Sc 28199

Manufactured by Santa Cruz Biotechnology

Sourced in United States

Sc-28199 is a laboratory product offered by Santa Cruz Biotechnology. It is a device used for scientific research and analysis. The core function of Sc-28199 is to provide a tool for researchers and scientists to conduct their experiments and studies.

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Anti-SNAIL (sc-28199, (3 citations)

5 protocols using sc 28199

Western Blot Analysis of EMT Markers

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Cells (1×10⁶ cells/ml) were dissolved using the radioimmunoprecipitation assay lysis buffer (Beyotime Institute of Biotechnology) and the lysate was centrifuged at 10,000 × g at 4°C for 30 min following incubation on ice for 50 min. Then, each sample containing 40 µg protein as subjected to 12% (w/v) SDS-PAGE and transferred to polyvinylidene fluoride transfer membranes (GE Healthcare, Chicago, IL, USA). Following blocking in 5% (w/v) skimmed milk (25°C for 1 h), membranes were incubated (4°C, overnight) with anti-E-cadherin antibody (1:1,000; sc-7870; Santa Cruz Biotechnology), anti-N-cadherin antibody (1:1,000; sc-7939; Santa Cruz Biotechnology), anti-Snail antibody (1:1,000; sc-28199; Santa Cruz Biotechnology), anti-HIF-1α antibody (1:1,000; sc-10790; Santa Cruz Biotechnology), anti-Notch1 antibody (1:1,000; sc-9170; Santa Cruz Biotechnology) and anti-GAPDH antibody (1:1,000; sc-25778; Santa Cruz Biotechnology), and then incubated (25°C for 1 h) with the horseradish peroxidase-conjugated anti-rabbit secondary antibody (1:5,000; sc-2004; Santa Cruz Biotechnology). Finally, proteins were detected by chemiluminescence (PerkinElmer, Inc., Waltham, MA, USA) and densitometric analysis used ImageJ software (1.8.0_112; National Institutes of Health, Bethesda, MD, USA).

Chen D.W., Wang H., Bao Y.F, & Xie K. (2018). Notch signaling molecule is involved in the invasion of MiaPaCa2 cells induced by CoCl2 via regulating epithelial-mesenchymal transition. Molecular Medicine Reports, 17(4), 4965-4972.

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Protein Extraction and Western Blot Analysis

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Total protein was extracted from the MPC5 cells using RIPA lysis buffer (Beyotime, Jiangsu, China) following the manufacturer's instructions. Western blot analysis was performed as previously described (25 (link)). The antibodies used were the following: rabbit anti-desmin (1:1,000; sc-14026), rabbit anti-snail (1:1,000; sc-28199) (all from Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA), rabbit anti-nephrin (1:2,000; ab136894; Abcam, Cambridge, MA, USA), rabbit anti-Wilms tumor 1 (WT1; 1:500; sc-192), rabbit anti-E-cadherin (1:1,000l; sc-7870) (both from Santa Cruz Biotechnology, Inc.), rabbit anti-GSK3β (1:2,000; ab18893; Abcam), rabbit anti-β-catenin [activated and unphosphorylated, 1:2,000; Netherlands Cancer Institute (NKI), Amsterdam, The Netherlands], mouse anti-β-tubulin (1:3,000; sc-80011), goat anti-rabbit IgG-HRP (1:3,000; sc-2004) and goat anti-mouse IgG-HRP (1:3,000; sc-2005) (all from Santa Cruz Biotechnology, Inc.). For the quantitative analysis of the western blots, the protein band intensities were quantified using Image J software and β-actin was used for normalization.

YANG X., WANG X., NIE F., LIU T., YU X., WANG H., LI Q., PENG R., MAO Z., ZHOU Q, & LI G. (2015). miR-135 family members mediate podocyte injury through the activation of Wnt/β-catenin signaling. International Journal of Molecular Medicine, 36(3), 669-677.

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Western Blot Analysis of EMT Markers

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Western blotting was conducted as previously described [25 (link)]. The primary antibodies used in this study are as follows: anti-CTSL (Abcam, ab6314), anti-p53 (Abcam, ab1101), anti-E-cadherin (Abcam, ab1416), anti-N-cadherin (Abcam, ab18203), anti-Vimentin (Abcam, ab92547), anti-Zeb1 (Abcam, ab203829), anti-Snail (Santa Cruz, sc-28199), anti-Egr-1 (Santa Cruz, sc-189), and loading control anti-β-actin (Abcam, ab8226). Finally, the primary antibodies were probed with rabbit or mouse secondary antibodies labeled with DyLight 800 (KPL) and scanned with the Odyssey Infrared Imaging System (LI-COR).

Wang W., Xiong Y., Ding X., Wang L., Zhao Y., Fei Y., Zhu Y., Shen X., Tan C, & Liang Z. (2019). Cathepsin L activated by mutant p53 and Egr-1 promotes ionizing radiation-induced EMT in human NSCLC. Journal of Experimental & Clinical Cancer Research : CR, 38, 61.

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Western Blot Analysis of Snail-1

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Briefly, protein content of cells was isolated by RIPA buffer (25 mM Tris HCl pH 7.6, 150 mM NaCl, 1% NP-40, 1% sodium deoxycholate, 0.1% SDS). Samples containing 50 μg total protein were separated on 12.5% SDS polyacrylamide gel, and then electroblotted to supported PVDF membranes (Roche Diagnostics GmbH). The membranes were then blocked overnight at room temperature with 3% bovine serum albumin (BSA) in TBS-T (1× Tris-Buffered Saline, 0.1% Tween-20). Primary Rabbit polyclonal antibodies were used for detection of Snail-1 (1:500, sc-28199, Santa Cruz Biotechnology) and β-actin (1:3000, monoclonal antibody, Abcam), which were diluted with 0.05% Tween-20 in PBS. After washing several times, the blots were incubated with horseradish peroxidase (HRP)-conjugated goat anti-rabbit secondary polyclonal antibody (1:3000, Santa Cruz Biotechnology). To assess the signals, ECL kit (Roche Diagnostics GmbH) was applied and quantified by NIH ImageJ 1.63 Software.

Hemmatzadeh M., Mohammadi H., Babaie F., Yousefi M., Ebrazeh M., Mansoori B., Shanehbandi D, & Baradaran B. (2018). Snail-1 Silencing by siRNA Inhibits Migration of TE-8 Esophageal Cancer Cells Through Downregulation of Metastasis-Related Genes. Advanced Pharmaceutical Bulletin, 8(3), 437-445.

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Immunohistochemical Analysis of EMT Markers

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5‐μm sections from formalin‐fixed, paraffin‐embedded (FFPE) tissue were stained by immunohistochemistry for Twist (rabbit polyclonal antibody [H‐81]: sc‐15393 [Santa Cruz Biotechnology, Santa Cruz, CA, USA]), Slug (monoclonal rabbit antibody [C19G7] [Cell Signaling Technology®, Danvers, MA, USA]), and Snail (polyclonal rabbit antibody [H‐130]: sc‐28199 [Santa Cruz Biotechnology]). Staining for Hif‐1α (monoclonal mouse antibody clone H1α67 [sc‐53546; Santa Cruz Biotechnology]) was performed according to previous protocols [40]. Further information regarding the immunohistochemical staining methods can be found in supplementary material, Table S2. Positive controls (multiorgan TMA sections with known expression of the relevant antigen) and negative controls (isotypic immunoglobulin or antibody diluent without the primary antibody) were included. In addition, smooth muscle cells in the intervening stroma served as the positive control for Slug.

Børretzen A., Gravdal K., Haukaas S.A., Mannelqvist M., Beisland C., Akslen L.A, & Halvorsen O.J. (2021). The epithelial–mesenchymal transition regulators Twist, Slug, and Snail are associated with aggressive tumour features and poor outcome in prostate cancer patients. The Journal of Pathology: Clinical Research, 7(3), 253-270.

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