Commun 12:102 (2021). PubMed: 33397994
  • Hogarth K  et al. Singular and short-term anesthesia exposure in the developing brain induces persistent neuronal changes consistent with chronic neurodegenerative disease. Sci Rep 11:5673 (2021). PubMed: 33707598
  • Hung CM  et al. AMPK/ULK1-mediated phosphorylation of Parkin ACT domain mediates an early step in mitophagy. Sci Adv 7:N/A (2021). PubMed: 33827825
  • Olson GS  et al. Type I interferon decreases macrophage energy metabolism during mycobacterial infection. Cell Rep 35:109195 (2021). PubMed: 34077724
  • Herrero M  et al. The Energy Status of Astrocytes Is the Achilles' Heel of eIF2B-Leukodystrophy. Cells 10:N/A (2021). PubMed: 34440627
View all Publications for this product

Customer reviews and Q&As

Show All Reviews Q&A
Submit a review Submit a question

1-10 of 41 Abreviews or Q&A

ADP/ATP Ratio Assay in M. tuberculosis

Excellent Excellent 5/5 (Ease of Use)
Abreviews
Abreviews
abreview image
We tested ab65313 ADP/ATP Ratio Assay in M. tuberculosis (Mtb) strains H37Rv and 18b. The main goal was to determine if the Nucelotide Releasing Buffer (NRB) was able to cause release of nucleotides from Mtb with or without additional disruption (bead beating).
Experimental set-up:
Cells: Mtb strains H37Rv and 18b were grown in 7H9 media and harvested at an OD600 of ~0.4. 175 Million total bacteria were used from each strain.
Control: 7H9 alone
Treatments:
- "Lysis": NRB (5-10 minutes)
- "Beads": NRB (5-10 minutes) + bead-beading (speed 6.60 - 3 x 30s runs)
Measurement:
ab65313 on a Pherastar Plus plate reader on bioluminescent setting
Standard Curve:
ATP disodium salt (ab120385) dilutions to create a standard curve.
Results:
Standard curve: log10(RLU) = 3.739 + 0.938 * log10(ATPconc)
Boxplot of ADP and ATP concentrations on two technical replicates on each of two biological replicates. #N.B. The gain setting on the luminometer resulted in ADP readings (Data D of the kit) for the bead-beating groups that were off-scale and thus non-quantifiable, but still plotted in the attached figure.
Discussion:
NRB alone did not lead to appreciable release of nucleotides from two strains of Mtb. The addition of bead beating to NRB led to significant increases in measured concentrations of both nucleotides in both strains.

Mr. Gregory Olson

Verified customer

Submitted Oct 06 2017

Very sensitive and easy to use

Excellent Excellent 5/5 (Ease of Use)
Abreviews
Abreviews
We have used this product extensively to measure changes in the ATP/ADP ratio in response to changes in glucose availability and during stress tests (H2O2 treatment). We have some recommendations below for use:

1) Use a black-walled 96-well plate to reduce plate phosphorescence
2) If possible, use ~1,000 cells per well. If you can do a seeding density curve, you will notice that the ATP/ADP ratio drops with increasing cell number. This is due to glucose depletion for the media. Less cells will give you a better charged cell.
3) Following on from point 2, change the media 1-2 hours before starting your experiment. This will replenish the glucose in the media. We found this to be very important when trying to utilise physiological glucose concentrations (2.5-10 mM glucose).

Reliable kit and easy to use.

DR. Craig Beall

Verified customer

Submitted Sep 17 2014

Question

I would like to use ab65313 kit on drug treated cells next week.

I am requested to represent my results per mg protein.

Does the buffer in which i lyes the cells (the nucleotide releasing buffer) allow performing a bradford assay (I use a sigma kit) after the luminescence reading?

Otherwise, would it allow reading hoechst fluorescence after the fact?

Please advise,





Read More

Abcam community

Verified customer

Asked on Dec 29 2015

Answer

The assay buffer has high concentration of detergent-hence Bradford assay is not recommended. We advise using the BCA assay for protein measurement when using this kit.
This assay is performed on cell lysates, so it would be difficult to label the cells with Hoechst after the assay.

Read More

Sam Washer

Abcam Scientific Support

Answered on Dec 29 2015

Question

Why is data C required after data B acquisition?
Why is it required to wait a few hours in step 3?
What is the best sample preparation protocol for pancreatic islet cells?

Read More

Abcam community

Verified customer

Asked on Nov 14 2014

Answer

Thank you for contacting us. Regarding your questions from yesterday:

1. Why is data C required after data B acquisition?

The two readings should be similar and perhaps data C is not necessary but the ATP signal may go down slightly while the ADP converting enzyme is prepared.

2. Why wait a few hours after adding the ATP Monitoring Enzyme and Nucleotide Releasing Buffer to the wells and before reading data A?

There may be bacterial contamination in the well or reagents that will produce some ATP signal, which should disappear after a few hours. Again, the wait may not be necessary if there is no contamination. (The note in step 3 about waiting a few hours should probably be at the start of step 3. There is no need to wait a few hours AFTER reading data A).

3. What is the best sample preparation protocol for pancreatic islet cells?

I recommend incubating each islet sample with 50ul of the Nucleotide Releasing Buffer, incubating for 5 minutes at room temperature with gentle shaking, and then adding this mixture to the ATP Monitoring Enzyme and Nucleotide Releasing Buffer mixture in the assay plate.

Read More

Tom Ruyle

Abcam Scientific Support

Answered on Nov 14 2014

Question

The protocol for this kit states to read the sample in a luminometer after about 2 minutes. Is it possible to wait longer (eg. 10 minutes), or will the signal fade quickly?

Read More

Abcam community

Verified customer

Asked on Aug 04 2014

Answer

The signal should be fairly stable as long as the sample is protected from light. But we suggest measuring after 2 mins to make sure that no light is quenched. Waiting for 10 mins is not advisable since some light can be quenched during this time. For the specific samples in question, if there is no significant light by 2 mins, then most likely more sample should be used in the well.

Read More

Jeremy Kasanov

Abcam Scientific Support

Answered on Aug 04 2014

Question

We are planning
to purchase your KIT #ab65313 in order to estimate ATP/ADP ratio in
tissue samples.
I have looked at the description of the assay at your web page
(https://www.abcam.com/ps/products/65/ab65313/documents/ab65313%20ADP%20ATP%20Ratio%20Assay%20Kit%20Bioluminescent%20v2%20%28website%29.pdf)
but samples preparation is described here only for cells. I would
greatly appreciate it if you provide me instruction for tissue samples
preparation for this assay.

Read More

Abcam community

Verified customer

Asked on Jul 30 2014

Answer


We can recommend two protocols:
1. Prepare a single cell suspension from the tissues of interest with any method desired (cells have to stay intact). Then treat the cells as instructed in the protocols section for adherent cells: Remove culture medium/ buffer and treat cells with 50 μl of Nucleotide Releasing Buffer for 5 minutes at room temperature with gentle shaking. Transfer into luminometer plate. (page 8, 4b).
2. Chop the tissues to the finest possible size (keep cool). Then treat cells with 50 μl of Nucleotide Releasing Buffer for 5 minutes at room temperature with gentle shaking. Transfer into luminometer plate.
Please note that the tissue preparation needs to be optimised by the end user since an increased number of dead cells may affect the results.

Read More

Anja Hoffmann

Abcam Scientific Support

Answered on Jul 30 2014

Question

I would like to investigate the ratio of ATP:ADP in nerve tissue, namely DRG and saphenous nerve. Could you please advise whether this kit would be suitable and if extra steps are required?

Read More

Abcam community

Verified customer

Asked on Jan 03 2014

Answer



Please see the following publication which may be of use to you:

Diguet N et al. Muscle creatine kinase deficiency triggers both actin depolymerization and desmin disorganization by advanced glycation end products in dilated cardiomyopathy. J Biol Chem 286:35007-19 (2011).

Protocol:
ATP and ADP/ATP ratios were measured using the bioluminescent ADP/ATP ratio assay kit from Abcam as indicated by the manufacturer with minor modifications. Briefly, cardiac tissue was frozen in liquid nitrogen and powdered with a mortar. Tissue powder was suspended in the provided lysis buffer (10 μl/mg of tissue powder) for 5 min exactly at room temperature, followed by centrifugation at 10,000 × g for 1 min to pellet insoluble material. We used 50 μl of supernatant per assay in a final volume of 100 μl. Luciferase units were converted to picomoles of the amount of ATP by plotting against a standard curve with serial dilution of ATP. Data were normalized by the amount of protein present in the supernatant as measured by the Bradford assay.

Read More

Elisa Thomas

Abcam Scientific Support

Answered on Jan 03 2014

Question

ADP/ATP Ratio Assay Kit (Bioluminescent) (ab65313)
Hello. I would like to know if I can ue your kit to measure ATP
content in mouse liver.
I need to measure heaptic ATP, ADP, and AMP. Any suggestion?
Thank you,

Read More

Abcam community

Verified customer

Asked on Mar 04 2013

Answer

Thank you for your inquiry.

I am happy to confirm that ab65313 can be used with mouse liver tissue samples.

This kit can be used to measure the ADP/ATP Ratio, AMP is not measured.

If you are planning to use this kit with tissue samples, I would like to note that the preparation of the samples need to be optimized according to the sample type etc..

A single cell suspension has to be prepared from the tissue and this can then be treated as the adherent cells in the prototocol.

Another possibility is to cut the tissue in very small pieces and then incubate for 5 minutes at room temperature in Nucleotide releasing buffer.

I hope this information is helpful and wish you good luck for your results.

Read More

Abcam Scientific Support

Answered on Mar 04 2013

Question

I see that the readout would be ATP in the media; this means that non-viable (or dead) cells would have lower ATP readings in the media compared to live cells.

Read More

Abcam community

Verified customer

Asked on Feb 20 2013

Answer


Yes, you are correct. Lower ATP readings would indicate a decrease in cell viability.

Read More

Abcam Scientific Support

Answered on Feb 20 2013

Question

Can you please suggest a product for which I can test cell viability using cell supernatants?

Read More

Abcam community

Verified customer

Asked on Feb 20 2013

Answer

Ab65314 is recommended for cell viability. For media, use the same protocol as mentioned for the suspension cells in step 2.a.

Read More

Abcam Scientific Support

Answered on Feb 20 2013

1-10 of 41 Abreviews or Q&A

  •  Previous
  • 1
  • 2
  • 3
  • 4
  • 5
  • Next 

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
For licensing inquiries, please contact partnerships@abcam.com

Get resources and offers direct to your inbox Sign up
A-Z by research area
  • Cancer
  • Cardiovascular
  • Cell biology
  • Developmental biology
  • Epigenetics & Nuclear signaling
  • Immunology
  • Metabolism
  • Microbiology
  • Neuroscience
  • Signal transduction
  • Stem cells
A-Z by product type
  • Primary antibodies
  • Secondary antibodies
  • Biochemicals
  • Isotype controls
  • Flow cytometry multi-color selector
  • Kits
  • Loading controls
  • Lysates
  • Peptides
  • Proteins
  • Slides
  • Tags and cell markers
  • Tools & Reagents
Help & support
  • Support
  • Make an Inquiry
  • Protocols & troubleshooting
  • Placing an order
  • RabMAb products
  • Biochemical product FAQs
  • Training
  • Browse by Target
Company
  • Corporate site
  • Investor relations
  • Company news
  • Careers
  • About us
  • Blog
Events
  • Tradeshows
  • Conferences
International websites
  • abcam.cn
  • abcam.co.jp

Join with us

  • LinkedIn
  • facebook
  • Twitter
  • YouTube
  • Terms of sale
  • Website terms of use
  • Cookie policy
  • Privacy policy
  • Legal
  • Modern slavery statement
© 1998-2022 Abcam plc. All rights reserved.