Protocol detail

Immunofluorescence Staining and Imaging

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Immunofluorescence staining was performed as reported [14 (link)]. Briefly, the cultured RAVSMCs and p-HAVSMCs were stained with Ki67 (ab16667, Abcam) antibody. Secondary antibody was incubated for 60 min and followed with DAPI staining. Images were taken by Olympus light microscope BX53 system. RAVSMCs infected with lenti-mCherry-GFP-LC3 were fixed with 4% paraformaldehyde for 15 min in darkness. After that, cells were taken to capture image by Olympus light microscope BX53 system directly.

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Chen T.Q., Guo X., Huo B., Zhong X.X., Wang Q.H., Chen Y., Zhu X.H., Feng G.K., Jiang D.S., Fang Z.M, & Wei X. (2023). BRD4770 inhibits vascular smooth muscle cell proliferation via SUV39H2, but not EHMT2 to protect against neointima formation. Human Cell, 36(5), 1672-1688.

Publication 2023

light microscope BX53 system

Antibody Cells Dapi Darkness Immunofluorescence Microscope light Paraformaldehyde

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Variable analysis

independent variables

Infection of RAVSMCs with lenti-mCherry-GFP-LC3

dependent variables

Ki67 expression in RAVSMCs and p-HAVSMCs
Autophagy in RAVSMCs infected with lenti-mCherry-GFP-LC3 (as indicated by mCherry-GFP-LC3 signal)

control variables

Cultured RAVSMCs
Cultured p-HAVSMCs

controls

No positive or negative controls were explicitly mentioned in the provided information.

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