Immunofluorescence Staining for NLRP3 Inflammasome

Immunofluorescence staining method was used according to our previous study (Wang R. et al., 2017 (link)). Specific primary antibodies were double stained with frozen aortic root sections for macrophage accumulation analysis, including goat anti-NLRP3 (Abcam) and rat anti-F4-80 (macrophage marker; Abcam) antibodies. goat anti-NLRP3 (Abcam) and rabbit anti-cleaved caspase-1 antibodies (ImmunoWay, TX, United States) were used for NLRP3 inflammasome activation analysis. Antibodies were double stained with α1-nAchR and CT-XB (lipid raft marker; Abcam) for correlation analysis. The analysis was imaged under a laser scanning confocal microscope (Zeiss, Germany).

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Duan F., Zeng C., Liu S., Gong J., Hu J., Li H, & Tan H. (2021). α1-nAchR-Mediated Signaling Through Lipid Raft Is Required for Nicotine-Induced NLRP3 Inflammasome Activation and Nicotine-Accelerated Atherosclerosis. Frontiers in Cell and Developmental Biology, 9, 724699.

Publication 2021

goat anti-NLRP3 rat anti-F4-80 α1-nAchR CT-XB laser scanning confocal microscope

Activation analysis Anti antibodies Antibodies Aortic root Caspase 1 Confocal laser scanning microscope Frozen sections Goat Immunofluorescence method Inflammasome Lipid Macrophage Rabbit

Corresponding Organization : Third Affiliated Hospital of Sun Yat-sen University

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Variable analysis

independent variables

Immunofluorescence staining method
Specific primary antibodies used (goat anti-NLRP3, rat anti-F4-80, goat anti-NLRP3, and rabbit anti-cleaved caspase-1)
Antibodies double stained with α1-nAchR and CT-XB (lipid raft marker)

dependent variables

Macrophage accumulation analysis
NLRP3 inflammasome activation analysis
Correlation analysis between antibodies and α1-nAchR and CT-XB

control variables

Frozen aortic root sections

controls

Positive control: Not specified
Negative control: Not specified

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