The proteins from the GC cells and tissues were extracted with M-per protein lysis buffer (Thermo Fisher Scientific, Waltham, MA, USA) supplemented with protease and phosphatase inhibitors (BBI Life Sciences Corporation, Shanghai, China) and subjected to western blotting [36 (link)] with the primary antibodies anti-GAPDH (No 7074 T; Cell Signaling Technology (CST), Danvers, MA, USA), anti-TMEM176B (No. PHC0758; Abmart, Wuhan, China), anti-ASNS (No. R22614; ZenBio, Chengdu, China), anti-AKT (No. R23411; ZenBio), anti-p-AKT (No. R381555; ZenBio), anti-PI3K (No. R381092; ZenBio), anti-p-PI3K (No. 310164; ZenBio), and anti-PI3K (No. R381092; ZenBio). The mTOR Substrates Antibody Sampler Kit (No. 9862 T) was sourced from CST.
Anti akt
Manufactured by ZenBio
Sourced in China
Anti-AKT is a laboratory reagent used for the detection and quantification of the AKT protein, which plays a critical role in cellular signaling pathways. It provides a tool for researchers to study the expression and activity of AKT in various biological systems.
Automatically generated - may contain errors
Lab products found in correlation
Anti p akt, by ZenBio (2 mentions)
Anti tmem176b, by Abmart (1 mentions)
M per protein lysis buffer, by Thermo Fisher Scientific (1 mentions)
Anti pi3k, by ZenBio (1 mentions)
Anti gapdh, by Cell Signaling Technology (1 mentions)
Enhanced chemiluminescence reagent, by Merck Group (1 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Anti gapdh, by ZenBio (1 mentions)
Anti erk, by Cell Signaling Technology (1 mentions)
Anti p akt, by Cell Signaling Technology (1 mentions)
Anti p erk, by Cell Signaling Technology (1 mentions)
Bca protein assay kit, by Beyotime (1 mentions)
Omin ecl ultra sensitive chemiluminescence detection kit, by Ipsen (1 mentions)
Anti gapdh, by Proteintech (1 mentions)
Anti bax, by ZenBio (1 mentions)
Anti cd36, by Proteintech (1 mentions)
Alexa fluor 555 labeled donkey anti rabbit igg h l secondary antibody, by Beyotime (1 mentions)
Anti phospho akt, by Proteintech (1 mentions)
4 protocols using anti akt
1
Protein Expression Analysis in GC Cells
2
Western Blot Analysis of Protein Expression
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The protein concentration was quantified by the BCA Protein Assay Kit (Beyotime) and normalized. Approximately 30 µg of protein extracts was analyzed by electrophoresis using a 12% SDS–PAGE gel and electroblotted onto polyvinylidene fluoride membranes (Millipore). The membranes were blocked with 5% BSA and incubated with specific primary antibodies overnight at 4 °C with gentle rotation. A horseradish peroxidase-labeled secondary antibody was added, incubated for 1 h with shaking and visualized using an enhanced chemiluminescence reagent (Millipore). The primary antibodies included anti-GAPDH (#200306-TE4, 1:2 000), anti-FAK (#860324, 1:1 000), anti-p-FAK (#381143, 1:1 000), anti-ITGB1 (#R24729, 1:1 000), anti-ITGA5 (#R24725, 1:1 000), and anti-ITGB3 (#384730, 1:1 000) from Zen BioScience and anti-AKT (#4691, 1:1 000), anti-p-AKT (#4060, 1:1 000), anti-ERK (#4695, 1:1 000), and anti-p-ERK (#4370, 1:1 000) from Cell Signaling.
3
Western Blot Analysis of Kidney Proteins
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Proteins supernatant was extracted from kidney and analyzed by Western blotting. Equal amounts of proteins were separated by SDS-PAGE and transferred to PVDF membranes. The membrane was then blocked with skimmed milk for 1 h, incubated with primary antibody overnight at 4 °C. After cleaning with TBST, the membranes were incubated with Goat Anti-rabbit HRP antibody (1:5000; Bioss, Beijing, China) at room temperature for 1 h and the signals were detected with the Omin-ECL ultra-sensitive chemiluminescence detection kit (EpiZyme, Shanghai, China), and their intensities were measured with Image J 6.0 software and analyzed by GraphPad Prism 8. Primary antibodies used in this study included anti-VEGF (1：1000，ZEN-BIOSCIENCE, China), anti-p-VEGFR (1:1000, Abclonal), anti-AKT (1:1000; ZEN-BIOSCIENCE, Chengdu, China), anti-p-AKT (1:1000; ZEN-BIOSCIENCE, Chengdu, China), anti-ERK1/2 (1:1000; ZEN-BIOSCIENCE, Chengdu, China), anti-p-ERK1/2 (1:1000; Boster, Wuhan, China; 1:1000,Abclonal) and anti-GAPDH (1:1000; Proteintech), GAPDH served as an internal control.
4
Recombinant CD5L Protein Signaling Pathways
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Recombinant human CD5L protein was purchased from R&D Systems (Minneapolis, USA). Anti-phospho-AKT, anti-CD36, and anti-CD55 primary antibody were purchased from proteintech (Chicago, USA). MEK inhibitor U0126 was purchased from Beyotime (Shanghai, China) and was diluent in DMSO. Anti-phospho-JAK, antiphospho-p38 MAPK, anti-phospho-ERK, anti-phospho-IκB-α, anti-AKT, anti-JAK, anti-p38 MAPK, anti-IκB-α, anti-Bax, anti-Bcl-2, HRP-conjugated rabbit secondary antibody, and goat secondary antibody were purchased from Zenbio (Chengdu, China). Rabbit derived anti-ERK was purchased from Wanleibio(Shenyang, China). Alexa Fluor 555 labeled donkey anti-rabbit IgG (H + L) secondary antibody was purchased from Beyotime (Shanghai, China). The cell culture system was purchased from Gibco (Carlsbad, USA).
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