Intracellular ROS Measurement Protocol

The DCFH-DA probe (S0033S, Beyotime Biotechnology, China) was used to examine the intracellular ROS level according to the manufacturer’s instructions and our published protocol (Zhang et al., 2020 (link)). In brief, NRCMs were incubated with DCFH-DA for 2 h at 37°C, and then the cells were washed with PBS three times in the dark. Photos were taken using an Olympus IX53 fluorescence microscope. Image Pro-Plus (version 6.0) was used to analyze images.

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Zhang N., Liao H.H., Feng H., Mou S.Q., Li W.J., Aiyasiding X., Lin Z., Ding W., Zhou Z.Y., Yan H., Chen S, & Tang Q.Z. (2021). Knockout of AMPKα2 Blocked the Protection of Sestrin2 Overexpression Against Cardiac Hypertrophy Induced by Pressure Overload. Frontiers in Pharmacology, 12, 716884.

Publication 2021

DCFH-DA probe IX53 fluorescence microscope

Cells Dcfh da Fluorescence microscope Intracellular

Corresponding Organization :

Other organizations : Wuhan University, Renmin Hospital of Wuhan University

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Variable analysis

independent variables

DCFH-DA probe

dependent variables

Intracellular ROS level

control variables

Incubation time (2 h)
Incubation temperature (37°C)
Washing with PBS (3 times)
Imaging with Olympus IX53 fluorescence microscope
Image analysis using Image Pro-Plus (version 6.0)

controls

No positive or negative controls were explicitly mentioned in the input protocol.

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