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

    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 assaycytotoxicity 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

  • 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.

Images

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

  • 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).

     

  • 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

References

This product has been referenced in:

  • Korski KI  et al. Hypoxia Prevents Mitochondrial Dysfunction and Senescence in Human c-Kit+ Cardiac Progenitor Cells. Stem Cells 37:555-567 (2019). Read more (PubMed: 30629785) »
  • Whang YM  et al. Rapamycin enhances growth inhibition on urothelial carcinoma cells through LKB1 deficiency-mediated mitochondrial dysregulation. J Cell Physiol 234:13083-13096 (2019). Read more (PubMed: 30549029) »
See all 51 Publications for this product

Customer reviews and Q&As

Filter by Ratings

1-2 of 2 Abreviews

ADP/ATP Ratio Assay in M. tuberculosis

Excellent Excellent 5/5 (Ease of Use)
Abreviews
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
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

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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