Total protein was extracted with a cell lysis reagent and the concentration was measured using a BCA protein assay kit (Beyotime Biotec, China). Protein samples were separated by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis. The separated proteins were transferred onto a polyvinylidene difluoride membrane (Millipore, MA, USA), which was then blocked with 5% skim milk powder dissolved in Tris-buffered saline-Tween. After incubation at room temperature for 1 h, the membrane was incubated overnight with primary antibodies at 4°C. The primary antibodies of anti-PPARG, anti-E-cadherin, anti-Vimentin, and anti-GAPDH were purchased from Abcam (MA, USA); anti-Snail from Gene Tex (CA, USA) and NF-κB from Cell Signaling Technology (MA, USA). The membranes were then incubated for 1 h with horseradish peroxidase-conjugated secondary antibody (Abcam, MA, USA) at room temperature. The proteins were detected using an ECL detection Kit (Millipore, MA, USA). The signal density of protein bands was quantified using the Tanon-5200 Imaging System (Tanon Science & Technology, China). GAPDH was used as a loading control.
Anti snail
Manufactured by GeneTex
Sourced in United States
Anti-Snail is a laboratory reagent used to detect the transcription factor Snail in biological samples. It is an antibody that specifically binds to the Snail protein, allowing for its identification and quantification through various experimental techniques.
Automatically generated - may contain errors
Lab products found in correlation
Anti e cadherin, by GeneTex (3 mentions)
Anti twist, by GeneTex (3 mentions)
Anti vimentin, by GeneTex (2 mentions)
Anti zeb1, by GeneTex (2 mentions)
Anti slug, by GeneTex (2 mentions)
Anti n cadherin, by GeneTex (2 mentions)
Anti vimentin, by Santa Cruz Biotechnology (2 mentions)
Horseradish peroxidase conjugated rabbit anti mouse or goat anti rabbit secondary antibodies, by Santa Cruz Biotechnology (2 mentions)
Anti e cadherin, by BD (2 mentions)
Polyvinylidene difluoride membrane, by Merck Group (1 mentions)
Anti pparg, by Abcam (1 mentions)
Horseradish peroxidase conjugated secondary antibody, by Abcam (1 mentions)
Ecl detection kit, by Merck Group (1 mentions)
Nf κb, by Cell Signaling Technology (1 mentions)
Tanon 5200 imaging system, by Shanghai Tanon Science & Technology (1 mentions)
Bca protein assay kit, by Beyotime (1 mentions)
Anti gapdh, by Abcam (1 mentions)
Anti e cadherin, by Abcam (1 mentions)
Anti vimentin, by Abcam (1 mentions)
Alexa fluor 488 goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
7 protocols using anti snail
1
Western Blot Analysis of Protein Expression
2
Immunostaining of Drosophila Larval Hearts
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Immunostaining on whole mount larvae or dissected larval hearts was performed as previously described (Yang and Xu, 2012 ). The primary antibodies used in this study included the following: anti-phospho-Smad3 (rabbit; Abcam, 52903), anti-phospho-Smad1/5/9 (rabbit; CST, 13820), anti-phospho-histone H3 (rabbit; Merck Millipore, 06570), anti-MHC (mouse; DSHB, MF20), anti-Twist (mouse; Santa Cruz, 81417), and anti-Snail (rabbit; Genetex, 125918). The secondary antibodies included the following: Alexa Fluor 488 goat anti-rabbit IgG, Alexa Fluor 488 goat anti-mouse IgG, Alexa Fluor 555 goat anti-mouse IgG, and Alexa Fluor 647 goat anti-rabbit IgG from Thermo Fisher Scientific.
3
Resveratrol and Viniferin Modulators
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Resveratrol (CAS Number 501-36-0, Purity: ≥99%), ε-viniferin (CAS Number 62218-08-0, Purity: ≥99.5%), SB 431542 (CAS Number 301836-41-9, Purity: ≥98%), and [3-(4, 5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide] (MTT) (CAS Number: 298-93-1) were purchased from Sigma-Aldrich, Burlington, MA, USA. α-Viniferin (CAS Number 62218-13-7, Purity: ≥98%) and kobophenol A were purchased from SunHank Technology, Tainan City, Taiwan. N-Acetyl-L-cysteine (CAS Number 616-91-1) was purchased from Merck Millipore, Burlington, MA, USA. Reactive oxygen species (ROS) Detection Assay Kits (Catalog No. K936-100) were purchased from BioVision. Anti-β-actin antibody (Cat No. GTX109639), anti-vimentin, anti-MMP2, anti-Slug, anti-Zeb1, anti-Snail, anti-E-cadherin, anti-p-SMAD2, anti-p-SMAD3 and ATP-binding cassette sub-family G member 2 (ABCG2) antibody were purchased from GeneTex.
4
Molecular Signaling Pathways in Cellular Processes
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Acrylamide, N,N′-methylenebis-Acrylamide, and RETROscript (reverse transcriptase [RT]-PCR kit; Ambion; ThermoFisher Scientific) were procured from Invitrogen BioServices India Pvt. Ltd. Taq-polymerase was procured from HiMedia. Ethidium bromide (EtBr), agarose, sodium arsenite (NaAsO2), diaminobenzidine (DAB), 2′,7′-dichlorofluorescin diacetate, and bovine serum albumin were obtained from Sigma-Aldrich. Goat Immunoglobulin G anti-rabbit and anti-mouse (alkaline phosphatase conjugated and horseradish peroxidase conjugated) were purchased from Genetex; 5-bromo-4-chloro-3-indolyl phosphate/nitro blue tetrazolium (BCIP-NBT) was bought from Santa Cruz Biotechnology; nitrocellulose membrane was purchased from GE HealthCare; Glycine, Tris and SDS were purchased from Amresco. Primary antibodies anti-TGF-β, anti-Smad2 (phospho [p]Ser467), anti-Smad3 (pSer423/425), anti-phosphatase and tensin homolog deleted on chromosome 10 (PTEN), anti-PI3Kp110α, anti-AKT (pSer473), anti-mTOR, anti-S6Kp70, anti-NF-κB/p65, anti-TAK1 (pThr184), anti-MAPKK3 (pSer189), anti-p38 MAPK, anti-MAPKK4 (pThr 261), anti-JNK, anti-E-cadherin, anti-desmoplakin, anti-vimentin, anti-N-cadherin, anti-Snail, anti-Slug, anti-Twist, anti-β-actin, and anti-Zeb1 were purchased from Genetex. Anti-Smad4 (pThr277) was purchased from ThermoFischer Scientific.
5
Quantitative real-time PCR and immunoblotting assay
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Assays for real-time PCR and immunoblotting were conducted as described previously.7 (link), 9 (link) These primers used in real-time PCR were shown in Supplementary Table 3 . The following primary antibodies were used in immunoblotting assay: anti-V5 (MCA1360; AbD Serotec, Raleigh, NC, USA), anti-NKX6.1 (generated by Yao-Hong Biotechnology Inc., Taipei, Taiwan), anti-E-cadherin (610404; BD Biosciences, San Jose, CA, USA), anti-vimentin (sc-6260; Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti-N-cadherin (610920; BD Biosciences), anti-SNAIL (GTX100754; GeneTex, Irvine, CA, USA), anti-TWIST (GTX127310; GeneTex) and anti-SLUG (sc-10436; Santa Cruz Biotechnology), anti-BAF155 (GTX114777; GeneTex) and anti-RBBP7 (sc-8272; Santa Cruz Biotechnology). Horseradish peroxidase-conjugated rabbit anti-mouse or goat anti-rabbit secondary antibodies (Santa Cruz Biotechnology) were used as appropriate.
6
Protein Expression Analysis by Western Blot
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Confluent cells were collected and lysed in RIPA buffer (Genestar Biotechnology, Taiwan). In total, 30 μg of proteins was separated by 10% sodium dodecylsulfate - polyacrylamide gel electrophoresis (SDS-PAGE) and transferred onto polyvinylidene difluoride (PVDF) membranes (Life Science, Carlsbad, CA, USA). After blocking, the membranes were hybridized with specific primary antibodies followed by incubation with secondary antibodies conjugated to horseradish peroxidase (HRP). The protein images were detected using the Western Blotting Plus Chemiluminescence Reagent (Thermo Scientific, Waltham, MA, USA). The density of each protein band was determined after normalization to an actin control band using the Gel Image System and Image J software (Scion Corporation, Torrance, MD, USA). The primary antibodies used included anti-ANXA2 (R&D System, Minneapolis, MN USA), anti-Akt, anti- Akt S473p, anti-Akt T308p, anti-Snail, anti-TWIST, anti-E-cadherin, and anti-N-cadherin (all from Genetex, Irvine, CA, USA). Protein images were detected using Western Blotting Plus Chemiluminescence Reagent (Thermo Scientific, Waltham, MA, USA). The density of each protein band was determined after normalization to an actin control band using the Gel Image System and Image J software.
7
Quantifying Protein Expression Levels
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Assays for real-time PCR and immunoblotting were conducted as described previously [53 (link),54 (link)]. The primers used in real-time PCR are shown in Supplementary Table S2 . The following primary antibodies were used in the immunoblotting assay: anti-NKX6.1 (AF5857; R&D systems, Minneapolis, MN, USA), anti-E-cadherin (610405; BD Biosciences, San Jose, CA, USA), anti-vimentin (sc-6260; Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti-SNAIL (GTX100754; GeneTex, Irvine, CA, USA), anti-TWIST (ab49254; Abcam, Cambridge, MA, USA), and anti-SLUG (#9585; Cell Signaling, Danvers, MA, USA). Horseradish peroxidase-conjugated rabbit anti-mouse or goat anti-rabbit secondary antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA) were used as appropriate.
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