The following materials were purchased from commercial companies: TNFα (PeroTech; Rocky Hill, NJ, USA); pan-caspase inhibitor z-VAD-fmk (Abcam, Cambridge, MA, USA). InSolution Akt Inhibitor viii isozyme-selective, Akti-1/2 and InSolution rapamycin were obtained from Calbiochem (San Diego, CA, USA). MitoSox Red was obtained from Invitrogen (Carlsbad, CA, USA). Hoechst 33258, butylated hydroxyanisole (BHA) and rotenone were obtained from Sigma (St. Louis, MO, USA). Nec-1 (5-(7-chloro-1H-indol-3-ylmethyl)-3-methylimidazolidine-2,4-dione), the inactive analog of necrostatin-1 analog (Nec-1i; 5-(7-chloro-1H-indol-3-ylmethyl)imidazolidine-2,4-dione),3 (link), 12 (link), 13 (link) was a kind gift from Dr. Greg Cuny. LOX-1 was obtained from Chembridge (compound 5680672; San Diego, CA, USA), and Bai was from Cayman Chemicals (Ann Arbor, MI, USA). Antibodies were obtained from commercial sources: anti-pAkt-473, anti-p–Akt-308, anti-p-GSK-3β, anti-p-mTOR, anti-p-S6, anti-mTOR, anti-Akt1, anti-Akt2, anti-Akt3, and anti-Akt from Cell Signaling (Danvers, MA, USA); anti-RIPK1 from BD Transduction Laboratories (Lexington, KY, USA); anti-RIPK3 from ProSci Incorporated (San Diego, CA, USA); anti-HMGB1 and β-actin were obtained from Abcam.
Anti p akt 308
Manufactured by Cell Signaling Technology
Sourced in United States
Anti-p-Akt-308 is a primary antibody that specifically recognizes the phosphorylated form of Akt at Serine 308. It is intended for use in various research applications, such as western blotting, immunohistochemistry, and flow cytometry, to detect and analyze the phosphorylation status of Akt in cellular samples.
Automatically generated - may contain errors
Lab products found in correlation
Anti pakt473, by Cell Signaling Technology (3 mentions)
Anti akt, by Cell Signaling Technology (2 mentions)
Anti akt1, by Cell Signaling Technology (1 mentions)
Butylated hydroxyanisole bha, by Merck Group (1 mentions)
Anti hmgb1, by Abcam (1 mentions)
Anti ripk1, by BD (1 mentions)
Anti mtor, by Cell Signaling Technology (1 mentions)
Anti p mtor, by Cell Signaling Technology (1 mentions)
Anti akt3, by Cell Signaling Technology (1 mentions)
Anti akt2, by Cell Signaling Technology (1 mentions)
Z vad fmk, by Abcam (1 mentions)
Anti p gsk 3β, by Cell Signaling Technology (1 mentions)
Anti ps6, by Cell Signaling Technology (1 mentions)
Mitosox red, by Thermo Fisher Scientific (1 mentions)
Hoechst 33258, by Merck Group (1 mentions)
Rotenone, by Merck Group (1 mentions)
β actin, by Abcam (1 mentions)
Anti ripk3, by ProSci (1 mentions)
Ab54481, by Abcam (1 mentions)
Ab110413, by Abcam (1 mentions)
4 protocols using anti p akt 308
1
Inhibition of Akt and Necroptosis Pathways
2
Protein Expression Analysis Protocol
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Tissues were dissolved in RIPA buffer (150 mM sodium chloride, 1.0% Triton X-100, 0.5% sodium deoxycholate, 0.1% SDS, 50 mM Tris, protease and phosphatase inhibitor mixture (Roche Diagnostics)). Protein concentrations were determined using a BCA assay kit (Pierce Diagnostics). Protein was separated by 10% (wt/vol) SDS/PAGE, transferred to a PVDF membrane (Millipore), blocked in 5% (wt/vol) skim milk in TBST (0.02 M Trisbase, 0.14 M Vehicle, 0.1% Tween 20, pH 7.4), and incubated with primary antibodies overnight at 4 °C and then incubated with secondary antibodies conjugated with HRP. The following primary antibodies were used: anti-UCP1 (ab10983, Abcam), anti-PGC1ɑ (ab54481, Abcam), anti-OXPHOS (ab110413, Abcam), anti-Mas1 (AAR-013, Alomone labs), anti-Akt (#9272, cell signaling technology), anti-p-Akt308 (#13038, cell signaling technology), anti-FoxO1 (#2880, cell signaling technology), anti-p-FoxO1 (#84192, cell signaling technology), anti-PKA (#4782, cell signaling technology), anti-p-PKA (#9621, cell signaling technology), anti-ACE2 (#92485, cell signaling technology), and actin (#4970, Cell Signaling Technology). Signals were detected with Super Signal West Pico Chemiluminescent Substrate (Pierce).
3
Evaluating AKT and MEK Signaling
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Anti-SCOP/PHLPP1 (COSMO Bio; Carlsbad, CA, USA), anti-pAKT473, anti-pAKT308, anti-AKT Total, anti-pMEK, anti-MEK Total, anti-α-Tubulin (Cell Signaling Technology; Danvers, MA, USA).
4
Protein Expression Analysis in VSMCs
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The protein levels of Rac1, MYH, SM22α, Akt, p-Akt (308) and p-Akt (473) in VSMCs were analysed by Western blot. Cells were lysed in RIPA Lysis Buffer (Beyotime Institute of Biotechnology, China), and protein was quantified using BCA protein assay reagent (Beyotime). Proteins (25 µg) were resolved on 6% or 10% SDS-polyacrylamide gels and transferred to polyvinylidene fluoride membranes, which were blocked with 5% BSA. The primary antibodies anti-Rac1/2/3(1:1000), anti-p-Akt (308; 1:1000), anti-p-Akt (473; 1:1000) (all from Cell Signaling Technology, USA), anti-MYH (1:500), anti-SM22α (1:1000) and anti-TRPC1 (1:1000) (all from Santa Cruz, USA) were incubated with polyvinylidene fluoride membranes overnight at 4°C. After successive washes, membranes were incubated for 1 h with HRP-conjugated sheep anti-rabbit IgG (1:1000) and horse anti-mouse IgG (1:1000) at room temperature (Zhongshan Golden Bridge Biotechnology Company, China). Enhanced chemiluminescence developing methods (Beyotime) were used to visualize immunolabelling. Image-Pro Plus was used to take measurements of band density.
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