The protocol and procedure for western blotting were as described in our previous reports 30 (link), 44 (link), 45 (link). The anti-p-YAP (Ser127), anti-YAP, anti-RalA and anti-β-actin primary antibodies were obtained from Cell Signalling Technology (Danvers, MA, USA). The anti-Wnt5a, anti-Wnt5b, anti-Aggrecan and anti-SOX9 primary antibodies were obtained from Abcam (Cambridge, MA, USA). The anti-CD63, anti-CD9, anti-CD81 and anti-Alix were obtained from System Biosciences (Palo Alto, CA, USA). Based on previous research 46 (link), 47 (link), collagen type II was analysed using 5% (wt/vol) SDS-PAGE with anti-collagen type II primary antibodies (Chemicon®, Merck-Millipore).
Anti wnt5b
Manufactured by Abcam
Sourced in United States, China
Anti-Wnt5b is a primary antibody that detects the Wnt5b protein. Wnt5b is a member of the Wnt family of signaling proteins involved in various cellular processes. This antibody can be used to detect and quantify Wnt5b expression in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Anti β actin, by Cell Signaling Technology (2 mentions)
Anti wnt16, by Abcam (2 mentions)
Anti wnt 2, by Abcam (2 mentions)
Radioimmunoprecipitation assay (ripa), by Beyotime (2 mentions)
Protease inhibitor mixture cocktail, by Roche (2 mentions)
Anti wnt5a, by Abcam (1 mentions)
Anti sox9, by Abcam (1 mentions)
Anti rala, by Cell Signaling Technology (1 mentions)
Anti cd9, by System Biosciences (1 mentions)
Anti cd63, by System Biosciences (1 mentions)
Anti cd81, by System Biosciences (1 mentions)
Anti aggrecan, by Abcam (1 mentions)
Anti rapsyn, by Abcam (1 mentions)
Anti bcr abl, by Abcam (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Anti β catenin, by Proteintech (1 mentions)
Anti c myc, by Proteintech (1 mentions)
Anti β tubulin, by Proteintech (1 mentions)
Anti src, by Abcam (1 mentions)
Anti e cadherin, by Abcam (1 mentions)
4 protocols using anti wnt5b
1
Western Blotting Protocol for Protein Analysis
2
Western Blot Analysis of Protein Expression
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Total proteins were obtained by cell lysis (Beyotime, Nanjing, China), and protein samples of the same concentration were electrophorized on 8% or 10% SDS-PAGE and then transferred to PVDF membranes (Millipore, USA). After incubating with anti-RAPSYN (Abcam, Cambridge, GB), anti-BCR-ABL (Abcam, Cambridge, GB), anti-Wnt5b (Abcam, Cambridge, GB), anti-c-Myc (Proteintech, Wuhan, China), anti-β-catenin (Proteintech, Wuhan, China), and anti-β-Tubulin (Proteintech, Wuhan, China) overnight at 4 ℃, then incubated with secondary antibody of goat anti-rabbit IgG/HRP (CST technology, Boston, USA) or goat anti-mouse IgG/HRP (CST technology, Boston, USA) at room temperature for 2 h. Finally, protein bands were visualized using ECL system (Tanon 3500).
3
Molecular Profiling of Epithelial-Mesenchymal Transition
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Experimental cancer cells (after culturing in HUVEC-CM for 21 days) and respective control cells were lysed on ice in a lysis buffer (RIPA, Beyotime, Shanghai, China) with a protease inhibitor mixture cocktail (Roche, Switzerland). Western-blot analysis was performed by established protocols 12 . Anti-α-catenin, anti-E-cadherin, anti-ZEB-1, ZEB-2, Snail, Slug, anti-ZO-1 and anti-Laminin A1 primary antibodies were purchased from (Abcam); anti-MMP-1, -2, -3, -11, -12, -13, -17 and -21, anti- ITGA6, B1, B3, B4, B7, anti-FAK, P-FAK-Y397, anti-Src, P-Src-Y418, P-Src-Y529, anti-fibronectin, anti-Laminin B3, anti-Wnt-2, anti-Wnt-5B, anti-Wnt-16, anti-TGF-β, anti-β-catenin and anti-N-cadherin primary antibodies were purchased from (Epitomics); anti-Cdc-2, P-Cdc-2-Tyr15, CDK4, CDK6, Cyclin A, Cyclin D1, Cyclin D3, Cyclin E2, P15, P16, P21, P27, P53, Rb, P-Rb-S811, P-Rb-S780; anti-Bcl-xl, Bad, P-Bad-Ser112, Bak, Mcl-1, Puma and anti-β-actin were from (Cell Signaling Technology).
4
Western Blot Analysis of Liver Cancer Cells
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Following culture in MRC-5-CM for 14 days, whole liver cancer cells were lysed on ice in a lysis buffer (RIPA, Beyotime, Shanghai, China) with a protease inhibitor mixture cocktail (Roche, Switzerland). After centrifugation at 12000rpm for 30 minutes at 4℃, the protein concentrations of supernatants in samples were measured by the BCA protein assay (Thermo scientific, Rockford, IL, USA). Equal amounts of protein (50μg) were separated by 10%-12% NUPAGE Bis-tris Gel (Invitrogen, CA, USA) electrophoresis (constant voltage: 120mv) and transferred onto polyvinylidene fluoride (PVDF, 0.45μm) membranes (constant current: 350mA for 70/120 min). After being blocked by Tris-buffered saline and Tween 20 (TBST) buffer containing 5% non-fat powder milk for 2h, the membranes were incubated with primary antibodies overnight on ice. After washing the membranes several times in TBST while agitating, detection was performed using the appropriate secondary HRP-conjugated anti-mouse or anti-rabbit antibody. Immunoreactive bands on the blots were visualized with enhanced chemiluminescence reagent ECL kit (Beit Haemek, Israel). Anti-GAPDH, anti-WNT-2, anti-WNT-5B, anti-WNT16, anti-TGFB1, anti-CTNNB1, anti-IL6, anti-Nanog, anti-OCT4 and anti-CK19 primary antibodies were purchased from (Epitomics).
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