Visualization and Quantification of ASC Specks in Macrophages

The WT alveolar macrophages and BMDMs were seeded on chamber slides. After LPS and poly(dA:dT) stimulation, cells were fixed with 4% paraformaldehyde and then incubated with polyclonal ASC antibody (ADI-905-173-100, Enzo Lifesciences) for 16 h and FITC goat anti-rabbit (IgG) secondary antibody (ab6717, Abcam (Cambridge, MA, USA)) for 1h followed by DAPI (P36962, ThermoFisher Scientific) staining [41 (link),42 (link),43 (link)]. The ASC specks were analyzed using a Zeiss LSM880 laser scanning confocal microscope and quantified using ImageJ software v1.52a (Bethesda, MD, USA). The graph in figure represents the quantification of percent of ASC speck-positive cells for each mouse.

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Cho S.J., Hong K.S., Jeong J.H., Lee M., Choi A.M., Stout-Delgado H.W, & Moon J.S. (2019). DROSHA-Dependent AIM2 Inflammasome Activation Contributes to Lung Inflammation during Idiopathic Pulmonary Fibrosis. Cells, 8(8), 938.

Publication 2019

ADI-905-173-100 ab6717 P36962

Alveolar macrophages Anti igg Antibody Dapi Fitc Goat Laser scanning microscope Mouse Paraformaldehyde Poly Rabbit

Corresponding Organization :

Other organizations : Cornell University, Presbyterian Hospital, New York Hospital Queens, NewYork–Presbyterian Hospital, Ewha Womans University, Soonchunhyang University

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Variable analysis

independent variables

LPS stimulation
Poly(dA:dT) stimulation

dependent variables

Percent of ASC speck-positive cells

control variables

WT alveolar macrophages and BMDMs were seeded on chamber slides
Cells were fixed with 4% paraformaldehyde
Cells were incubated with polyclonal ASC antibody for 16 h
Cells were incubated with FITC goat anti-rabbit (IgG) secondary antibody for 1h
Cells were stained with DAPI

controls

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

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