Cell lines were acquired from the American Type Culture Collection (ATCC) and cell culture media were obtained from Lonza. All media were supplemented with 10% fetal bovine serum (FBS). HEK-293 cells were grown in DMEM, while 32Dcl3 cells were grown in RPMI containing 10 ng/ml of murine IL-3. The cells were grown at 37 °C with 5% of CO2 and maintained according to manufacturer instructions. For transfection experiments, HEK293 cells were plated at 70% confluency. We utilized the calcium-phosphate method [12 (link)] with 10 μg of corresponding plasmids. The medium was changed after 8 h and the cells were incubated for 36 h. Nuclear extracts were made using Dignam’s protocol [13 (link)]. For stable transfection of 32Dcl3 cells the NEPA21 Electroporator was used (NEPA GENE). 32Dcl3 cells and immortalized NUP98-NSD1/FLT3-ITD primary murine cells were lysed using 4X Laemmli buffer, after which the lysate was boiled at 95 °C for 5 min. Patient cells were obtained from the lab of Prof. Olaf Heidenreich at Princes Maxima Center for Pediatric Oncology in Utrecht, The Netherlands. NUP98-NSD1+ patient cells were grown in StemSpan™ (STEMCELL Technologies) supplemented with 10 ng/mL IL-3, 10 ng/mL FLT3 ligand, 10 ng/mL GM-CSF, 150 ng/mL SCF, 100 ng/mL TPO.
Cell culture media
Manufactured by Lonza
Sourced in United States, Switzerland
Cell culture media is a liquid or powder formulation designed to support the growth and maintenance of cells in a laboratory setting. It provides the necessary nutrients, vitamins, and other components required for optimal cell health and proliferation.
Automatically generated - may contain errors
Lab products found in correlation
Amphotericin b, by Lonza (3 mentions)
Penicillin, by Lonza (3 mentions)
Streptomycin, by Lonza (3 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Anti phospho nf κb p65, by Abcam (2 mentions)
Anti phospho c jun n terminal kinase jnk, by Abcam (2 mentions)
Anti phospho extracellular signal regulated kinase erk1 2, by Abcam (2 mentions)
Anti phospho p38, by Abcam (2 mentions)
Horseradish peroxidase conjugated anti rabbit antibody, by Abcam (2 mentions)
Proteinase k, by Merck Group (2 mentions)
α amylase, by Merck Group (2 mentions)
Phosphate buffered saline (pbs), by Lonza (2 mentions)
Stemspan, by STEMCELL (1 mentions)
Nepa21 electroporator, by Nepa Gene (1 mentions)
Anti cd11b m1 70, by Thermo Fisher Scientific (1 mentions)
Draq5 fluorescent probe, by Thermo Fisher Scientific (1 mentions)
Anti smad2 3 3102, by Cell Signaling Technology (1 mentions)
Anti β tubulin, by Merck Group (1 mentions)
Anti cd68 ebm11, by Agilent Technologies (1 mentions)
Cd140a pe, by BD (1 mentions)
23 protocols using cell culture media
1
Transfection and Lysis of Cell Lines
2
Molecular Signaling Pathways Profiling
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Cell culture media and its supplements were purchased from Lonza, Inc. (Walkersville, MD, USA). The major enzymes such as α-amylase and proteinase K were attained from Sigma-Aldrich, St. Louis, MO, USA. The primary antibodies such as anti-phospho-NF-κB p65, anti-phospho-c-Jun N-terminal kinase (JNK), anti-phospho-extracellular signal-regulated kinase (ERK1/2), and anti-phospho p38, and secondary antibody (horseradish peroxidase-conjugated anti-rabbit antibody) were obtained from Abcam, Cambridge, UK. The antibodies for flow cytometry analysis such as anti-CD40-APC (1C10) and anti-CD11b (M1/70) were procured from ThermoFisher Scientific, Waltham, MA, USA.
3
Immunofluorescence and Western Blot Analysis
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Cell culture media, serum, phosphate-buffered saline and trypsin were purchased from Lonza (Verviers, Belgium) and cell culture reagents were from Sigma-Aldrich Chimie (Saint-Quentin Fallavier, France).
Human antibodies for immunofluorescence analysis were purchased as indicated: anti-αSMA (#A5228) from Sigma-Aldrich, anti-CD68 (#EBM11) from Dako (Glostrup, Denmark), anti-MR (#ab8918) from Abcam (Cambridge, United-Kingdom), anti-CD140a (#3174 T) from Cell Signaling (Ozyme, St Quentin en Yvelines, France). Human antibodies for western blot analysis were purchased as indicated: anti-phospho-Smad2 (Ser465/467, #3101) and anti-Smad2/3 (#3102) from Cell Signaling, and anti-β-tubulin from Sigma-Aldrich. Human antibodies against CD56-APC (#341027) and CD140a-PE (#556002) were purchased from BD-Biosciences (Le Pont de Claix, France). Human recombinant IL-4 (#200-04) and IL-1β (#200-01B) were purchased from Peprotech (Neuilly-Sur-Seine, France) and used at 15 ng/ml. SB431542 (#S4317) was purchased from Sigma-Aldrich Chimie and used at 5 µM. DRAQ5 fluorescent probe (#62254) was purchased from Thermofisher Scientific and used at 20 µM.
Human antibodies for immunofluorescence analysis were purchased as indicated: anti-αSMA (#A5228) from Sigma-Aldrich, anti-CD68 (#EBM11) from Dako (Glostrup, Denmark), anti-MR (#ab8918) from Abcam (Cambridge, United-Kingdom), anti-CD140a (#3174 T) from Cell Signaling (Ozyme, St Quentin en Yvelines, France). Human antibodies for western blot analysis were purchased as indicated: anti-phospho-Smad2 (Ser465/467, #3101) and anti-Smad2/3 (#3102) from Cell Signaling, and anti-β-tubulin from Sigma-Aldrich. Human antibodies against CD56-APC (#341027) and CD140a-PE (#556002) were purchased from BD-Biosciences (Le Pont de Claix, France). Human recombinant IL-4 (#200-04) and IL-1β (#200-01B) were purchased from Peprotech (Neuilly-Sur-Seine, France) and used at 15 ng/ml. SB431542 (#S4317) was purchased from Sigma-Aldrich Chimie and used at 5 µM. DRAQ5 fluorescent probe (#62254) was purchased from Thermofisher Scientific and used at 20 µM.
4
eNOS Regulation in Bovine Aortic Cells
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Bovine aortic endothelial cells (BAECs) and cell culture media were obtained from Lonza. (Walkersville, MD). A monoclonal antibody against eNOS and the antibodies against eNOS Ser1179 phosphorylation and eNOS Thr497 phosphorylation were obtained from Upstate Biotechnology Inc. (Lake Placid, NY). Protein phosphatase 2A (PP2A) and Protein phosphatase 1 (PP1) kit was obtained from Millipore (Billerica, MA). N-methyl-D-glucamine dithiocarbamate/ferrous sulfate (MGD-Fe) and 5-(Diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide (DEPMPO) were purchased from Alexis. CaM, NADPH, L-arginine, BH4, N-nitro-L-arginine methyl ester (L-NAME), and other reagents were purchased from Sigma Chemical Co. (St. Louis, MO), unless otherwise indicated. Both siRNA scramble control (siRNA control) and smart pool siRNA of heat shock protein 90 (siHsp90) were purchased from Dharmacon (GE Health Life Science, Pittsburgh, PA).
5
Skeletal Muscle Cell Characterization
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Cell culture media was obtained from Lonza. FBS and trypsin-EDTA were obtained from Life Technologies (Paisley, UK). Chick embryo extract was purchased from Sera Labs International (Sussex, UK). Phospho-AMPKThr172 (40H9) and total AMPKα (F6) antibodies were obtained from New England Biolabs (Herts, UK). Anti-myosin, skeletal fast (clone MY-32) and β-actin (clone AC-15) antibodies were purchased from Sigma. Monoclonal mouse anti-human desmin (D33) antibody was obtained from DAKO. Vector VIP HRP-substrate kit was obtained from Vector Laboratories. 2-Deoxy-D-[2,6-3H]glucose was purchased from Hartmann Analytic (Germany). IL6 ELISA kits were obtained from Qiagen (Sussex, UK).
6
EGFR Signaling Pathway Peptide Assay
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All protected amino acids, Rink amide MBHA resin and PAL-NovaPEGresin were purchased from Novabiochem unless otherwise noted. HCTU was purchased from Peptides International. All synthesis reagents and solvents were purchased from Fisher, Sigma-Aldrich, or Acros and used without further purification. Cell culture media and phosphate buffer saline were obtained from Lonza, fetal bovine serum from Thermo, penicillin/streptomycin and bovine serum albumin from Amresco, trypsin EDTA from Cellgro, and EGF from Abcam. Antibody to EGFR p-Y1068 was purchased from Abcam or Pierce, total EGFR from Santa Cruz Biotech, , and α-tubulin from the University of Iowa. PVDF membrane was purchased from Millipore. Immobilized chymotrypsin was obtained from Proteochem, and chemiluminescent substrate and immobilized trypsin were obtained from Pierce. Peptide characterization and purification were performed on a Zorbax SB-C18, 5 μm HPLC column using an Agilent 1200 series HPLC system coupled to the Agilent 6120 quadrupole LC/MS. Peptides and viability assays were quantified using the Biotek Synergy 2 microplate reader. C57BL/6J mice obtained from Jackson Laboratories were maintained in a local breeder colony. All mouse housing and handling procedures were approved by the University of Georgia Institutional Animal Care and Use Committee.
7
Damnacanthal Biological Protocol
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Damnacanthal was purchased from Calbiochem (Darmstadt, Germany). Supplements and other chemicals not listed in this section were obtained from Sigma Chemicals Co. (St. Louis MO, USA). Cell culture media, penicillin, streptomycin and amphotericin B were purchased from Biowhittaker (Walkersville, MD, USA). Fetal bovine serum (FBS) was a product of Harlan-Seralb (Belton, United Kingdom). Plastics for cell culture were supplied by NUNC (Roskilde, Denmark) and VWR (West Chester, Pennsylvania, USA). Collagen was provided by SERVA Electrophoresis (Heidelberg, Germany). Antibodies used in this work were purchased from Cell Signaling Technology (Danvers, MA, USA).
8
Cell Culture Reagents and Antibodies
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(+)-Apl-1 was provided by Instituto Biomar (León, Spain) and was dissolved in DMSO and stored at −20 °C. Supplements and other chemicals not listed in this section were obtained from Sigma Chemicals Co. (St. Louis, MO, USA). Cell culture media, penicillin, streptomycin and amphotericin B were purchased from Biowhittaker (Walkersville, MD, USA). Fetal bovine serum (FBS) and human serum were products of Harlan-Seralb (Belton, UK). Antibodies (anti-TXNRD1, anti-TXNDC5, anti-PYCR1, and anti-PRX IV) were acquired from AbCam (Cambridge, UK). Antibodies (anti-GAPDH, anti-p-Iκκα/β, anti-Iκκβ) were purchased from Cell Signaling Technology (Danvers, MA, USA). Antibodies (anti-Nrf2) were purchased from Santa Cruz Biotechnologies (Dallas, TX, USA). Secondary antibodies (anti-rabbit IgG, horseradish peroxidase-linked whole antibody, and anti-mouse IgG, horseradish peroxidase-linked whole antibody) were acquired from GE Healthcare (Buckinghamshire, UK). Plastics for cell culture were supplied by NUNC (Roskilde, Denmark) and VWR (West Chester, PA, USA).
9
Silver Nitrate Antimicrobial Activity
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Silver nitrate (AgNO3; Sigma-Aldrich) and cell culture media (Lonza) where obtained from commercial sources, while microbial strains were clinically isolated, cultured, and tested for antibiotic resistance at the biopharmaceutical laboratory, SRTA, Alexandria, Egypt. Arthrospira sp was cultured on a modified Zarrouk medium at ~ 25 ± 2°C. All used cell lines were acquired from ATCC. E. coli K12 AG100 and E. coli HB101 were obtained from the Department of Pharmaceutical Microbiology, Faculty of Pharmacy, Tanta University.
10
Rat C6 and Human U87 Glioma Cell Culture
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The rat C6 glioma cell line [66 (link)] was purchased from the European Collection of Animal Cell Culture (ECACC) (Salisbury, UK). The human U87 MG glioblastoma cell line was obtained from the American Type Culture Collection (Manassas, VA, USA). C6 cells were utilized from passage 9 to 20, to avoid the possible loss in these cells of efficient coupling of the VIP and PACAP receptors to adenylate cyclase at late passages reported by other investigators [33 (link)]. In standard monolayer conditions, the cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) with GlutamaxTM I, and supplemented with 10% fetal bovine serum, 100 U/mL penicillin, and 100 µg/mL streptomycin. Cells were incubated in humidified 95% air, 5% CO2 at 37 °C. Medium was changed twice a week, and cells were subcultured once a week using trypsin-EDTA solution. Cell culture media, supplements and reagents were from Lonza, Levallois-Perret, France.
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