Antibodies against E-cadherin and N-cadherin were obtained from BD Bioscience (NJ, USA); ZEB1/TCF8, MMP-2, MMP-9, phospho-Akt (Ser 473), phospho-Akt (Thr 308), Akt, phospho-GSK3β (Ser9), GSK3β, phospho-PI3K, PI3K p85α, β-catenin, β-actin and lamin A/C antibodies were from Cell Signaling Technology (MA, USA); fibronectin and α-smooth muscle actin (SMA) antibodies were from Sigma-Aldrich Biotechnology (LP, USA); vimentin, green fluorescent protein (GFP), Twist, and Slug antibodies were from Santa Cruz Biotechnology (CA, USA); and TCTP antibodies were obtained from LabFrontier (Seoul, Korea). 2-(4-Morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002), rapamycin, and PP242 were obtained from Sigma-Aldrich Biotechnology (LP, USA).
Pi3k p85α
Manufactured by Cell Signaling Technology
Sourced in United States
PI3K p85α is a regulatory subunit of the phosphoinositide 3-kinase (PI3K) enzyme complex. It plays a core function in the activation of the PI3K signaling pathway, which is involved in various cellular processes such as cell growth, proliferation, and survival.
Automatically generated - may contain errors
Lab products found in correlation
Gsk3β, by Cell Signaling Technology (2 mentions)
P akt, by Cell Signaling Technology (2 mentions)
Nf κb p65, by Cell Signaling Technology (2 mentions)
Irdye 800cw donkey anti mouse igg, by LI COR (2 mentions)
Irdye 680rd donkey anti rabbit igg, by LI COR (2 mentions)
Stat3, by Cell Signaling Technology (2 mentions)
β tubulin, by Santa Cruz Biotechnology (2 mentions)
Phospho gsk 3β ser9, by Cell Signaling Technology (1 mentions)
Phospho pi3k, by Cell Signaling Technology (1 mentions)
Antibodies against e cadherin, by BD (1 mentions)
N cadherin, by BD (1 mentions)
β actin, by Cell Signaling Technology (1 mentions)
β catenin, by Cell Signaling Technology (1 mentions)
Lamin a c, by Cell Signaling Technology (1 mentions)
Mmp 2, by Cell Signaling Technology (1 mentions)
Mmp 9, by Cell Signaling Technology (1 mentions)
Control sirna, by Santa Cruz Biotechnology (1 mentions)
Anti gapdh antibody, by Abcam (1 mentions)
Horseradish peroxidase conjugated goat anti mouse or anti rabbit secondary antibodies, by Cytiva (1 mentions)
Phosphoserine 473 akt, by Cell Signaling Technology (1 mentions)
7 protocols using pi3k p85α
1
Investigating EMT Signaling Pathways
2
Tks5 Knockdown and Protein Detection
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siRNA targeting human Tks5, control siRNA, GST, and Myc (clone 9E10) antibodies, goat anti-Tks5 polyclonal antibody (M-20), and p21 antibody were from Santa Cruz Biotechnology (Santa Cruz, CA). Rabbit anti-Tks5 polyclonal antibody was a gift of S. Courtneidge (Sanford/Burnham Medical Research Institute, La Jolla, CA). XB130 monoclonal antibody was generated as previously described (Xu et al., 2007 (link)). XB130 siRNA and control siRNA were from GE Healthcare Dharmacon (Lafayette, CO). RGS-His antibody to detect 6xHis-tagged proteins was from Qiagen (Valencia, CA). Src (clone GD11) monoclonal antibody, phosphotyrosine (clone 4G10) monoclonal antibody, and GADPH antibody were from Upstate Biotechnology (Lake Placid, NY). Horseradish peroxidase–conjugated goat anti-mouse or anti-rabbit secondary antibodies were from Amersham Pharmacia Biotech (Piscataway, NJ). PI3K p85α, phosphotyrosine 458 PI3K, Akt, and phosphoserine 473 Akt antibodies were from Cell Signaling Technology (Beverly, MA). Anti-GAPDH antibodies were from Abcam (Cambridge, United Kingdom). Human Ki67 antibody and rabbit anti-Tks5 (SH3#1) were from EMD Millipore (Merck, Darmstadt, Germany).
3
Immunoblotting Analysis of Cellular Signaling
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Whole cell lysates were prepared and the immunoblotting was done as previously described [3 (link)] in Triton X 100-containing lysis buffer (50 mM Tris–HCl, pH 7.5, 10% glycerol, 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, 50 mM NaF). For Western blotting, the following antibodies were used: pAKT (S473, #6942), AKT (#9272), Bcl-xL (#2764), pERK1/2 (#9101), MEK1/2 (#4694), pMEK1/2 (#9121), ERK1/2 (#4696), NF-κB p65 (#6956), pNF-κB p65 (#3033), PARP (#9532), PI3K p85α (#13,666), pPI3K p85/p55 (#4228), PTK7 (#25618), Rac-1 (#4651), RhoA (#2117), ROR1 (#16,540), ROR2 (#88,639), Src (#2109), STAT3 (#9139), pSTAT3 (#9145) from Cell Signaling Technology (CST, Danvers, MA, USA); anti-pTYR 4G10 (#05–321) from Merck Millipore (Burlington, MA, USA); β-tubulin (#sc-166729) from Santa Cruz Biotechnology (Dallas, TX, USA); HA (#901,513) from BioLegend (San Diego, CA, USA). As secondary antibodies, IRDye® 800CW Donkey anti-Mouse IgG or IRDye® 680RD Donkey anti-Rabbit IgG (LI-COR, Lincoln, NE, USA) were used at 1:10 000 dilution.
4
Evaluation of Apoptosis-Related Proteins
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APG-2575 was provided by Ascentage Pharma (Suzhou) Co., Ltd and ABT-199 was purchased from Selleck Chemicals (Houston, TX, USA). HHT was purchased from Sigma-Aldrich (St. Louis, MO, USA). The antibodies GAPDH (#5174), PARP (#9532), Caspase3 and Cleaved-Caspase3 (Cleaved-C3) (#9662), Caspase7 and Cleaved-Caspase7 (Cleaved-C7) (#9494), PI3K (p85α) (#4257), AKT (#4691), AKT (Ser473) (#4060), GSK3β (#9315), GSK3β (Ser9) (#9323), MCL-1 (#94296), MCL-1 (Thr163) (#14765) and MCL-1 (Thr163/Ser159) (#4579) were purchased from Cell Signaling Technology (CST, Beverly, MA, USA).
5
Protein Expression Analysis in Skeletal Muscle
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Total protein was extracted from skeletal muscle by the protein extraction Kit (KeyGEN). Protein concentration was determined with BCA Protein Assay Kit (Takara), and an equal amount of protein was separated on SDS-polyacrylamide gel electrophoresis. After transferring to a polyvinylidene fluoride (PVDF) membrane (Millipore), the primary antibody and the corresponding secondary antibody were used to detect protein of interest. The antibody used in this study were as follows: Atrogin-1 (Abclonal); MuRF-1 (Abclonal); LC3 (Abclonal); P62 (Abclonal); Ndufb2 (Abclonal); Clusterin (Abclonal); IGF1 (Abclonal); PI3K(p85α) (Abclonal); P-mTOR (Abclonal); mTOR (Abclonal); P-P70S6K (Abclonal); P70S6K (Abclonal); P-FOXO3A (Abclonal); FOXO3A (Abclonal); P-EIF4EBP1 (Abclonal); EIF4EBP1 (Abclonal); P-AKT (Cell Signaling Technology); AKT (Proteintech); and GAPDH (Bioworld Technology). The protein was visualized using High-sig ECL Western Blotting Substrate (Tanon) and imaged using the Tanon-5200S Chemiluminescent Imaging System (Tanon).
6
Protein Expression Analysis of Frozen Tumors
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Frozen tumor pieces were thawed, washed 2X with cold PBS and pestered to lyse in lysis buffer (50 mM Tris–HCl pH 7.5, 10% glycerol, 150 mM NaCl, 1 mM EDTA, 1% Triton-x-100, 50 mM NaF) supplemented with protease and phosphatase inhibitor cocktails (Bimake, Houston, TX, USA). Lysates were mixed with 4X Laemmli loading buffer and subjected to SDS-PAGE gel electrophoresis and Western blotting. The primary antibodies used were as following: pAkt S473 (#4060), pMEK1/2 S217/221 (#9121), NF-κB p65 (#6956), pPI3K p85 Y458/p55 Y199 (#4228), PI3K p85α (#13666), Rac-1 (#4651), pSTAT3 Y705 (#9145), STAT3 (#9139), Wnt5a/b (#2530) (Cell Signaling Technology, Danvers, MA, USA); Akt (#sc-5298), Bcl-2 (#sc-7382), MEK1/2 (#sc-6250), β-tubulin (#sc-166,729) (Santa Cruz, Dallas, TX, USA); ROR1 6D4 (Dr. Riddel lab, ref. Balakrishana et al. 2016); ROR2 (#565550, BD Biosciences, San Jose, CA, USA). Secondary antibodies: IRDye® 800CW Donkey anti-Mouse IgG, or IRDye® 680RD Donkey anti-Rabbit IgG (LI-COR, Lincoln, NE, USA). Blots were scanned and quantified using Odyssey CLx and Image Studio software (LI-COR). Protein quantification was normalized to β-tubulin expression level for each sample.
7
Protein Expression Analysis in Rat Tissues
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After the rats were scarified, the gastrocnemius muscle and liver were quickly frozen in liquid nitrogen, and stored at −80 °C. The tissues were homogenized with the T-PER™ tissue protein extraction buffer (Pierce Biotechnology, Thermo-Fisher, Rockford, IL, USA) containing 1% protease inhibitor cocktail and 1% phosphatase inhibitor cocktail (Calbiochem, Merck Millipore, Darmstadt, Germany). The protein samples in the supernatants were collected after centrifugation at 15,000× g at 4 °C for 10 min, and analyzed by Western blotting with primary antibodies detecting IRS-1 (1:1000, GeneTex, San Antonio, TX, USA), PI3-k p85 α (1:1000; Cell Signaling Technology, Beverly, MA, USA), SREBP1c, CD36 (1:1000; Novus Biologicals, Littleton, CO, USA), and Glyceraldehyde 3-phosphate dehydrogenase (GAPDH, 1:20,000; Merck Millipore), and then detected with secondary antibodies (anti-rabbit or anti-mouse IgG HRP-conjugated). The blots were quantified by chemiluminescence with MiniChemi I system (Beijing Sage Creation Science, Beijing, China), and then normalizing with GAPDH. The relative protein intensity was expressed as folds of the content in the NC group.
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