IHC staining of paraffin-embedded human or mice tumor samples were conducted according to the manufacturer’s instructions. Samples were deparaffinized, rehydrated, subjected to antigen retrieval, and blocked with 3% hydrogen dioxide, followed by incubating with the primary antibodies recognizing human AIM2 (dilution 1:200; #ab93015, Abcam) or human Gli1 (dilution 1:200; #ab15179, Abcam) overnight at 4° C. Next day, the sections were incubated with secondary antibody and were visualized using a tissue staining kit (Zhongshan Biotechnology, Beijing, China). Staining intensity was classified as 0 (lack of staining), 1 (mild staining), 2 (moderate staining) or 3 (strong staining); staining percentage was designated as 1 (<25%), 2 (25%-50%), 3 (51%-75%), or4 (>75%). The final staining score was calculated by the multiple of color intensity and positive cell percentage, which ranged from 0 to 12. Scores 0-4 were described as none or low expression, while 5-12 as high expression.
Ab15179
Manufactured by Abcam
Sourced in China
Ab15179 is a laboratory equipment product offered by Abcam. It is designed for general laboratory use. No further details about its core function or intended use are available.
Automatically generated - may contain errors
2 protocols using ab15179
1
Immunohistochemical Staining of Tumor Samples
2
Protein Extraction and Western Blot Analysis
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Whole protein extracts were lysed using RIPA lysis buffer (Beyotime, Beijing, China) supplemented with protease inhibitors (Roche, CA, USA) according to manufacturer’s protocol. The proteins were separated by SDS-PAGE and transferred onto PVDF membranes, which were then blocked with TBS buffer containing 5% skim milk for 1 h at room temperature. After incubation with primary and secondary antibodies, the proteins were visualized by chemiluminescence. Antibodies used in this study are as follows: anti-YAP (dilution 1:1000; #14074, CST), anti-p-AKT (Ser473) (dilution 1:1000; #4058, CST), anti-AKT (dilution 1:1000; #9272, CST), anti-p-mTOR (dilution 1:1000; #9205, CST), anti-mTOR (dilution 1:1000; #2983, CST), anti-E-cadherin (dilution 1:500; #3195, CST), anti-Vimentin (dilution 1:500; #5741, CST), anti-Gli1 (dilution 1:1000; #ab15179, Abcam), anti-SMO (dilution 1:1000; #ab38686, Abcam) and anti-glyceraldehyde 3-phosphate dehydrogenase (dilution 1:5000; GAPDH, #AG019, Beyotime).
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