Quantification of Cellular Reactive Oxygen Species

ROS was quantified using 2′,7′ –dichlorofluorescin diacetate (DCFDA) fluorescence (Sigma-Aldrich, D6883, St Louis, MO, USA) normalized to cell number [14 (link),17 (link)]. Twenty-five mM glucose (Sigma Aldrich, D8270) and 50 µM tert-butyl hydrogen peroxide (TBHP) (Sigma Aldrich, 416665) were used as positive controls. The free radical scavenger Tempol (50 µM; Sigma Aldrich SML0737) was used to manipulate ROS generation and served as a control [14 (link)].

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McCambridge G., Agrawal M., Keady A., Kern P.A., Hasturk H., Nikolajczyk B.S, & Bharath L.P. (2019). Saturated Fatty Acid Activates T Cell Inflammation Through a Nicotinamide Nucleotide Transhydrogenase (NNT)-Dependent Mechanism. Biomolecules, 9(2), 79.

Publication 2019

tert-butyl hydrogen peroxide Tempol

Dichlorofluorescin Fluorescence Free radical scavenger Glucose Tempol Tert butyl hydrogen peroxide

Corresponding Organization : University of Kentucky

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Variable analysis

independent variables

25 mM glucose (Sigma Aldrich, D8270)
50 µM tert-butyl hydrogen peroxide (TBHP) (Sigma Aldrich, 416665)
50 µM Tempol (Sigma Aldrich SML0737)

dependent variables

ROS quantified using 2′,7′ –dichlorofluorescin diacetate (DCFDA) fluorescence (Sigma-Aldrich, D6883, St Louis, MO, USA) normalized to cell number

control variables

Cell number

positive controls

25 mM glucose (Sigma Aldrich, D8270)
50 µM tert-butyl hydrogen peroxide (TBHP) (Sigma Aldrich, 416665)

negative control

50 µM Tempol (Sigma Aldrich SML0737)

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