Primary cultures of astrocytes were prepared from the cerebral cortices of 1-day-old postnatal Institute of Cancer Research/ KOATEC, Pyeongtaek-si, Gyeonggi-do, Korea (ICR) mice, according to the method described by Shano et al. The cells were seeded in culture plates coated with poly-L-lysine hydrobromide (100 μg/ml; Sigma-Aldrich) and grown in Dulbecco Modified Eagle Medium (DMEM) supplemented with 10% Fetal Bovine Serum (FBS), 100 units/mL penicillin, and 100 μg/mL streptomycin in a humidified atmosphere containing 5% CO2 at 37°C. When the cells reached confluence in 5 to 7 days, they were harvested with 0.05% trypsin/EDTA solution (Life technologies, Carlsbad, CA), seeded in culture plates previously coated with poly-L-lysine hydrobromide (100 μg/ml; Sigma-Aldrich), and re-grown in DMEM supplemented with 10% FBS, 100 units/mL penicillin, and 100 μg/mL streptomycin, for performing experiments.
Poly l lysine hydrobromide
Manufactured by Merck Group
Sourced in United States, Germany, Poland, Italy, United Kingdom
Poly-L-lysine hydrobromide is a polycationic polymer used in various laboratory applications. It has a high density of positive charges, which allows it to interact with negatively charged surfaces or molecules. The primary function of Poly-L-lysine hydrobromide is to facilitate cell adhesion and improve cell growth in cell culture systems.
Automatically generated - may contain errors
Lab products found in correlation
Penicillin streptomycin, by Thermo Fisher Scientific (13 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (11 mentions)
Neurobasal a medium, by Thermo Fisher Scientific (9 mentions)
Neurobasal medium, by Thermo Fisher Scientific (8 mentions)
Bovine serum albumin (bsa), by Merck Group (7 mentions)
Papain, by Merck Group (7 mentions)
Poly l glutamic acid sodium salt, by Merck Group (5 mentions)
Direct q 5uv purification system, by Merck Group (5 mentions)
Trypsin, by Merck Group (4 mentions)
Poly sodium 4 styrenesulfonate, by Merck Group (4 mentions)
Sodium borohydride, by Merck Group (4 mentions)
Poly l lysine (pll), by Merck Group (4 mentions)
Sterilin petri plastic dishes, by Sarstedt (4 mentions)
Trypsin inhibitor, by Merck Group (4 mentions)
Horse serum, by Thermo Fisher Scientific (4 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (4 mentions)
L glutamine, by Thermo Fisher Scientific (4 mentions)
Lsm 780, by Zeiss (4 mentions)
Sodium chloride (nacl), by Merck Group (4 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (4 mentions)
140 protocols using poly l lysine hydrobromide
1
Astrocyte Primary Culture from Mice
2
Extracellular Flux Analysis of B Cells
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OCR and ECAR were measured in XF media (unbuffered RPMI 1640 containing 1% FBS, 11 mM glucose, 2 mM glutamine and 1 mM pyruvate) under basal conditions and in response to 1 μM oligomycin (O4876; Sigma), 1 μM carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone and 1 μM antimycin A (A0149; Sigma) on the XF96 extracellular flux analyzer (Seahorse Bioscience, Billerica, MA, USA). Basal OCR was calculated by subtraction of the residual rate after antimycin A treatment. After culture with LPS and IL-4 for 2.5 days, B cells were stained with MitoTracker Green and DeepRed and sorted using FACSAria. Sorted cells were cultured for 24 h under the same conditions to allow recovery from the stress of sorting. These cells were resuspended in XF media and were plated onto Seahorse cell plates (1 × 105 cells per well) coated with poly-L -lysine hydrobromide (P2636; Sigma).
To assess the effects of reagents on ECAR and OCR, we prepared cells treated with various reagents under the conditions described in ‘Reagents used in in vitro culture.' After culture with LPS and IL-4 for 48 h, cells were resuspended in XF media and were plated onto Seahorse cell plates (1 × 105 cells per well) coated with poly-L -lysine hydrobromide (P2636; Sigma). OCR and ECAR were measured as described above.
To assess the effects of reagents on ECAR and OCR, we prepared cells treated with various reagents under the conditions described in ‘Reagents used in in vitro culture.' After culture with LPS and IL-4 for 48 h, cells were resuspended in XF media and were plated onto Seahorse cell plates (1 × 105 cells per well) coated with poly-
3
Enterococcus faecalis Adhesion Protocol
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E. faecalis ATCC 29212 (American Type Culture Collection) was maintained by weekly subculturing on trypticase soy agar (TSA) plates (Scharlau, Sentmenat Barcelona, Spain). A single colony was inoculated in 40 mL of tryptic soy broth (TSB) medium and incubated at 37 °C. After 24 h of incubation, the culture was diluted 100 times in fresh TSB, and adjusted spectrophotometrically (Unicam UV-2 at 600 nm) at OD600 = 1.3 (i.e., 7.8 × 108 colony-forming units CFU/mL). Root surfaces were coated with 0.01% (w/v) poly-L-lysine hydrobromide (Sigma-Aldrich, Dorset, UK) to enhance bacterial adhesion and inoculated with 10 µl of bacterial culture using a 30-gauge syringe and needle (Becton Dickinson, Madrid, Spain). The dental roots were placed in Eppendorf tubes and incubated at 37 °C for 10 days. Re-inoculation at days 1, 4 and 7 were performed to ensure the presence of live bacteria during the incubation period [20 (link)]. Finally, the inner part of the root canal was gently washed with 1 mL of Ringer’s ¼ solution to remove the free-floating microbes and liquids.
4
Primary Cortical Neuron Isolation from WT and Mecp2 KO Mice
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Primary cortical neurons were prepared from WT and Mecp2 null mouse embryos at 15.5 days. Embryos were sacrificed by decapitation and brains were removed under a microscope and immersed in ice‐cold Hank's Buffered Salt Solution (HBSS; Life Technologies). Meninges were removed, and cerebral cortex was rapidly dissected and maintained in cold HBSS. Tissues were incubated with 0.25% trypsin/EDTA for 7 min at 37°C and the digestion was blocked with 10% FBS in DMEM (Life Technologies). Cortices were then mechanically dissociated by pipetting in Neurobasal medium, containing 10% FBS, 2% B27 (Life Technologies), 1% l ‐glutamine (Sigma‐Aldrich) and 0.5% P/S (Sigma‐Aldrich). For immunofluorescence experiments, neurons were seeded on coated glass coverslip (1 mg/ml poly‐L‐lysine hydrobromide; Sigma‐Aldrich) in 24‐well dishes at the density of 50,000 cells/well or at low density (20,000 cells/well) for morphological analysis.
5
Cell Culture Protocols for Hepatocyte Models
6
Metabolic Profiling of Splenic CD4+ T Cells
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Oxygen consumption rate (OCR, in pmol/min) and extracellular acidification rate (ECAR, in mpH/min) of splenic CD4+ T cells were analyzed using a Seahorse XF-24 metabolic extracellular flux analyzer (Seahorse Bioscience, Billerica, MA, USA) as we previously reported [12 (link)]. CD4+ T cells were first treated with the indicated reagents for 24 h and then harvested and resuspended in XF Base Medium (Seahorse Bioscience, USA) supplemented with 11 mM glucose, 1 mM pyruvate, and 2 mM glutamine. CD4+ T cells were then seeded at a density of 106 cells/well in 24-well XF microplates coated with poly-L-lysine hydrobromide (Sigma Chemical Co., USA). After incubation in a CO2-free incubator at 37 °C for 1 h, basal metabolism and metabolism in response to mitochondrial inhibitors [1 µM oligomycin, 1 µM FCCP, and 1 µM Rot+1 µM Anti A (Sigma Chemical Co., USA)] were analyzed. The assay conditions were as follows: 3 min of mixing; a 2 min wait; and 3 min of measurement. Metabolic parameters were then calculated. OCR and ECAR were normalized to the cell number in each well.
7
Synthesis and Characterization of Functional Polymeric Biomaterials
8
Synthesis and Characterization of Fluorescent Oligonucleotides
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2-Cyanoethyl diisopropylchlorophosphoramidite was received from Wako Chemicals and used without further purification. Poly-L-lysine hydrobromide (MW 30,000-70,000) and poly-L-lysine-FITC label (MW 30,000-70,000) were purchased from Sigma-Aldrich Chemicals Co. and used as received. OxiRedTM probe (also marketed and more commonly known as Amplex RedTM) was purchased from BioVision and used as received. N,N-diisopropylethylamine (DIPEA) was purchased from Nacalai and used as received. Nuclease-free water was purchased from Life Technologies Corporation and used as received. All other chemicals and solvents were purchased from Sigma-Aldrich Chemicals Co., Wako Pure Chemical Ind. Ltd., TCI, or Kanto Chemical Co. Inc. and used without further purification. Glass slides (24 × 60 mm, thickness 0.13-0.17 mm) and cover slips (18 × 18 mm, thickness 0.12–0.17 mm) were purchased from Matsunami Glass and used as received.
Glen-Pak™ DNA and RNA cartridges columns were purchased at Glen Research. ODN was obtained from Sigma Genosys. Water was deionized (specific resistance of ∼18.2 MW cm at 25°C) by a Milli-Q system (Millipore Corp.). All reactions were carried out under an argon atmosphere unless otherwise stated.
Glen-Pak™ DNA and RNA cartridges columns were purchased at Glen Research. ODN was obtained from Sigma Genosys. Water was deionized (specific resistance of ∼18.2 MW cm at 25°C) by a Milli-Q system (Millipore Corp.). All reactions were carried out under an argon atmosphere unless otherwise stated.
9
Cellular Uptake of Pharmaceutical Agents
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Quercetin dihydrate, phenolsulfonphthalein (PSP), probenecid, rifampicin, vincristine, Hank's balanced salts, minimum essential medium (MEM) non-essential amino acid solution, poly-L -lysine hydrobromide, D -glucose, HEPES, and sodium bicarbonate were purchased from Sigma (St. Louis, MO). Docetaxel trihydrate was obtained from Shin Poong Pharmaceutical Co. Ltd. (Ansan, Republic of Korea). Zoletil® 50 (a mixture of 25 mg/ml of zolazepam and tiletamine each) was obtained from Virbac (Carros, France). All other chemicals and reagents were of analytical or HPLC grade as appropriate.
10
Glioblastoma Cell Encapsulation Protocol
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1,4-butanediol diacrylate, 4-amino-1-butanol, dimethyl sulfoxide (DMSO), triethylamine (TEA), pyridine (Py), dialysis bag 10 kDa, sodium chloride (NaCl), poly-L-lysine hydrobromide (PLL), poly-L-γ-glutamate sodium salt (PGA), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT assay), DAPI, phosphate buffer Saline (PBS), Eagle’s minimum essential medium (EMEM), fetal bovine serum (FBS), ampicillin sodium salt and opti-MEM 1 reducing serum media were purchased from Sigma Aldrich. NH2-PEG-COOH was purchased from Laysan Bio. The human glioblastoma (U87MG) cell line was purchased from ATCC. Rhodamine B Isothiocyanate was purchased from CHEM-IMPEX International Inc. Eosin-5-Isothiocyanate was obtained from Chemodex. Ultrapure Agarose, Lipofectamine 3000, lysogeny broth (LB), and SYBR Safe DNA gel stain were acquired from Thermo Fisher Scientific. Tris/Borate/EDTA (TBE) Buffer was obtained from Alfa Aesar. Plasmid PiggyBac transposon (PBCAG-eGFP, 6.34 kb) was purchased from Addgene as bacterial agar stab. Bacterial amplification was performed using a sterile LB solution which contained ampicillin and was purified utilizing an endotoxin-free DNA purification kit from QIAGEN.
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