Ab79849

HGFs cells were planted on the plate and treated with drugs. The cells were fixed with 4% paraformaldehyde, and permeabilized with Triton X-100. PBS was rinsed and sealed with 5% BSA. Then anti-HSP90α (abcam, ab79849, 1:100), anti-HSP90β (abcam, ab53497, 1:100) antibody were added and incubated at 37 °C for 3 h. After washing with PBS, the secondary antibody IgG H & L (abcam, ab150080, 1:500) was incubated for 30 min. DAPI was added dropwise to dye the nucleus under dark conditions, and sealed slices, then the collected image was observed under the fluorescence microscope.

HSP90AA1 (Heat Shock Protein 90 Alpha Family Class A Member 1) protein levels were analyzed in tumor and peritumor using standard immunohistochemical methods. Briefly, slides were coated with primary antibody against HSP90AA1 (mouse ID:ab79849, Abcam) and incubated overnight at 4 °C, then incubated with secondary antibody for 30 min at room temperature. The slides were stained with DAB solution for 10 min, and the intensity of HSP90AA1 staining was scored, shown below. 0, 1, 2, 3 representing negative, weak, moderate, and strong; The evaluation of the tumor cell positivity rate was as follows. 1, 2, 3, 4 representing 0-25%, 26-50%, 51-75% and 76-100%. The staining intensity was multiplied by the positive rate score to obtain the IHC score. Two pathologists reviewed sections.

Antibodies against pro–IL-1β (12242) and cleaved IL-1β (83186) were purchased from Cell Signaling Technology. Antibodies against tubulin (AG-27B-0005-C100), caspase-1 (AG-20B-0048-C100), and NLRP3 (AG-20B-0014) were purchased from Adipogen. Antibodies against gasdermin-D (ab210070), HSP90β (ab53497), and HSP90α (ab79849) were purchased from Abcam. Anti-SGT1 (612104) was purchased from BD Biosciences. Anti–vesicular stomatitis virus glycoprotein (VSVg) (V4888) was purchased from Sigma-Aldrich. Anti-ASC (sc-22514) was purchased from Santa Cruz Biotechnology. Anti-mouse–horseradish peroxidase (HRP) (115-035-146), anti-rabbit–HRP (111-035-144), and anti-human–HRP (709-035-149) were purchased from the Jackson Laboratory. Anti-rabbit–Alexa Fluor 647 was purchased from Thermo Fisher Scientific.