The rats were randomly divided into 7 groups (8 rats per group, survived after treatment for 24 h) as follows: (I) the sham (control) group was operated with no ischemia; (II) the MCAO group was operated with MCAO; (III) the ulinastatin group was operated with MCAO and given ulinastatin at a dose of 300,000 U/kg (AmyJet Scientific Inc., Wuhan, China) (17 (link)); (IV) the ML-385 group was administered ML-385 (50 pmol/5 µL; AOBIOUS, MA, USA) and operated with MCAO (18 (link)); (V) the ulinastatin + ML-385 group was administered ulinastatin + ML-385 and then operated with MCAO; (VI) the Tert-butylhydroquinone (TBHQ) group was administered TBHQ (50 mg/kg; Sigma Chemical Co., St. Louis, MO, USA) and operated with MCAO (18 (link)); (VII) the ulinastatin + TBHQ group was administered ulinastatin + TBHQ and then operated with MCAO. ulinastatin was stored at 4 °C and dissolved in 0.9% saline upon use. ulinastatin was injected intraperitoneally 15 min after MCAO (17 (link)). TBHQ and ML-385 were diluted in dimethyl sulfoxide (DMSO) before intracerebroventricular (i.c.v.) injection and administered 24 hours before MCAO (18 (link)). Upon completion of the above experiments, the blood and brain tissue of each rat was collected for analysis.
Ml385
Manufactured by Merck Group
Sourced in United States, Germany
The ML385 is a laboratory instrument designed for performing various analytical and scientific procedures. It is a versatile and reliable piece of equipment that can be used in a wide range of research and testing applications. The core function of the ML385 is to provide accurate and consistent measurements, enabling researchers and scientists to gather reliable data for their studies.
Automatically generated - may contain errors
Lab products found in correlation
N acetylcysteine nac, by Merck Group (2 mentions)
Dimethyl fumarate, by Merck Group (2 mentions)
Lipopolysaccharides (lps), by Merck Group (2 mentions)
Antimycin a, by Merck Group (2 mentions)
Fluorescent mounting media, by Solarbio (1 mentions)
Double antibiotics, by Solarbio (1 mentions)
Mtt kit, by Solarbio (1 mentions)
Nuclear and cytoplasmic protein extraction kit, by Solarbio (1 mentions)
Sulforaphane sfn, by Merck Group (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Solarbio (1 mentions)
Ros assay kit, by Solarbio (1 mentions)
Glutathione (gsh), by Merck Group (1 mentions)
Mitoquinone mesylate, by MedChemExpress (1 mentions)
Macs microbead, by Miltenyi Biotec (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
L glutamine, by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions)
Phorbol myristate acetate (pma), by Merck Group (1 mentions)
Isoproterenol, by Merck Group (1 mentions)
32 protocols using ml385
1
Neuroprotective Agents in Ischemic Stroke
2
Nrf2 Inhibitor Evaluation in Oxidative Stress
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3,4-DHAP was purchased from Jinan Luxin Chemical Technology Co., Ltd. Sulforaphane (SFN), an antioxidant reagent that was used as a positive control, was obtained from Sigma-Aldrich; Merck KGaA. ML385, a novel and specific Nrf2 inhibitor, was purchased from Selleck Chemicals. ML385 is an inhibitor of Nrf2 activity and nuclear translocation, which has been confirmed to affect the expression of downstream genes. The ROS assay kit, Nuclear and Cytoplasmic Protein Extraction Kit, double antibiotics, MTT kit, DAPI and Fluorescent Mounting Media were obtained from Beijing Solarbio Science & Technology Co., Ltd.
3
Isolation and Culture of T-cell Subsets
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Human CD4+ and CD8+ T cells were isolated from the PBMCs of patients with RRMS and healthy donors by immunomagnetic separation using CD4+ or CD8+-labelled magnetic-activated cell sorting (MACS) microbeads (Miltenyi Biotec), respectively. For the purification of the T-cell subsets [memory (TM) and naïve (TN)], CD4+ or CD8+ T cells were enriched using a negative selection technique (Miltenyi Biotec) and subsequently TM cells (CD45RO+) and TN cells (CD45RO−) were isolated using CD45RO+-labelled MACS microbeads. All T cells were cultured in X-VIVOTM Media 15 (Lonza). When indicated, 5 µg/ml DMF, 5 µg/ml monomethyl fumarate, 250 nM N-acetylcysteine (NAC), 250 µM glutathione (all from Merck) and 100 µM mitoquinone mesylate (MitoQ; MedChem Express) were used. Sulphoraphane (500 nM, Merck) was applied for pharmacological activation of Nrf2.23 (link) For pharmacological inhibition of Nrf2, 5 µg/ml ML385 (Merck) was used.24 (link) Buthionine sulphoximine (3 mM, Merck) was utilized as a specific inhibitor of glutamate cysteine ligase, the rate-limiting step in glutathione synthesis.25 (link) For a detailed description see the Supplementary material .
4
Macrophage Differentiation and Ox-LDL Exposure
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THP-1 monocyte-like cells (American Type Culture Collection) were cultured in RPMI-1640 medium supplemented with 10% FBS, 2 mM L-glutamine and 1% penicillin/streptomycin (all Gibco) at 37°C with 5% CO2. THP-1 cells were differentiated into macrophages by exposure to 100 nM PMA (Sigma-Aldrich) as previously described [21 (link)] for indicated hours. Then cells were incubated with 50 μg/ml ox-LDL for 24 h. For NRF2 inhibition, 2 μM ML385 (Sigma-Aldrich) was used to treat cells for 24 h.
5
Ginsenoside Rc Regulates Oxidative Stress
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Ginsenoside Rc was purchased from Yuanye Bio-technology Co., Ltd (Lot number: M27GB141849, purity > 98%). Isoproterenol and ML385 (Nrf2 inhibitor) were bought from Sigma Co. (St. Louis, MO, USA). Carboxymethyl cellulose sodium salt (CMC-Na) was from Shanghai Jinshan Chemical Co., Ltd (Shanghai, China). Creatine kinase (CK-MB, Lot number: 20210518), malondialdehyde (MDA, Lot number: 20210621) and glutathione (GSH, Lot number: 20210624) biochemical assay kits were obtained from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). TNF-α (Lot number: M21015903) and IL-1β (Lot number: V21013165) ELISA kits were from Wuhan Huamei Biotech Co., Ltd (Wuhan, China). Troponin T ELISA kit (Lot number: 20210618) was purchased from Solarbio Science & Technology Co., Ltd (Beijing, China). Nuclear and cytoplasmic protein extraction kit, BCA protein kit and ECL detection reagents were from Beyotime Institute of Biotechnology (Shanghai, China). Other reagents were of commercially analytical grade.
6
Modulation of Lymphocyte Oxidative Stress
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Lymphocytes were pretreated for 18 hours with Auranofin (AUF, with the concentration 1 µg/mL for NK cells and 0,5 µg/mL for TIL and CAR T cells, if not indicated differently), DL-sulforaphane (SUL) or dimethyl fumarate (DMF) (all Sigma-Aldrich) at indicated concentrations. As all compounds were reconstituted in DMSO, control NK cells were pretreated with DMSO with an equivalent volume to the highest compound concentration. 100 U/mL Catalase (Sigma-Aldrich) were added to control lymphocytes just before exposure to oxidative stress. For N-acetylcysteine (NAC, Invitrogen) treatment, DMSO pretreated cells were incubated for 1 hour with 5 or 10 mM NAC prior to H2O2 treatment. For Nrf2 inhibition, NK cells were pretreated with 50 µM ML-385 (Sigma-Aldrich) in parallel to AUF. For H2O2 treatment, lymphocytes were washed and resuspended to 1×106 cells/mL in RPMI containing the indicated H2O2 (Sigma-Aldrich) concentration for 1 hour at 37°C 5% CO2. Cells were washed with medium and then used for further experiments.
7
Chitosan Oligosaccharide Attenuates Liver Injury
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D‐gal and ML385 were obtained from Sigma‐Aldrich Chemical Co. COS (average molecular weight <1,000, DD. 91.3%) was obtained from Dalian Glycobio Co., Ltd. A certain amount of COS was weighed accurately and dissolved in ultrapure water at room temperature to prepare two different concentrations: 15 and 60 mg/ml. Liver function biomarkers assay kits were obtained from BioVision (Cat. No: K553, E4320, and E4324, Inc.). The MTT cell assay kit, oxidative stress, and antioxidant biomarkers assay kits were obtained from Nanjing Jiancheng Bioengineering Institute: MTT (cat. no. G020‐1‐1), AGEs (cat. no. abx512406), 8‐OH‐dG (cat. no. SKT‐120‐480), ROS (cat. no. E004‐1‐1), GSH‐Px (cat. no. A005‐1‐2), SOD (cat. no. A001‐3‐2), and CAT (cat. no. A007‐1‐1). The following antibodies were used: GAPDH (#5174) and Nrf2 (#12721) (Cell Signaling Technology). All other chemicals and reagents used in the study were purchased from Sigma‐Aldrich.
8
Exogenous FGF10 Modulates Inflammation and Oxidative Stress
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The standard reference airborne (1649b) was purchased from NIST (Gaithersburg, MD, USA). For in vivo animal experiments, exogenous recombinant FGF10 was obtained from the School of Pharmaceutical Science affiliated with Wenzhou Medical University. For in vitro cell experiments, exogenous recombinant FGF10 was purchased from PeproTech (Shanghai, China). Antibodies against FGF10, phospho-p65, p65, phospho-IκBα, IκBα, Nrf2, Kelch-like ECH-associated protein 1 (KEAP1), heme oxygenase 1 (HO-1), and Quinone Oxidoreductase 1 (NQO1) were purchased from Cell Signaling Technology (Beverly, MA, USA). The enzyme linked immunosorbent assay (ELISA) kits for mouse FGF10, mouse IL-6, mouse IL-8, human IL-6, and human IL-8 were purchased from Boyun Biotechnology (Shanghai, China). The compound BAY11-7082 was purchased from Selleck (Houston, Texas, USA). N-acetylcysteine (NAC) and ML385 were purchased from Sigma-Aldrich (Shanghai, China).
9
Fumarate and Maleate Compound Assays
10
Antibody Validation for ER Stress Pathway
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Tun, thapsigargin, ML385 and common reagents were from Sigma. GSK606414 was from APExBio and CCT020312 from Calbiochem. Mouse anti-actin and rabbit anti-calnexin antibodies were from Sigma. Mouse anti-eIF2α was from Cell Signaling. Rabbit anti-phospho-eIF2α (Ser51) was from MBL. Rabbit anti-PERK was from Abcam. Rabbit anti-ATF4 was a kind gift from David Ron (Univ. Cambridge). Goat anti-mouse IgG conjugated to 488 DyeLight, goat anti-rabbit IgG conjugated to 564 DyeLight, and goat anti-rabbit and anti-mouse IgG conjugated to HRP were from Jackson Labs. Normal goat serum was from Vector Laboratories (Burlingame, CA).
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