D arginine
Manufactured by Merck Group
Sourced in United States
D-arginine is a lab equipment product manufactured by Merck Group. It is a naturally occurring amino acid that serves as a precursor for the synthesis of various biomolecules in living organisms. D-arginine is commonly used in biochemical and pharmaceutical research applications.
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Lab products found in correlation
L arginine, by Merck Group (7 mentions)
Glycine, by Merck Group (3 mentions)
L citrulline, by Merck Group (3 mentions)
D proline, by Merck Group (2 mentions)
D alanine, by Merck Group (2 mentions)
D methionine, by Merck Group (2 mentions)
D tyrosine, by Merck Group (2 mentions)
D histidine, by Merck Group (2 mentions)
D isoleucine, by Merck Group (2 mentions)
D leucine, by Merck Group (2 mentions)
D phenylalanine, by Merck Group (2 mentions)
D threonine, by Merck Group (2 mentions)
D tryptophan, by Merck Group (2 mentions)
D valine, by Merck Group (2 mentions)
Sodium chloride (nacl), by Merck Group (2 mentions)
Vinblastine, by Selleck Chemicals (2 mentions)
Sorafenib, by Selleck Chemicals (2 mentions)
Bazedoxifene, by Selleck Chemicals (2 mentions)
Camptothecin, by Selleck Chemicals (2 mentions)
Rapamycin, by Thermo Fisher Scientific (2 mentions)
13 protocols using d arginine
1
Amino Acid Synthesis and Characterization
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d -Alanine, d -Arginine, d -Asparticacid, d -Asparagine, d -Glutamicacid, d -Histidine, d -Isoleucine, d -Leucine, d -Lysine, d -Methionine, d -Phenylalanine, d -Proline, d -Serine, d -Threonine, d -Tryptophan, d -Tyrosine, d -Valine, KBr (spectral grade), bovine serum albumin (BSA), glutathione and 3,3′,5,5′-tetramethylbenzidine (TMB) were purchased from Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). d -Cystine and d -Glutarnine were purchased from Aladdin Reagent Co, Ltd. (Shanghai, China). All other chemicals and reagents were of analytical grade. All aqueous solutions were prepared with Milli-Q water.
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2
Apoptosis and Cell Cycle Analysis of Fibroblast-Like Synoviocytes
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RA FLSs in the logarithmic growth phase were plated at 2 × 105 cells/well in 6-well plates and cultured under normoxic or hypoxic conditions for 24 h. D-Arginine (Sigma-Aldrich, MO, USA) at different concentrations (2 mM, 4 mM, 8 mM, 16 mM, and 32 mM) was added, and the control was set at the same time. After culturing for 48 h, the cells were collected and washed twice with PBS. Each sample was stained with 5 μL Annexin V and 5 μL PI solution (BD Bioscience, NJ, USA) at 4℃ for 30 min. The apoptosis rate of FLSs was detected using a BD Aria II flow cytometer and analyzed using Diva software.
Cells were collected using the same method described above, and 1 × 105 FLSs were fixed in 1 mL 70% ethanol solution at 4℃ overnight. Next, the samples were centrifuged at 1000 rpm for 5 min, and the fixation solution was removed. After washing with precooled PBS 3 times, 100 μL of 100 μg/mL RNase (BD Bioscience, NJ, USA) was added to each tube. The samples were incubated with RNase at 37℃ for 30 min to degrade RNA, and then 100 μL of 50 μg/mL PI (BD Bioscience, NJ, USA) staining solution was added and incubated at 4℃ for 30 min. The cell cycle was detected using a BD FACSCalibur flow cytometer, and results were analyzed using ModFit software.
Cells were collected using the same method described above, and 1 × 105 FLSs were fixed in 1 mL 70% ethanol solution at 4℃ overnight. Next, the samples were centrifuged at 1000 rpm for 5 min, and the fixation solution was removed. After washing with precooled PBS 3 times, 100 μL of 100 μg/mL RNase (BD Bioscience, NJ, USA) was added to each tube. The samples were incubated with RNase at 37℃ for 30 min to degrade RNA, and then 100 μL of 50 μg/mL PI (BD Bioscience, NJ, USA) staining solution was added and incubated at 4℃ for 30 min. The cell cycle was detected using a BD FACSCalibur flow cytometer, and results were analyzed using ModFit software.
3
Cell Line Identification and Culture
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The TNBC cell lines MDA-MB-231 and MDA-MB-468 and the NSCLC cell line H460 were purchased from the American Type Culture Collection (ATCC). The MDA-MB-231-BrM2 cell line was obtained from J. Massague (Memorial Sloan Kettering Cancer Center, New York, NY). The mouse TNBC cell line 4T1 was obtained from Vivek Mittal (Weill Cornell Medicine, New York, NY). Identification was done once a year by genotyping, and mycoplasma was tested quarterly by polymerase chain reaction (PCR). Cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) (human and murine TNBC cells) or RPMI 1640 medium (NSCLC cells) supplemented with fetal calf serum and antibiotics (penicillin and streptomycin) or with a low-buffered DMEM medium with no Hepes and supplemented with 1 mM phosphate buffer (pH 7.2) in certain experiments. l -Arginine and d -arginine were purchased from Sigma-Aldrich and dissolved in RPMI 1640 (adjusting the final pH to 6.8 to 7.2). The competitive NOS inhibitor NG-monomethyl-l -arginine and the NOS2 selective inhibitor 1400W dihydrochloride were purchased from Sigma Chemical and Enzo Life Science, respectively. Compound solutions were prepared fresh for each experiment.
4
Cellular Adhesion and Imaging Protocol
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The following chemicals were purchased from Sigma and used without further purification: d -arginine, glycine, l -aspartic acid, NaCl, Hoechst (bisbenzimide H33342 trihydrochloride) and indocyanine green (ICG). NHS-PEG, Mw 750. NHS-PEG, Mw 5000. HUVEC cells and their EBM-2 medium were purchased from Lonza. PBS and XTT kit were ordered form Biological Industries.
5
Amino Acid Analogs and Inhibitors
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L-arginine, D-arginine, Pentamidine, Canavanine, N-Methyl L-arginine acetate (NMLAA), Nω-Nitro-L-arginine methyl ester (L-NAME), Nω-Nitro-L-arginine (L-NNA), L-citrulline, 3-Ureidopropionic acid and 4-{[5-(4-aminophenoxy)pentyl]oxy}phenylamine were obtained from Sigma-Aldrich, USA. All other materials used in this study were of analytical grade and commercially available.
6
Compound Library Screening for Bioactive Inhibitors
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An 1,833-member bioactive compound library, and an independent 86-member PI3K signaling inhibitor library comprising mTOR, PI3K and AKT pathway inhibitors, were obtained from Selleck Chemicals and stored at −80°C. Erastin2, annotated as compound 35MEW28 (Ref.51 (link)) and ML162 were synthesized by Acme Bioscience. Erastin was the kind gift of B. Stockwell (Columbia). Chemical used (supplier) were: DMSO, ferrostatin-1, thapsigargin, tunicamycin, cycloheximide, L-arginine, D-arginine and L-citrulline (Sigma-Aldrich Corporation); bortezomib, rapamycin, etoposide and buthionine sulfoximine (Thermo-Fisher Scientific); INK 128, AZD8055, vinblastine, camptothecin, sorafenib, bazedoxifene, raloxifene and JTC-801 (Selleck Chemicals); and GCN2iB (MedChemExpress). buthionine sulfoximine was dissolved directly into cell media. All other drugs were prepared as stock solutions in DMSO. Stock solutions were stored at −20˚C.
7
Conjugation of TRAIL with PEG-Maleimide
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NHS-PEG (3500 MW) maleimide ester was purchased from Advanced BioChemicals (Lawrenceville, GA, USA). The following chemicals were obtained from Sigma and used without further purification: D-arginine, L-arginine, glycine, NaCl, bicarbonate buffer (BB, 0.1 M), 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), N-hydroxysulfosuccinimide (sulfo-NHS) and 2-morpholinoethanesulfonic acid (MES, pH = 6). Doxorubicin HCl was purchased from Tzamal D-Chem laboratories (Petah-Tikva, Israel), and human TRAIL was acquired from Peprotech (Rehovot, Israel). CAOV-3 ovarian cell lines were purchased from ATCC (Manassas, VA, USA) and their medium was purchased from Lonza (Basel, Switzerland). PBS (0.01 M) and XTT kits were obtained from Biological Industries (Beit Haemek, Israel). Double distilled water (DDW) was purified by passing deionized water through an Elgastat Spectrum reverse osmosis system (Elga Ltd., High Wycombe, UK). An ELISA kit for TRAIL was purchased from Biotest Ltd. (Beit Haemek, Israel).
8
Amino Acid Synthesis and Purification
9
T Cell Proliferation Assay with CFSE
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PBMC isolated after Ficoll density gradient were labeled with carboxyfluorescein succinimidyl ester (CFSE, 200 nM; interchim, Montluçon, France). The quantification of T cells was obtained by staining PBMC with an anti-CD3 APC antibody (Becton Dickinson). PBMC were seeded in 96-well round-bottom plates at a concentration of 1 × 105 T cells per well. Cells were cultured in RPMI 1640 supplemented with 10% human AB serum (Biowest, Nuaillé, France) and anti-CD3 and anti-CD28 monoclonal antibodies (0.6 μg/mL, Sanquin, Amsterdam, Netherlands). When indicated, cells were cultured in presence of the following chemical inhibitors or their controls: (i) L-arginine (1 mM, Sigma-Aldrich, St Louis, MO, USA) or with control D-arginine; (ii) coptisine (an IDO inhibitor) (50 nM, Sigma-Aldrich, St Louis, MO, USA) or vehicle; and (iii) PD-L1 blocking antibodies (10 μM, ebioscience, San Diego, CA, USA) or IgG. After 4 days of culture, cells were harvested and labeled with FVS 780 (Becton Dickinson) for cell viability, CD2 PC7 and CD8 APC (Beckman Coulter), CD4 BUV496 and CD14 BV605 (Becton Dickinson). CFSE dilution was assessed on FVSneg viable T cells by flow cytometry on Fortessa X20 (Becton Dickinson) and results were analyzed with ModFit LT software (Verity Software, Topsham, ME).
10
Preparation of Amino Acid Solutions
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D-methionine, D-tryptophan, D-leucine, D- or L-cysteine, D- or L-aspartic acid, D- or L-glutamic acid, D-alanine, D-threonine, D-valine, D-isoleucine, D-glutamine, D-phenylalanine, D-histidine, D-proline, D-lysine, D-arginine, and glycine (Sigma-Aldrich, St. Louis, MO, USA) were all prepared at a concentration of 40 mM, and D-tyrosine was prepared at a concentration of 0.8 mM due to its low solubility. Nisin powder (2.5% purity, Sigma-Aldrich, St. Louis, MO, USA) was dissolved in a solution at pH 2 to obtain a stock solution with a concentration of 1 g/L.
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