EnzyLight™ ADP/ATP Ratio Assay Kit

Protocol SDS

Application

• For quantitative bioluminescent assay for ADP: ATP ratio (apoptosis) in cells and screen for modulators.

Key Features

• Safe. Non-radioactive assay.
• Homogeneous and convenient. “Mix-incubate-measure” type assay. No wash and reagent transfer steps are involved.
• Robust and amenable to HTS: Z factors of 0.5 and above are routinely observed in 96-well and 384-well plates. Can be readily automated on HTS liquid handling systems for processing thousands of samples per day.

Method

• Luminescence

Samples

• Cells etc

Species

• All

Procedure

• 20 min

Size

• 100 tests

Shelf Life

• 12 months

More Details

Changes in the ADP/ATP ratio have been used to differentiate modes of cell death and viability. Increased levels of ATP and decreased levels of ADP signify proliferating cells. Conversely, decreased levels of ATP and increased levels of ADP represent apoptotic or necrotic cells where the decrease in ATP and increase in ADP is much more pronounced in necrosis versus apoptosis. BioAssay Systems’ EnzyLight™ ADP/ATP Ratio Assay Kit provides a rapid method to measure ADP and ATP levels for the screening of apoptosis, necrosis, and cell proliferation in mammalian cells. The assay involves two steps. In the first step, the working reagent lyses cells to release ATP and ADP. In the presence of luciferase, ATP immediately reacts with the Substrate D-luciferin to produce light. The light intensity is a direct measure of intracellular ATP concentration. In the second step, the ADP is converted to ATP through an enzyme reaction. This newly formed ATP then reacts with the D-luciferin as in the first step. This non-radioactive, homogeneous cell-based assay is performed in microplates. The reagent is compatible with all culture media and with all liquid handling systems for high-throughput screening applications in 96-well and 384-well plates.

We would like to measure ADP, ATP and ADP/ATP Ratio in serum or plasma. This kit seems to be suitable for cell samples, but is it compatible with serum or plasma?

The ADP/ATP ratio assay was developed as a cell based assays and is useful to follow changes in the ADP/ATP ratio as a marker for viability in cells in the course of a drug treatment or during culture. The assay also works with serum samples but because it is not quantitative this may limit its usefulness for serum analysis. Depending on the exact nature of your experiment, our EATP-100 and EADP-100 assay kits may be the better choice.

Fang, Y., et al (2020). Melatonin inhibits formation of mitochondrial permeability transition pores and improves oxidative phosphorylation of frozen-thawed ram sperm. Frontiers in Endocrinology, 10. Assay: ATP/ADP in ram semen.

Zhu, J., et al (2020). Altered energy metabolism during early optic nerve crush injury: Implications of warburg-like aerobic glycolysis in facilitating retinal ganglion cell survival. Neuroscience Bulletin, 36(7), 761-777. Assay: ATP/ADP in mouse tissue.

Yuan, G., et al. (2020). RGS12 is required for the maintenance of mitochondrial function during skeletal development. Cell Discovery, 6(1) Assay: ATP/ADP in mouse cells.

Hong, Z., et al (2020). Upregulation of DPY30 promotes cell proliferation and predicts a poor prognosis in cholangiocarcinoma. Biomedicine & Pharmacotherapy, 123, 109766. Assay: ATP/ADP in human cancer cells.

Fan, X., et al (2020). Genome reduction enhances production of polyhydroxyalkanoate and alginate oligosaccharide in pseudomonas mendocina. International Journal of Biological Macromolecules, 163, 2023-2031 Assay: ATP/ADP in bacteria cells.

Jayawardhane, J., et al (2020). Roles for plant mitochondrial alternative oxidase under Normoxia, hypoxia, and Reoxygenation conditions. Frontiers in Plant Science, 11. Assay: ATP/ADP in plant tissue.

Gordillo, G. M., et al (2021). Mitochondria as target for tumor management of Hemangioendothelioma. Antioxidants & Redox Signaling, 34(2), 137-153. Assay: ATP/ADP in mouse cells.

Cao, D., et al (2019). ACE overexpression in myeloid cells increases oxidative metabolism and cellular ATP. Journal of Biological Chemistry, 295(5), 1369-1384. Assay: ATP/ADP in mouse white blood cells.

Le, Q. G., et al (2021). The ADP-binding kinase region of Ire1 directly contributes to its responsiveness to endoplasmic Reticulum stress. Scientific Reports, 11(1). Assay: ATP/ADP in yeast cells.

Zafari, S., et al (2020). Transcriptional and metabolic changes associated with Phytoglobin expression during germination of Barley seeds. International Journal of Molecular Sciences, 21(8), 2796. Assay: ATP/ADP in plant tissue.

Dae-Ho So (2018). The role of SIRT1 in avoiding AIM2-mediated antiviral defense in cervical cancer (Doctoral dissertation). Assay: ADP/ATP ratio in human cells.

Dyczynski, M., Vesterlund, M., Bjorklund, A. C., Zachariadis, V., Janssen, J., Gallart-Ayala, H. & Tamm, K. P. (2018). Metabolic reprogramming of acute lymphoblastic leukemia cells in response to glucocorticoid treatment. Cell death & disease, 9(9), 846. Assay: ADP/ATP ratio in human cells.

Flood, J. J., & Copley, S. D. (2018). Genome-Wide Analysis of Transcriptional Changes and Genes That Contribute to Fitness during Degradation of the Anthropogenic Pollutant Pentachlorophenol by Sphingobium chlorophenolicum. mSystems, 3(6), e00275-18. Assay: ADP/ATP ratio in S. chlorophenolicum cells.

Ghatak, P. D., Mathew-Steiner, S. S., Pandey, P., Roy, S., & Sen, C. K. (2018). A surfactant polymer dressing potentiates antimicrobial efficacy in biofilm disruption. Scientific reports, 8(1), 873. Assay: ADP/ATP ratio in bacterial cells.

Jian, Z., Cheng, T., Zhang, Z., Raulefs, S., Shi, K., Steiger, K. & Bruns, P. (2018). Glycemic variability promotes both local invasion and metastatic colonization by pancreatic ductal adenocarcinoma. Cellular and Molecular Gastroenterology and Hepatology, 6(4), 429-449. Assay: ADP/ATP ratio in murine cells.

Kleme, M., Sane, A., Garofalo, C., Seidman, E., Brochiero, E., Berthiaume, Y., & Levy, E. (2018). CFTR Deletion Confers Mitochondrial Dysfunction and Disrupts Lipid Homeostasis in Intestinal Epithelial Cells. Nutrients, 10(7), 836. Assay: ADP/ATP ratio in human cells.

Dominguez Sanchez, J. M. (2017). Role of CLK3 in HIF1alpha driven cardiac hypertrophy (Doctoral dissertation, ETH Zurich). Assay: ADP/ATP ratio in mouse cells.

Jian, Z. (2017). Dominant oncogenic signal-mediated nutritional dependency affects malignant behaviours of pancreatic cancer (Doctoral dissertation, Technische Universitat Munchen). Assay: ADP/ATP ratio in murine cells.

Morten, K. J., Potter, M., Badder, L., Sivathondan, P., Dragovic, R., Neumann, A. & Lodge, T. A. (2017). Insights into pancreatic beta cell energy metabolism using rodent beta cell models. Wellcome Open Research, 2. Assay: ADP/ATP ratio in rodent cells.

Nadeau, L. (2017). Investigating IL-15 Metabolic Impact and its Mechanism of Action in Skeletal Muscle Cells (Doctoral dissertation, Universite d’Ottawa/University of Ottawa). Assay: ADP/ATP ratio in rat cells.

Rajamani, U., Gross, A. R., Ocampo, C., Andres, A. M., Gottlieb, R. A., & Sareen, D. (2017). Endocrine disruptors induce perturbations in endoplasmic reticulum and mitochondria of human pluripotent stem cell derivatives. Nature communications, 8(1), 219. Assay: ADP/ATP ratio in human cells.

Villa-Bellosta, R., Hamczyk, M. R., & Andres, V. (2017). Novel phosphate-activated macrophages prevent ectopic calcification by increasing extracellular ATP and pyrophosphate. PloS one, 12(3), e0174998. Assay: ADP/ATP ratio in mice cells.

Amantini, C., Morelli, M. B., Nabissi, M., Cardinali, C., Santoni, M., Gismondi, A., & Santoni, G. (2016). Capsaicin triggers autophagic cell survival which drives epithelial mesenchymal transition and chemoresistance in bladder cancer cells in an Hedgehog-dependent manner. Oncotarget, 7(31), 50180. Assay: ADP/ATP ratio in human cells.

Lezi, E., & Swerdlow, R. H. (2016). Lactate’s effect on human neuroblastoma cell bioenergetic fluxes. Biochemical pharmacology, 99, 88-100. Assay: ADP/ATP ratio in human cells.

Liu, L., Wang, Y., Bai, R., Yang, K., & Tian, Z. (2016). MiR-186 inhibited aerobic glycolysis in gastric cancer via HIF-1alpha regulation. Oncogenesis, 5(5), e224. Assay: ADP/ATP ratio in human cells.

Schilf, P. (2016). Interplay of mtDNA, metabolism and microbiota in the pathogenesis of AIBD (Doctoral dissertation, Universitat zu Lubeck). Assay: ADP/ATP ratio in mice cells.

Son, S. W., Kim, S. H., Moon, E. Y., Kim, D. H., Pyo, S., & Um, S. H. (2016). Prognostic significance and function of the vacuolar H+-ATPase subunit V1E1 in esophageal squamous cell carcinoma. Oncotarget, 7(31), 49334. Assay: ADP/ATP ratio in human cells.

Taneike, M., Nishida, K., Omiya, S., Zarrinpashneh, E., Misaka, T., Kitazume-Taneike, R. & Shah, A. M. (2016). mTOR hyperactivation by ablation of tuberous sclerosis complex 2 in the mouse heart induces cardiac dysfunction with the increased number of small mitochondria mediated through the down-regulation of autophagy. PLoS One, 11(3), e0152628. Assay: ADP/ATP ratio in mice tissues.

Chen, R. & Jeong, SS (2013). Apyrase therapy for bleeding conditions. WO/2011/088231. Assay: ADP/ATP ratio in human cell.

Kwon DH et al (2012).Dietary protein restriction induces steatohepatitis and alters leptin/signal transducers and activators of transcription 3 signaling in lactating rats. J Nutr Biochem 23(7):791-9. Assay: ADP/ATP ratio in human cell.

Chen, R. and Jeong, S. (2011). Pyrase Therapy For Bleeding Conditions. WO/2011/088231. Assay: ADP/ATP ratio in rat bronchoalveolar fluid.

Choi EM (2011). Glabridin protects osteoblastic MC3T3-E1 cells against antimycin A induced cytotoxicity. Chem Biol Interact. 193(1):71-8. Assay: ADP/ATP ratio in mouse cell line.

Choi EM (2011). Luteolin protects osteoblastic MC3T3-E1 cells from antimycin A-induced cytotoxicity through the improved mitochondrial function and activation of PI3K/Akt/CREB. Toxicol In Vitro. 25(8):1671-9. Assay: ADP/ATP ratio in mouse cell line.

Choi EM (2011). Luteolin protects osteoblastic MC3T3-E1 cells from antimycin A-induced cytotoxicity through the improved mitochondrial function and activation of PI3K/Akt/CREB. Toxicol In Vitro. Assay: ADP/ATP ratio in rat bronchoalveolar fluid.

Narain, NR, Mccook, JP, Sarangarajan, R. et al (2011). Methods for treatment of metabolic disorders using epimetabolic shifters, multidimensional intracellular molecules, or environmental influencers. US Patent Appl. 20110020312. Assay: ADP/ATP ratio in human cell, environmental.

Rink, C et al (2011). Oxygen-inducible glutamate oxaloacetate transaminase as protective switch transforming neurotoxic glutamate to metabolic fuel during acute ischemic stroke. Antioxid Redox Signal 14(10):1777-85. Assay: ADP/ATP ratio in mouse cell line.

Sarangarajan, R. (2011). Methods for treatment of a sarcoma using an epimetabolic shifter (Coenzyme q10). US 2011/0064747. Assay: ADP/ATP ratio in human cell.

Bekeredjian R, Suhr ST et al (2010). Conditional HIF-1alpha expression produces a reversible cardiomyopathy. PLoS One 5(7):e11693. Assay: ADP/ATP ratio in human cell line.

Bekeredjian R. et al. (2010). Conditional HIF-1alpha expression produces a reversible cardiomyopathy. PLoS One 5(7):e11693. Assay: ADP/ATP ratio in mouse heart lysate.

Chandak PG et al (2010). Efficient phagocytosis requires triacylglycerol hydrolysis by adipose triglyceride lipase. J Biol Chem. 285(26):20192-201. Assay: ADP/ATP ratio in rat total ATP/ADP in mitochondria.

Chandak PG, Suhr ST, et al. (2010). Efficient phagocytosis requires triacylglycerol hydrolysis by adipose triglyceride lipase. J Biol Chem. 285(26):20192-201. Assay: ADP/ATP ratio in mouse cells.

Saito A, Castilho RF (2010). Inhibitory effects of adenine nucleotides on brain mitochondrial permeability transition. Neurochem Res. 35(11):1667-74. Assay: ADP/ATP ratio in mouse heart lysate.

Suhr ST et al (2010). Mitochondrial rejuvenation after induced pluripotency. PLoS One 5(11):e14095. Assay: ADP/ATP ratio in rat brain tissue.

Suhr ST, et al. (2010). Mitochondrial rejuvenation after induced pluripotency. PLoS One 5(11):e14095. Assay: ADP/ATP ratio in human cell line.

Schafer ZT, et al (2009). Antioxidant and oncogene rescue of metabolic defects caused by loss of matrix attachment. Nature. 461(7260):109-113. Assay: ADP/ATP ratio in human cell.

Sugimoto S et al (2009). Apyrase treatment prevents ischemia-reperfusion injury in rat lung isografts. J Thorac Cardiovasc Surg.138(3):752-9. Assay: ADP/ATP ratio in mouse cells.

Sugimoto S, et al. (2009). Apyrase treatment prevents ischemia-reperfusion injury in rat lung isografts. J Thorac Cardiovasc Surg.138(3):752-9. Assay: ADP/ATP ratio in rat lung tissue.

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If you or your labs do not have the equipment or scientists necessary to run this assay, BioAssay Systems can perform the service for you.

– Fast turnaround
– Quality data
– Low cost

Please email or call 1-510-782-9988 x 2 to discuss your projects.