Immunofluorescence Staining of DNA Damage

Immunofluorescence staining was performed as described previously (6 (link)). Suspended cells were fixed onto glass using 1% paraformaldehyde and ethanol (70%) at the treatment endpoint. The cells were incubated with primary antibodies against γH2AX (1:500, CST 2577), RAD51 (1:500, Abcam ab133534), RPA (1:200, Abcam ab2175), and p-CHK1 (1:200, Ser317, CST 12302), then incubated with secondary antibody. The cells were visualized using a Zeiss fluorescence microscope.

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Bian X., Wang X., Zhang Q., Ma L., Cao G., Xu A., Han J., Huang J, & Lin W. (2020). The MYC Paralog-PARP1 Axis as a Potential Therapeutic Target in MYC Paralog-Activated Small Cell Lung Cancer. Frontiers in Oncology, 10, 565820.

Publication 2020

ab133534

Antibodies Antibody Cells Ethanol Fluorescence microscope Immunofluorescence Paraformaldehyde

Corresponding Organization : High Magnetic Field Laboratory

Other organizations : University of Science and Technology of China, Anhui Provincial Hospital, Zhejiang University

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Variable analysis

independent variables

Treatment endpoint

dependent variables

Expression of γH2AX
Expression of RAD51
Expression of RPA
Expression of p-CHK1

control variables

Suspended cells
Glass slide fixation with 1% paraformaldehyde and 70% ethanol
Primary antibody concentrations (1:500 for γH2AX and RAD51, 1:200 for RPA and p-CHK1)
Visualization using Zeiss fluorescence microscope

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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