Dulbecco s modified eagle s medium dmem culture medium

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Dulbecco's modified Eagle's medium (DMEM) is a cell culture medium designed to support the growth of a wide variety of cell types. It provides a balanced salt solution, amino acids, vitamins, and other nutrients essential for cell growth and maintenance.

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10 protocols using dulbecco s modified eagle s medium dmem culture medium

Fluorenylmethyloxy carbonyl peptide synthesis

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The fluorenylmethyloxy carbonyl(Fmoc) amino acids and the Fmoc-Lys(Boc)-Wang Resin were purchased from AAPPTec (Louisville, KY, USA). All the other chemicals were purchased from Sigma-Aldrich (St. Louis, MO, USA) and used without further purification. Chlorin e6 (Ce6) was purchased from Frontier-Scientific (Logan, UT, USA). Water was obtained from Millipore Q3 system (Billerica, MA, USA).
Dulbecco’s modified Eagle’s medium (DMEM) culture medium, fetal calf serum, and penicillin–streptomycin were purchased from Gibco (Life Technologies, NY, USA). Lysotracker green, mitotracker green, ER tracker green, HPF, Amplex Red assay, and Calcein AM were purchased from Thermo Fisher Scientific.

Gilson R.C., Tang R., Gautam K.S., Grabowska D, & Achilefu S. (2019). Trafficking of a Single Photosensitizing Molecule to Different Intracellular Organelles Demonstrates Effective Hydroxyl Radical-Mediated Photodynamic Therapy in the Endoplasmic Reticulum. Bioconjugate chemistry, 30(5), 1451-1458.

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Culturing Epidermoid and Breast Cancer Cells

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Dulbecco’s modified Eagle’s medium (DMEM) culture medium, fetal calf serum, and penicillin-streptomycin were purchased from Gibco, (Life Technologies, NY, USA). Vectashield was purchased from Vector Laboratories (CA, USA).
Human epidermoid cancer (A431) cells were obtained from ATCC (VA, USA) and 4T1/luc murine breast cancer cells were obtained from David Piwnica-Worms, Washington University in St. Louis.^{27 (link)} Cells were cultured using DMEM supplemented with 10% FBS, and 1% penicillin streptomycin in a humidified incubator at 5% CO₂ balanced with air at 37°C. The cells were kept at 50–80% confluence.
Female BALB/c and female athymic NCr-nu/nu mice were obtained from Charles River Laboratories (MA, USA). Female PyMT mice were bred in-house. All animal studies were performed with accordance with protocols approved by the Washington University School of Medicine Animal Studies Committee.

Gilson R.C., Tang R., Som A., Klajer C., Sarder P., Sudlow G.P., Akers W.J, & Achilefu S. (2015). Protonation and Trapping of a Small pH-Sensitive Near-Infrared Fluorescent Molecule in the Acidic Tumor Environment Delineate Diverse Tumors In Vivo. Molecular pharmaceutics, 12(12), 4237-4246.

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Engineered IFN-γ and OmpA Peptide Protocol

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The IFN-γ gene and OmpA peptide gene sequence was obtained from GenBank and synthesized by Sangon bioengineering Co.(Shanghai, China). The pBV220 plasmid was purchased from Miaoling Bioscience & Technology Co.(Wuhan, China). E. coli DH5α strain and E. coli MG1655 strain were purchased from Angyubio Biotechnology Co. (Shanghai, China). The mouse breast cancer cells (4T1) were obtained from American Type Culture Collection. Dulbecco’s modified Eagle’s medium (DMEM) culture medium, fetal bovine serum (FBS), penicillin and streptomycin solution, trypsin containing 0.25% EDTA were purchased from Gibco (USA). 1,1-dioctadecyl-3,3,3,3-tetramethylindotricarbocyanine iodide (DiR), live & dead cell viability detection kit and CCK-8 assay kit were obtained from Beyotime Biotechnology Co. (Shanghai, China). Murine IFN-γ, TNF-α, IL-10, IL-1β and TGF-β enzyme-linked immunosorbent assay (ELISA) kits were obtained from Dakewe Bioengineering Co. (Shenzhen, China).

Chen Y., Du M., Yuan Z., Chen Z, & Yan F. (2022). Spatiotemporal control of engineered bacteria to express interferon-γ by focused ultrasound for tumor immunotherapy. Nature Communications, 13, 4468.

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AB23A and LPS Cytotoxicity Evaluation

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Chengdu Must Biotechnology Co., Ltd. (Chengdu, China) and Beyotime Biotechnology (Jiangsu, China) provided AB23A (purity > 98%) (Supplementary Figure S1) and LPS from E. coli (0111: B4), respectively. AB23A stock solutions were made in 0.1% DMSO, and working solutions were made in a culture medium followed by filter sterilization (0.2 mm) just prior to utilization. Gibco Life Technologies provided Dulbecco’s Modified Eagle’s Medium (DMEM) culture medium and fetal bovine serum (FBS). All biochemical indicator kits and other chemicals used in the study were commercially available unless otherwise mentioned.

Xia F., Li Y., Deng L., Ren R., Ge B., Liao Z., Xiang S, & Zhou B. (2022). Alisol B 23-Acetate Ameliorates Lipopolysaccharide-Induced Intestinal Barrier Dysfunction by Inhibiting TLR4-NOX1/ROS Signaling Pathway in Caco-2 Cells. Frontiers in Pharmacology, 13, 911196.

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Evaluation of miR-329-3p in Cell Proliferation and Apoptosis

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RPMI-1640 and Dulbecco’s Modified Eagle’s Medium (DMEM) culture medium, fetal bovine serum (FBS), trypsin, penicillin, streptomycin, dimethyl sulfoxide (DMSO) and phosphate buffered saline (PBS) were purchased from Gibco (Carlsbad, CA, USA). miR-329-3p mimic, miR-329-3p inhibitor, mimic negative control (NC) and inhibitor NC were synthesized in GenePharma (Shanghai, China). Lipofectamine 3000, TRIzol isolation kit and cDNA synthesis kit were purchased from Invitrogen (Carlsbad, CA, USA). The Cell Counting Kit-8 (CCK-8) was purchased from Sigma (St. Louis, MO, USA). The Annexin V-FITC Apoptosis Detection kit was obtained from BD Pharmingen (San Diego, CA, USA). Enzyme-linked immunosorbent assay (ELISA) kits were purchased from R & D System (Minneapolis, USA). The miRNeasy Mini kit was purchased from Qiagen (Hilden, Germany), and miRNA Complete Labeling and Hybridization kit and miRNA Microarray Release were purchased from Agilent (Santa Clara, CA, USA). The SYBR premix Ex Taq II kit was purchased from TaKaRa (Dalian, China) Primary antibodies and secondary antibodies for western blotting were purchased from Cell Signaling Technology (Danvers, MA, USA). Western ECL detection reagent was purchased from Beyotime Biotechnology (Shanghai, China). TAB (purity > 98%) was dissolved by DMSO at high concentration, which was prepared and stored at −20 ℃ for further experiments.

Chen J., Feng S., Li L., Qiu S., Jin Y, & Sun Y. (2022). Protective role of trametenolic acid B against sevoflurane-induced cognitive impairments by its different regulatory modalities of mir-329-3p in neurons and microglia. Molecular Medicine, 28, 77.

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Autophagy Regulation in Hepatocellular Carcinoma

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Dulbecco’s Modified Eagle’s Medium (DMEM) culture medium and fetal bovine serum (FBS) were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Lipo2000, Trizol and High Capacity cDNA Reverse transcription Kit were purchased from Thermo Fisher Scientific. Rapamycin and 3-methyl adenine (3-MA) were purchased from Sigma-Aldrich Co. (St Louis, MO, USA). FastStart Universal SYBR Green Master and protease inhibitor were purchased from Hoffman-La Roche Ltd. (Basel, Switzerland). Mouse anti-human polyclonal antibody against GPC3 was purchased from R&D Systems, Inc., (Minneapolis, MN, USA). Mouse anti-human monoclonal antibody against c-myc was purchased from Santa Cruz Biotechnology Inc., (Dallas, TX, USA). Rabbit anti-human monoclonal antibodies against LC3, Beclin-1, β-catenin and cyclin D1 were obtained from Cell Signaling Technology (Boston, MA, USA). Immobilon Western Chemiluminescent HRP substrate was purchased from EMD Millipore (Billerica, MA, USA). 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl-2H tetrazolium bromide (MTT) cell proliferation and cytotoxicity assay kit was purchased from Beyotime (Shanghai, China).

Hu P., Cheng B., He Y., Wei Z., Wu D, & Meng Z. (2018). Autophagy suppresses proliferation of HepG2 cells via inhibiting glypican-3/wnt/β-catenin signaling. OncoTargets and therapy, 11, 193-200.

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Synthesis and Characterization of Doxorubicin-Loaded Pluronic Nanoparticles

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Pluronic F127 (PEO₁₀₆PPO₇₀PEO₁₀₆) was purchased from BASF Co., Ltd (Hanover, Germany). DSPE-mPEG2000 was purchased from Avanti Polar Lipids, Inc (Alabaster, AL, USA). DOX and fluorescein isothiocyanate were supplied by Dalian Meilun Biology Co. Ltd (Dalian, People’s Republic of China). Other chemical reagents, such as phenol, formalin aqueous solution, NaOH, hydrogen peroxide, and chloroform, were purchased from Shanghai Chemical Regent Co (Shanghai, People’s Republic of China). The Dulbecco’s Modified Eagle’s Medium (DMEM), culture medium, fetal bovine serum (FBS), and Cell Counting Kit-8 (CCK-8) were obtained from Thermo Fisher Scientific (Waltham, MA, USA).

Wang H., Li X., Ma Z., Wang D., Wang L., Zhan J., She L, & Yang F. (2016). Hydrophilic mesoporous carbon nanospheres with high drug-loading efficiency for doxorubicin delivery and cancer therapy. International Journal of Nanomedicine, 11, 1793-1806.

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Peptide-Mediated Gene Delivery System

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All of the peptides were
custom-synthesized with >95% purity grade from G.L. Biochem (Shanghai)
Ltd. The plasmid used for this study, pMIR-REPORT Luciferase (pDNA),
was maintained in E. coli DH5α
cells and purified using GenElute HP Endotoxin-Free Plasmid MaxiPrep
Kit (Sigma). Polyethyleneimine (PEI, MW ∼ 25 kDa, branched)
was purchased from Sigma-Aldrich. Lipofectamine 2000, heparin salt
was purchased from Invitrogen. Primers for q-PCR were designed using
Primer3 Input version 0.4.0 and synthesized through (Integrated DNA
Technologies) IDT (Supporting Information Table S1). Fluorescein DNA labeling kits were purchased from Mirus
Bio Corporation. Cell viability assay kit CellTiter Glow was obtained
from Promega, and 35 mm glass-bottom imaging dishes for confocal microscopy
were purchased from ibidi cells in focus. KAPA SYBR fast 5× was
purchased from Sigma-Aldrich. Dulbecco’s modified Eagle’s
medium (DMEM) culture medium, phosphate-buffered saline (PBS), calf
fetal serum, and penicillin/streptomycin antibiotics mixture were
purchased from Invitrogen.

Dahiya U.R., Mishra S., Chattopadhyay S., Kumari A., Gangal A, & Ganguli M. (2019). Role of Cellular Retention and Intracellular State in Controlling Gene Delivery Efficiency of Multiple Nonviral Carriers. ACS Omega, 4(24), 20547-20557.

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Cytotoxicity Evaluation of Fusarium Mycotoxins

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Deoxy Nivalenol (DON), 3-acetylDeoxy Nivalenol (3-ADON), 15-acetylDeoxy Nivalenol (15-ADON), Fusarenon-X (FX), Nivalenol (NIV), and Deoxy Nivalenol-3-glucoside (D3G) were purchased from Sigma (St. Louis, MO, USA). Dulbecco’s Modified Eagle’s Medium (DMEM) culture medium, Fetal Bovine Serum and penicillin/streptomycin were purchased from Invitrogen (Carlsbad, CA, USA). Cell survival assay was performed using the cell counting kit-8 (DOJINDO, Kumamoto, Japan) according to the manufacturer’s instructions.

Yang Y., Yu S., Tan Y., Liu N, & Wu A. (2017). Individual and Combined Cytotoxic Effects of Co-Occurring Deoxynivalenol Family Mycotoxins on Human Gastric Epithelial Cells. Toxins, 9(3), 96.

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Optimized Chromatographic Separation Protocol

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HiPrep CM FF 16/10 column was purchased from GE Healthcare. SunFire C18 column (19×150 mm) was purchased from Waters. Penicillin-Steptomycin, MTT, dimethyl sulfoxide (DMSO), Diethylpyrocarbonate (DEPC) was obtained from Sigma Chemical Co. (St. Louis, MO, USA). Fetal calf serum and Dulbecco's Modified Eagle's Medium (DMEM) culture medium supplemented with Glutamine were purchased from Gibco Invitrogen (Carlsbad, CA, USA). Bicinchoninic acid (BCA) protein assay kit was obtained from Thermo scientific (Pierce Inc., Rockford, IL, USA). Other commercially available chemicals and reagents were analytical grade. All reagents were prepared using deionized MilliQ water.

Zheng L., Yi Y., Liu J., Lin X., Yang K., Lv M., Zhou X., Hao J., Liu J., Zheng Y, & Sun M. (2014). Isolation and Characterization of Marine Brevibacillus sp. S-1 Collected from South China Sea and a Novel Antitumor Peptide Produced by the Strain. PLoS ONE, 9(11), e111270.

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