Armenian hamster igg isotype control

Liver paraffin sections from 12 male mice (3 animals per age group and genotype) were labelled. Antigen retrieval was done by microwaving the sections 4x 5 min in 0.01 M citrate buffer (pH = 6) with 0.05% Tween. All antibodies were diluted in 2% BSA in PBS. Endogenous peroxidase was quenched in 3% H₂O₂ in methanol, and endogenous biotin with Biotin-Blocking System (Agilent, Cat. No X0590). Sections were incubated overnight at 4°C with Armenian hamster anti-mouse CD54 (ICAM-1) (5 μg/ml; Invitrogen—Thermo Fisher Scientific, Cat. No MA5405) or Armenian hamster IgG isotype control (5 μg/ml; Invitrogen—Thermo Fisher Scientific, Cat. No 14-4888-81). After rinsing in PBS with 0.05% Tween, sections were incubated for 30 min at RT with biotin-streptavidin-conjugated goat anti-Armenian hamster IgG (H+L, eBioscience, Cat. No 13–411385; 1.25 μg/ml), washed and incubated for 30 min with Streptavidin peroxidase (Agilent, Cat. No P0397). A diaminobenzidine substrate chromogen system kit (BD Pharmingen, Cat. No 550880) was used to visualize a positive reaction. Sections were counter-stained with haematoxylin, and images taken in a Nikon Eclipse TE2000-U Inverted Microscope.

Liver paraffin sections from 12 male mice (3 animals per age group and genotype) were labelled. Antigen retrieval was done by microwaving the sections 4x 5 min in 0.01 M citrate buffer (pH = 6) with 0.05% Tween. All antibodies were diluted in 2% BSA in PBS. Endogenous peroxidase was quenched in 3% H₂O₂ in methanol, and endogenous biotin with Biotin-Blocking System (Agilent, Cat. No X0590). Sections were incubated overnight at 4°C with Armenian hamster anti-mouse CD54 (ICAM-1) (5 μg/ml; Invitrogen—Thermo Fisher Scientific, Cat. No MA5405) or Armenian hamster IgG isotype control (5 μg/ml; Invitrogen—Thermo Fisher Scientific, Cat. No 14-4888-81). After rinsing in PBS with 0.05% Tween, sections were incubated for 30 min at RT with biotin-streptavidin-conjugated goat anti-Armenian hamster IgG (H+L, eBioscience, Cat. No 13–411385; 1.25 μg/ml), washed and incubated for 30 min with Streptavidin peroxidase (Agilent, Cat. No P0397). A diaminobenzidine substrate chromogen system kit (BD Pharmingen, Cat. No 550880) was used to visualize a positive reaction. Sections were counter-stained with haematoxylin, and images taken in a Nikon Eclipse TE2000-U Inverted Microscope.

L-Cysteine (L-Cys), aminooxyacetic acid (AOAA, CBS inhibitor), DL-propargylglycine (PAG, CSE inhibitor), lipopolysaccharide (LPS), and MCC950 (NLRP3 inflammasome inhibitor) were purchased from Sigma-Aldrich (St. Louis, MO). Control liposomes and clodronate liposomes were purchased from FormuMax (Silicon Valley, CA). Deoxyribonuclease Ⅰ was purchased from Beijing Solarbio Science and Technology Co. (Beijing, China). 2-Deoxy-D-glucose was purchased from J&K Scientific (Beijing, China). Armenian Hamster IgG Isotype Control and IL-1 beta Monoclonal Antibody were purchased from Invitrogen Life Technology (Foster City, CA). Antibodies for βⅢ-tubulin, Nestin, and IL-1β were purchased from Abcam (Cambridge, United Kingdom). Antibodies for cleaved caspase-1 and NLRP3 were purchased from Cell Signaling Technology (Danvers, MA). Monoclonal mouse anti-β-actin was purchased from Zhongshan Golden Bridge Biotechnology (Beijing, China). The secondary antibodies were purchased from Abcam (Cambridge, United Kingdom), Invitrogen Life Technology (Foster City, CA), and Zhongshan Golden Bridge Biotechnology (Beijing, China). Mouse ELISA kits were purchased from Dakewe Biotech (Shenzhen, China) and Boster Biological Technology (Wuhan, China). Dulbecco’s modified Eagle’s medium (DMEM), 10% heat-inactivated FBS, and 1% penicillin–streptomycin solutions were purchased from Gibco (Foster City, CA).