Plumbagin (5-hydroxy-2-methyl-1,4-naphthoquinone) was purchased from Sigma-Aldrich (St. Louis, MO, USA). Cisplatin was purchased from Qilu Pharm (Jinan, China). 3-Methyladenine (3-MA) and N-acetylcysteine (NAC) were obtained from MedChem Express (Shanghai, China). Fetal bovine serum (FBS) and Dulbecco's modified Eagle's medium (DMEM) high glucose were purchased from Gibco (Carlsbad, CA, USA). The antibodies used included Bax (5023), Bcl-2 (4223), cleaved caspase-3 (9664), Beclin-1 (3495), LC3B (3868), SQSTM1/p62 (39749), SAPK/JNK (9252), phospho-SAPK/JNK (Thr 183/Try 185, 4668), phospho-AKT (Ser 473, 4060), and phospho-mTOR (Ser 2448, 5536), all of which were acquired from Cell Signaling Technology (Danvers, MA, USA). Mouse anti-human GAPDH, rabbit anti-human β-actin, and secondary antibodies including goat anti-rabbit and goat anti-mouse were purchased from Proteintech (Wuhan, China).
Mouse anti human gapdh
Manufactured by Proteintech
Sourced in China, United States
The Mouse anti-human GAPDH is a primary antibody that recognizes the GAPDH protein, which is a widely expressed and constitutively active enzyme involved in glycolysis. This antibody can be used in various immunodetection techniques to identify and quantify the GAPDH protein in human samples.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Rabbit anti human β actin, by Proteintech (2 mentions)
3 methyladenine 3 ma, by MedChemExpress (1 mentions)
N acetylcysteine, by MedChemExpress (1 mentions)
Plumbagin, by Merck Group (1 mentions)
High glucose dulbecco s modified eagle s medium dmem, by Thermo Fisher Scientific (1 mentions)
Cd105, by Abcam (1 mentions)
Supersignal west pico plus chemiluminescent substrate, by Thermo Fisher Scientific (1 mentions)
Anti oct4, by Cell Signaling Technology (1 mentions)
Mouse anti human β actin monoclonal antibody, by Merck Group (1 mentions)
Cd144, by Cell Signaling Technology (1 mentions)
Hrp 60004, by Proteintech (1 mentions)
Ripa lysis buffer, by Beyotime (1 mentions)
Phenyl methyl sulfonyl fluoride (pmsf), by Beyotime (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
Glutamine, by Thermo Fisher Scientific (1 mentions)
Cycloheximide sc 3508, by Santa Cruz Biotechnology (1 mentions)
Polyvinylidene fluoride (pvdf), by Bio-Rad (1 mentions)
Enhanced chemiluminescent substrate, by Bio-Rad (1 mentions)
6 protocols using mouse anti human gapdh
1
Plumbagin Modulates Cisplatin-Induced Apoptosis
2
Western Blot Protein Expression Analysis
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Western blotting (WB) was carried out routinely. The primary antibodies used here included anti-Oct4 (1:1000, #2750; Cell Signaling), CD144 (1:1000, #2500; Cell Signaling), CD105 (1:1000, ab11414; Abcam), CD31 (1:1000, #3528; Cell Signaling), mouse anti-human GAPDH (1:10000, HRP-60004; proteintech) and mouse anti-human β-actin monoclonal antibody (1:10000, A5316; Sigma). Protein bands were visualized with SuperSignalTM West Pico PLUS Chemiluminescent Substrate (#34580; Thermo).
3
Quantitative Protein Analysis of HK-2 Cells
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After co-culturing for 2, 4, and 6 h, total protein was, respectively, extracted from cultured HK-2 cells only in H, H + A and H + M + A group using the RIPA Lysis Buffer (Beyotime) containing 2% PMSF (Beyotime) according to the manufacturer’s instructions, and then collected protein samples and subjected to western blotting performed essentially according to an established procedure [23 (link)]. The primary antibodies used were as follows: mouse anti-human GAPDH (1:10,000, ProteinTech, Chicago, IL, USA), mouse anti-human Fetuin-A, and Anti-Osteopontin (1:2000, Abcam, Cambridge, MA, USA), and BCL-2 (1:2000, D55G8) Rabbit mAb, BAX Antibody (1:2000). We performed quantifications by measuring band intensities using AlphaView and Image lab analysis software.
4
Cell Line and Antibody Analysis
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Cycloheximide (sc-3508), Rabbit anti-FOXK1 (H-140), E-cadherin (H-108) and mouse immunoglobulin G (nm IgG) were purchased from Santa Cruz (Santa-Cruz, CA, USA). Rabbit anti-Vimentin (Ag0489), MMP2 (Ag0549) and MMP9 (Ag0552), and mouse anti-human GAPDH were purchased from Proteintech (Wuhan, China). Rabbit anti-Erk1/2 (137F5), p- Erk1/2 (Thr202/Tyr204), Akt (C-terminal), and p-Akt (Ser473) were purchased from CST (CST, MA, USA). Anti-RIPX (Anti-RUFY3, ab89147) was purchased from Abcam (Cambridge, UK). LoVo and SW1116 cells were grown in RPMI 1640 containing 10% fetal bovine serum (Life Technologies, Monza, Italy), 1% glutamine (Life Technologies) and 1% penicillin/streptomycin (Life Technologies) in a humidified incubator at 37 °C with an atmosphere of 5% CO2.
5
Western Blot Analysis of Protein Expression
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Western blotting procedures were performed following a standard protocol. The protein was solubilized with loading buffer (5×) and heated at 100°C for 10 min. Proteins were separated by SDS-PAGE using 10% gels and transferred to a polyvinylidene fluoride membrane (PVDF, Bio-Rad Laboratories Inc, California, USA). The membranes were incubated overnight at 4°C with primary antibodies: rabbit anti-human CD63 (1:500, SBI, USA), rabbit anti-human β-actin (1:5000, Proteintech, USA), mouse anti-human GAPDH (1:2000, Proteintech, USA), and rabbit anti-human ephrinA2 (1:200, DF3027, Affinity Biosciences, USA) followed by horseradish peroxidase-conjugated secondary anti-rabbit and anti-mouse (1:2000, CoWin Biosciences, China) for 2 h. The membranes were visualized with enhanced chemiluminescent substrate (Bio-Rad Laboratories Inc, California, USA).
6
Western Blot Analysis of Peroxisomal Proteins
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Fibroblasts were lysed in RIPA buffer containing protease inhibitors (EDTA-free cOmplete tablets, Roche, Basel, Switzerland). Next, samples were mixed with 5× loading buffer, and proteins were separated on 10% polyacrylamide gel by SDS-PAGE. Separated proteins were transferred to a nitrocellulose membrane using semidry transfer (BioRad transfer system), followed by blocking with 4% non-fat milk powder. The following primary monoclonal antibodies were applied overnight at 4 °C to detect selected proteins: mouse anti-human ABCD1 (Euromedex ALD-1D6-AS, clone 2AL-1D6, 1:6000), mouse anti-human GAPDH (Proteintech, 1:50,000), mouse anti-human PLIN2 (Progen, 1:200), mouse anti-human β-actin (Chemicon, 1:500,000). The secondary antibody, goat anti-mouse conjugated with horseradish peroxidase (Dako, 1:20,000), was added for 1 h at room temperature. Chemiluminescent HRP Substrate (Immobilon®Western, Milipore, Burlington, MA, USA) was used for the detection of chemiluminescent signal using the Chemidoc XRS detection system (Bio-Rad). Quantitative analysis was completed using Image Lab 6.1 software (Bio-Rad).
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