Ferrostatin-1 (347174-05-4), liproxstatin-1 (950455-15-9), trolox (53188-07-1), deferoxamine (138-14-7), RSL-3 (1219810-16-8), erastin (571203-78-6), sulfasalazine (599-79-1), atorvastatin (599-79-1), KU60019 (925701-46-8) were purchased from Cayman. The concentration used is shown in Supplementary Table. S1 . C11BODIPY (D3861) was purchased from Invitrogen. Mouse recombinant IFNγ (485-MI-100), anti-mouse IFNγ block antibody (MAB485), human recombinant IFNγ (285-IF-100) were purchased from R&D. Anti-mouse IFNGR1 antibody (16-1193-85) was purchased from Thermo Fisher. The following antibody were used for immunoblots: anti-human SLC7A11 antibody (12691), anti-β-actin (12262), anti-GAPDH (2118), anti-STAT1 (9172), anti-phospho-STAT1 (9167), anti-ATM (2873) and anti-ACSL4 (4047) were bought from Cell Signaling Technology. Anti-GPX4 (ab41787) was from Abcam. Additionally, IgG (BE0087, BE0090), anti-mouse CTLA4 (BE0131), anti-mouse PD-L1 (BE0101) and anti-mouse CD8 (BE0117) were purchased from BioXcell. 4-HNE antibody (ab46545) was purchased from Abcam. Cyst(e)inase was generated as previously described (18 (link)).
Anti acsl4
Manufactured by Cell Signaling Technology
Sourced in United States
Anti-ACSL4 is a primary antibody product produced by Cell Signaling Technology. It is designed to detect the ACSL4 protein, which is involved in long-chain fatty acid activation.
Automatically generated - may contain errors
Lab products found in correlation
Anti gpx4, by Abcam (2 mentions)
4 hne antibody ab46545, by Abcam (1 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Goat anti rabbit igg antibody, by Cell Signaling Technology (1 mentions)
Ecl western blotting detection system, by Merck Group (1 mentions)
Anti 4 hydroxynonenal, by Abcam (1 mentions)
Anti actin, by Cell Signaling Technology (1 mentions)
Bca assay kit, by Beyotime (1 mentions)
Radioimmunoprecipitation assay (ripa), by Beyotime (1 mentions)
Ab46545, by Abcam (1 mentions)
Polyvinylidene difluoride membrane, by Merck Group (1 mentions)
Anti slc7a11, by Cell Signaling Technology (1 mentions)
Anti p ampk, by Cell Signaling Technology (1 mentions)
Anti sod1, by Cell Signaling Technology (1 mentions)
Anti sod2, by Cell Signaling Technology (1 mentions)
Anti klf4, by Cell Signaling Technology (1 mentions)
Anti ho 1, by Cell Signaling Technology (1 mentions)
Anti ampk, by Cell Signaling Technology (1 mentions)
Anti β actin, by Cell Signaling Technology (1 mentions)
Bicinchoninic acid protein assay kit, by Beyotime (1 mentions)
3 protocols using anti acsl4
1
Ferroptosis-Inhibition Compound Evaluation
2
Western Blot Analysis of Ferroptosis Markers
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Tissues and Cells were lysed in the lysis buffer (RIPA, Beyotime, China). The protein concentrations were measured by the BCA assay kit (Beyotime, China). Equal amounts of protein were loaded in 10% sodium dodecyl sulfate-polyacrylamide gel (SDS-PAGE) and were then transferred from the gel to polyvinylidene fluoride membranes (Millipore, USA). After blocking in a solution of 5% bovine serum albumin (BSA, Solarbio, China) for 1 h, the membranes were washed with TBST and then incubated with primary antibodies overnight at 4 °C. The following antibodies were used: anti-GPX4(1:2000), anti-4 Hydroxynonenal(1:1000, ab46545, Abcam, USA), anti-ACSL4(1:2000), anti-actin(1:2000, 4967, Cell Signaling Technology, USA). After washing, the membranes were incubated with horseradish peroxidase (HRP)-conjugated secondary antibodies for 1 h at 37 °C: goat anti-rabbit IgG antibody (1:2000, #7074, Cell Signaling Technology, USA). Bound antibodies were detected using the ECL western blotting detection system (Millipore, USA).
3
Molecular Mechanisms of HUVEC Responses
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After different treatments, the HUVECs were washed twice with cold PBS. Total cellular proteins were extracted using radioimmunoprecipitation assay buffer (RIPA) buffer. Protein concentrations were determined using a bicinchoninic acid protein assay kit (Beyotime Institute of Biotechnology). Equal amounts of proteins were separated using 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The separated proteins were transferred onto polyvinylidene difluoride membranes (Millipore, Billerica, MA, USA). Then, the membranes were blocked with 5% skimmed milk in TBST at room temperature for 1 h. The blots were incubated at 4°C overnight with the following primary antibodies from Abcam: anti-GPX4 (1: 5000), anti-SLC7A11 (1: 1000), anti-ACSL4 (1: 10000), anti-HO-1 (1: 2000), anti-KLF4 (1: 1000), and anti-β-actin (1: 1000), and with the following primary antibodies from Cell Signaling Technology: anti-p-AMPK (1: 1000), anti-AMPK (1: 1000), anti-SOD-1(1: 1000), and anti-SOD-2 (1: 1000). Then, the blots were incubated with HRP-conjugated secondary antibodies (1: 5000) for 1 h at room temperature. Subsequently, blots were developed using enhanced chemiluminescence (Model 6600; Tanon, Shanghai, China).
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