Understand the mechanism of mitochondrial toxicity using Cyprotex’s mitochondrial respiratory complex assay using permeabilized cells (Seahorse XFe96).
Cyprotex’s mitochondrial respiratory complex assay utilising permeabilised cells on the Seahorse XFe analyzer is in Cyprotex’s portfolio of in vitro toxicology services for measuring potential mitochondrial toxicity. Cyprotex deliver consistent, high quality data with the flexibility to adapt protocols based on specific customer requirements.
Drug-induced mitochondrial toxicity is rapidly gaining recognition within the pharmaceutical industry as a contributor to compound attrition and post-market drug withdrawals.
1 Nadanaciva S and Will Y (2011) Current Pharmaceutical Design 17; 2100-2112
|Cell Type||HepG2 (others available on request)|
|Analysis Platform||Seahorse XFe96 flux analyser
|Analysis Method||Use of solid state fluorescent sensors to measure oxygen consumption rate (OCR)|
|Test Article Requirements||50 µL of a DMSO stock solution to achieve 100x Cmax (200x top concentration to maintain 0.5% DMSO) or equivalent amount in solid compound|
|Test Article Concentration*||7 point dose response curve with top concentration based on 100x Cmax orsolubility limit|
|Number of Replicates*||3 replicates per concentration|
|Quality Controls*||Negative control: 0.5% DMSO (vehicle)
Positive control: Assay appropriate control
|Data Delivery||Minimum effective concentration (MEC) and AC50 values with dose response curves for each measured parameter|
|Glucose/galactose mitochondrial toxicity assay
HCS based mitochondrial toxicity assay
Functional mitochondrial toxicity assay (Seahorse XFe96)
Known mitochondrial toxicants and non-toxicants were screened in the mitochondrial respiratory complex assay. The identified mechanisms of action were compared to those published in the literature.
|Pyruvate Respiration||Succinate Respiration||Ascorbate Respiration|
|Compound||Mechanism||MEC (µM)||AC50 (µM)||MEC (µM)||AC50 (µM)||MEC (µM)||AC50 (µM)|
|Rotenone||Complex I inhibitor||0.006||0.033||No response||No response||No response||No response|
|Ketoconazole||Complex I inhibitor||13.3||42.1||No response||No response||No response||No response|
|Carboxine||Complex II inhibitor||No response||No response||2.43||23.3||No response||No response|
|Thenoyltrifluoro-acetone (TTFA)||Complex II inhibitor||605||>2000||45.6||151||688||1810|
|Antimycin A||Complex III inhibitor||0.019||0.027||0.012||0.019||No response||No response|
|Sodium azide||Complex IV inhibitor||177||654||79.1||411||23.5||156|
|Streptomycin||No effect||No response||No response||No response||No response||No response||No response|
|Betaine||No effect||No response||No response||No response||No response||No response||No response|
1 Nadanaciva S and Will Y (2011) New insights in drug-induced mitochondrial toxicity. Current Pharmaceutical Design 17; 2100-2112
Learn more about toxicology in our popular Mechanisms of Drug-Induced Toxicity guide