Concentrated as well as unconcentrated LV and IDLV supernatants were used to transduce different cell lines. Cells were washed with Dulbecco’s PBS (1×) (Biowest), counted, and plated on 48-well plates and incubated for 5 hr with different LV and IDLV particles at different MOIs (from 0.3 to 10). Media were changed after 5 hr. NPCs and 293T cells were washed with Dulbecco’s PBS (1×) (Biowest), dissociated with TrypLE (Gibco), and plated on 24-well plates in the presence of the fresh viral particles at different TU/cells. The cell line PBMC1-iPS4F1 was incubated for 5 hr on the day of passage with a concentrated virus in the presence of 8 μg/mL Polybrene and 10 μM Y-27632 (Sigma-Aldrich).
Dulbecco s pbs
Manufactured by Biowest
Sourced in France
Dulbecco's Phosphate-Buffered Saline (Dulbecco's PBS) is a balanced salt solution commonly used in cell culture applications. It is a sterile, isotonic buffer that maintains the pH and osmotic balance of cells in vitro. Dulbecco's PBS is a versatile solution that can be used for various purposes, such as washing cells, diluting reagents, and maintaining cell viability during experimental procedures.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 3000, by Thermo Fisher Scientific (2 mentions)
Hepes, by Merck Group (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Biowest (1 mentions)
100 μm cell strainer, by BD (1 mentions)
Liberase tl research grade, by Roche (1 mentions)
Tryple, by Thermo Fisher Scientific (1 mentions)
Y 27632, by Merck Group (1 mentions)
Polybrene, by Merck Group (1 mentions)
Phosphate buffered saline (pbs), by Biowest (1 mentions)
5 protocols using dulbecco s pbs
1
Transduction of Cell Lines with LV and IDLV
2
Microscopic Analysis of Cellular Osmotic Stress
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DM cells stably expressing H2B-mRFP and mGFP [1 (link),3 (link)] were kindly provided by Dr. Maeshima (National Institute of Genetics, Mishima, Japan). DM cells were cultured at 37 °C in an atmosphere containing 5% CO2 in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 20% fetal bovine serum (FBS), 100 U/mL penicillin, and 100 U/mL streptomycin. HeLa cells were cultured at 37 °C in an atmosphere containing 5% CO2 in DMEM supplemented with 10% FBS, 100 U/mL penicillin, and 100 U/mL streptomycin. For live cell microscopy, stably-transfected DM cells were plated in Tomo-dishes (Tomocube, Daejeon, Korea) or Lab-Tek 8-well chambered coverglasses (Nunc, Rochester, NY, USA). To observe HeLa cells expressing H2B-mRFP, cells were transiently transfected with the H2B vector using Lipofectamine 3000 (Thermo Fisher Scientific, Waltham, MA, USA).
To investigate the influence of osmotic stress, DM cells, which are normally cultured and maintained in a DMEM with high osmolarity (i.e., refractive index), containing 20% FBS, were incubated with phosphate-buffered saline (PBS) (hypoosmolar) or DMEM containing 0.2 M sucrose and 20% FBS (hyperosmolar) (Table 1 ). Media were warmed to 37 °C before use. For HeLa cells, PBS or phenol-free DMEM supplemented with 10% FBS and 0.2 M sucrose was added before imaging. Dulbecco’s PBS (Biowest, Nuaille, France) was used to wash DM and HeLa cells.
To investigate the influence of osmotic stress, DM cells, which are normally cultured and maintained in a DMEM with high osmolarity (i.e., refractive index), containing 20% FBS, were incubated with phosphate-buffered saline (PBS) (hypoosmolar) or DMEM containing 0.2 M sucrose and 20% FBS (hyperosmolar) (
3
Isolation of Stromal Vascular Fraction Cells
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Stromal vascular fraction (SVF) cells were isolated by a previously described methodology20 (link). Briefly, small pieces of adipose tissue (1–2 g of SAT or MAT) (avoiding blood vessels) were added to tubes containing Hanks’ balanced salt solution supplemented with 4% BSA, 10 mM HEPES and Liberase™ TL Research Grade (Roche Diagnostics, Risch-Rotkreuz, Switzerland) and incubated in a water bath up to 60 min at 37 °C, with manual shaking each 10 min. After water bath incubation, digested samples were homogenized to single-cell suspensions, passed through a 100-μm cell strainer (BD Biosciences Pharmingen, San Diego, CA) and centrifuged at 280 × g for 10 min at 4 °C. Cells at the bottom, corresponding to the SVF were resuspended in Dulbecco’s PBS, supplemented with 2% FBS (Biowest, Nuaillé, France), 2 mM EDTA and 10 mM HEPES (all from Sigma-Aldrich) for flow cytometry studies and complete RPMI medium for cell sorting experiments.
4
Cellular Permeability Assay with FD4
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The cell wall permeability for larger molecular weights was tested with FD4 (Sigma-Aldrich), a FITC-labeled dextran with a molecular size of 4 kDa. According to Ginzburg et al. (1999) (link), cells were incubated with FD4 for a minimum of 1 h. As stock solution 4 mg FD4 mL-1 was prepared with Dulbecco‘s PBS (Biowest, Nuaillé, France). Final concentration was 1 mg FD4 mL-1 at a cell concentration of 1 × 106 cells mL-1 Potential intracellular FITC-fluorescence was examined with an ImageStream®X Mk II. Compensation for non-FD4-deriving fluorescence was performed with unstained cells. 1000–2000 cells were recorded for each treatment and species.
5
Deciduous Tooth Stem Cell Extraction
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A total of 28 deciduous teeth indicated for extraction were collected from 25 patients at the Pediatric Dentistry Department in Faculty of Dental Medicine, Cairo University. Patient age ranged from 7 to 12 years. Collection was done at the pediatric clinic over 3 days, we looked for deciduous teeth indicated for extraction due to their natural shedding time in order to make room for their permanent successors, so no ethical concerns would arise. Deciduous tooth collection was conducted after obtainment of the guardians’ written informed consent at Pediatric Dentistry Department in the Faculty of Dental Medicine Cairo University, with the approval of the Ethics Committee of the Faculty of Oral and Dental Medicine, Cairo University. Subjects were identified by their treating physician, following which we contacted the guardians of the subjects for consent to use the extracted teeth. Stem cell propagation (at the Medical Biochemistry Department in the Faculty of Medicine Cairo University) was performed in accordance with recommendations and with the approval of the Ethics Committee of the Faculty of Oral and Dental Medicine, Cairo University.
Deciduous tooth surfaces were washed several times with Dulbecco’s PBS (Biowest, USA). Dental pulp was extracted delicately from teeth using a sterile endodontic barbed broach and placed in falcon tube containing PBS (Biowest, USA).
Deciduous tooth surfaces were washed several times with Dulbecco’s PBS (Biowest, USA). Dental pulp was extracted delicately from teeth using a sterile endodontic barbed broach and placed in falcon tube containing PBS (Biowest, USA).
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