@article{45f93c85665f4b2b9594f00b930f4dc0,
title = "Development of a 96-well based assay for kinetic determination of catalase enzymatic-activity in biological samples",
abstract = "Oxidative stress biomarkers are powerful endpoints in toxicological research. Cellular reductive/oxidative balance affects numerous signaling pathways involving H2O2. Detoxification and control of H2O2 levels results mainly from catalase activity. The aim of this work was to develop a precise, simple, cost-effective microassay to measure catalase activity in small tissue samples and cell extracts. We developed a protocol that quantifies H2O2 decomposition by intrinsic catalase in biological samples. Catalase activity was calculated based on rate of decomposition of H2O2, following absorbance at 240 nm. We developed a multi-well spectroscopic approach, reducing sample quantity requirements and allowing simultaneous assessment of large number of samples. The protocol is sensitive across a wide range of catalase activity (11.5–7575 U). The assay presents a 95% confidence interval with an intra-assay coefficient of variation of 3.7%, an inter-assay coefficient of variation of 6.2% and good correlation with a commercial kit. The assay was established and validated for different biological samples, including sheep hepatic tissue and human tumor and non-tumor cell lines. This high-throughput method is robust, sensitive, time-saving and cost-effective, generating highly reproducible results with precision and good correlation with a commercial kit reinforcing the method's validity for research and toxicological applications.",
keywords = "Antioxidant defense, Catalase, Hydrogen peroxide, Oxidative stress, Toxicity screening",
author = "Grilo, {Lu{\'i}s F.} and Martins, {Jo{\~a}o D.} and Cavallaro, {Chiara H.} and Nathanielsz, {Peter W.} and Oliveira, {Paulo J.} and Pereira, {Susana P.}",
note = "Funding Information: Funded by ERDF funds through the Operational Programme Competitiveness Factors ? COMPETE2020 and national funds by Foundation for Science and Technology under FCT-Post-doctoral Fellowship (SPP, SFRH/BPD/116061/2016), project grant PTDC/DTP-DES/1082/2014(POCI-01-0145-FEDER-016657), PTDC/BTM-SAL/29297/2017-POCI-01-0145-FEDER-029297, PTDC/MED-FAR/29391/2017-POCI-01-0145-FEDER-029391, PTDC/BIA-MOL/28607/2017, POCI-01-0145-FEDER-028607, LFG was supported by project ?Summer Course in Interdisciplinary Research, Development and Innovation in Cellular and Molecular Metabolism? (15-20/7/245), funded by the Portuguese Foundation for Science and Technology (FCT) and Directorate General for Higher Education (DGES), strategic project UIDB/04539/2020, and NIH grant R01HD070096-01A1. The funding agencies had no role in study design, data collection and analysis, decision to publish, or preparation of this document. Funding Information: Funded by ERDF funds through the Operational Programme Competitiveness Factors – COMPETE2020 and national funds by Foundation for Science and Technology under FCT-Post-doctoral Fellowship ( SPP, SFRH/BPD/116061/2016 ), project grant PTDC/DTP-DES/1082/2014(POCI-01-0145-FEDER-016657) , PTDC/BTM-SAL/29297/2017-POCI-01-0145-FEDER-029297 , PTDC/MED-FAR/29391/2017-POCI-01-0145-FEDER-029391 , PTDC/BIA-MOL/28607/2017 , POCI-01-0145-FEDER-028607 , LFG was supported by project “Summer Course in Interdisciplinary Research, Development and Innovation in Cellular and Molecular Metabolism” (15-20/7/245), funded by the Portuguese Foundation for Science and Technology (FCT) and Directorate General for Higher Education (DGES), strategic project UIDB/04539/2020 , and NIH grant R01HD070096-01A1 . The funding agencies had no role in study design, data collection and analysis, decision to publish, or preparation of this document. Publisher Copyright: {\textcopyright} 2020 Elsevier Ltd",
year = "2020",
month = dec,
doi = "10.1016/j.tiv.2020.104996",
language = "English (US)",
volume = "69",
journal = "Toxicology in Vitro",
issn = "0887-2333",
publisher = "Elsevier Limited",
}