Sterile skin biopsies were obtained from the forearm of subjects after informed consent, in accordance with guidelines set by the local ethics committee. Fibroblast cell cultures were established at the Sheffield Institute for Translational Neuroscience. Monolayers of primary fibroblast cell cultures were routinely maintained in T75 flasks in fibroblast cell culture medium (Lonza) supplemented with 10% foetal calf serum (Labtech), 2 mM glutamine (Lonza BE17–605 E), 50 µg/ml uridine (Sigma U3003), vitamins (Lonza 13–607 C 1/100 dilution), amino acids (Lonza BE13–114E 1/100 dilution), 1 mM sodium pyruvate (Lonza BE13–115E) in humid incubators at 37°C supplemented with 5% CO2.
Be13 114e
Manufactured by Lonza
Sourced in Italy
The BE13-114E is a piece of lab equipment manufactured by Lonza. It is designed to perform a specific function within a laboratory setting. The core function of this product is to facilitate essential laboratory processes, but a detailed description cannot be provided while maintaining an unbiased and factual approach.
Automatically generated - may contain errors
Lab products found in correlation
Foetal calf serum, by EQT (1 mentions)
Be13 115e, by Lonza (1 mentions)
Alum adjuvant, by Thermo Fisher Scientific (1 mentions)
Pfhm 2, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Penicillin, by Euroclone (1 mentions)
Streptomycin, by Euroclone (1 mentions)
75 cm2 flask, by Thermo Fisher Scientific (1 mentions)
Be12 604f, by Lonza (1 mentions)
L glutamine, by Euroclone (1 mentions)
Penicillin streptomycin, by Merck Group (1 mentions)
4 protocols using be13 114e
1
Establishing Primary Fibroblast Cultures
2
SARS-CoV-2 Monoclonal Antibody Generation
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C57BL/6 mice were sequentially immunized in 2 weeks intervals with purified SARS-CoV-2 S1 and S2 proteins. Antigens were injected at 30 μg/mouse using Alum Adjuvant (Thermo Fisher Scientific Inc.) freshly prepared according to the manufacturer's instruction for first and boosting injection. Fourteen days after the last injection, spleen and lymph nodes are harvested, and hybridomas are made by a standard method using a myeloma cell line (RRID:CVCL_2199; ATCC CRL-1581) as a fusion partner. Hybridomas were screened in antigen-specific ELISA, and those selected for further development were subcloned and produced on a small scale (100 mL of medium). For this purpose, hybridomas are cultured in serum- and protein-free medium for hybridoma culturing (PFHM-II, Thermo Fisher Scientific Inc.) with addition of non-essential amino acids (BE13-114E, Biowhittaker Lonza, Basel, Switzerland). Monoclonal antibodies were purified from hybridoma culture supernatants using Protein-G affinity chromatography (16-266, Merck KGaA). Purified antibodies were stored at -80°C until use.
3
Caco-2 Cell Culture Protocol
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Wild-type Caco-2 (Caco-2/wt, passages 33-43, ATCC HTB-37, Manassas, VA) and modified cell lines were grown in 75-cm 2 flasks (Nunc, Roskilde, Denmark) at 37 C in 5% CO 2 atmosphere and subcultured twice a week. Standard growth medium was Dulbecco's modified Eagle's medium (Biowhittaker BE12-604F, Verviers, Belgium), supplemented with 10% fetal bovine serum (Invitrogen, Gaithersburg, MD), 2 mM L-glutamine (Euroclone, Pero, Italy), 100 U/mL penicillin to 100 mg/mL streptomycin (Euroclone), and 1 Â non-essential amino acids (Biowhittaker BE13-114E).
4
Isolation and Culture of Fetal Cardiac Progenitor Cells
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Human fetal CPCs were isolated from tissue harvested under standard informed consent procedures and prior approval of the ethics committee of the University Medical Center Utrecht. All cell studies were done in compliance with the UC San Diego Human Research Protections Program. CPCs were isolated from human fetal hearts for selection with Sca-1+ magnetic bead sorting (Miltenyi Biotech, 130-091-176) and cultured on 0.1% gelatin in growth media. Growth media consists of 25% EGM-2 (Cambrex, CC-3156) supplemented with EGM-2 Single Quots (Cambrex, CC-4176), 10% fetal bovine serum, 1× penicillin/streptomycin (Sigma, P4458), and 1× minimum essential medium (MEM) non-essential amino acids (BioWhittaker, BE13-114E) in M199 (BioWhittaker, BE12-119F). Cells at passages 3–5 after isolation were tested for mycoplasma, showing no contamination. Cells at passages 17–21 were used for all experiments.
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