Colon adenocarcinoma cells DLD-1 (Horizon Discovery) were cultured in a 1:1 mixture of Roswell Park Memorial Institute-1640 medium (Sigma-Aldrich) and Ham's F-10 nutrient mix (Lonza), supplemented with 10% fetal calf serum (FCS; BioWest), and 1% penicillin/streptomycin (Sigma-Aldrich). Human embryonic kidney cells HEK-293T, osteosarcoma cells U2OS, hTERT-immortalized human fibroblasts VH10, SV40-transformed human fibroblasts C5RO, XP4PA (XPC-deficient) (19 (link)), CS1AN (CSB-deficient) (20 (link)), CS1AN complemented with YFP-CSBdel (21 (link)), and XRCC1-YFP expressing MRC-5 were cultured in a 1:1 mixture of Dulbecco's Modified Eagle medium (Lonza) and Ham's F-10, supplemented with 10% FCS, 1% penicillin/streptomycin, 2 mM UltraGlutamine (Lonza), and 0.1 mM MEM Eagle non-essential amino acid solution (Lonza). All cells were cultured at 37°C, in 20% O2 and 5% CO2, except for VH10 cells that were cultured at 3% O2. To generate MRC-5 cells expressing XRCC1-YFP, XRCC1-YFP cDNA (a gift from Anna Campalans) (22 (link)) was cloned into pLenti-CMV-Puro-DEST (a gift from Eric Campeau & Paul Kaufman) (23 (link)). Following lentiviral transduction, MRC-5 cells stably expressing XRCC1-YFP were selected by puromycin and fluorescence-activated cell sorting.
Ham s f 10 nutrient mix
Manufactured by Lonza
Sourced in Switzerland
Ham's F-10 nutrient mix is a cell culture medium designed for the growth and maintenance of a variety of cell types. It provides essential nutrients, vitamins, and salts required for cell proliferation and survival.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Dld 1, by Horizon Discovery (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Roswell park memorial institute 1640 medium, by Merck Group (1 mentions)
Fetal calf serum (fcs), by Lonza (1 mentions)
Ham s f10, by Lonza (1 mentions)
Mem eagle non essential amino acid solution, by Lonza (1 mentions)
Penicillin streptomycin, by Merck Group (1 mentions)
Penicillin streptomycin, by Lonza (1 mentions)
Ultraglutamine, by Lonza (1 mentions)
Dulbecco s modified eagle medium, by Lonza (1 mentions)
Fetal calf serum (fcs), by Biowest (1 mentions)
Penicillin streptomycin p s, by Merck Group (1 mentions)
Dulbecco s modified eagle media, by Merck Group (1 mentions)
Hct116, by Thermo Fisher Scientific (1 mentions)
Hct116, by Horizon Discovery (1 mentions)
T 25 cm2 cultureflask, by Corning (1 mentions)
Heracell 150i co2 incubator, by Thermo Fisher Scientific (1 mentions)
L glutamine, by Thermo Fisher Scientific (1 mentions)
Phosphate buffered saline (pbs), by Merck Group (1 mentions)
4 protocols using ham s f 10 nutrient mix
1
Culture of Cell Lines for DNA Repair Research
2
Colon Cancer Cell Culture Conditions
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All cells were maintained at 37°C, in 20% O2 and 5% CO2 and regularly checked for mycoplasma. Colon cancer cells DLD-1 and HCT116 were purchased from Horizon Discovery. HT-29, HCT-15, COLO 205, LoVo, Caco-2 and SW620 were a kind gift from Riccardo Fodde (Erasmus MC Rotterdam, The Netherlands) and RKO cells were a kind gift from Georg Winter, CeMM Vienna, Austria). Cells were cultured in a 1:1 mixture of Roswell Park Memorial Institute-1640 medium (RPMI, Sigma-Aldrich) and Ham's F-10 nutrient mix (Lonza), supplemented with 10% fetal bovine serum (FBS; Gibco), and 1% penicillin/streptomycin (PS; Sigma-Aldrich). Flag-GFP-XPC knock-in (KI) HCT116 cells (hereafter referred to as GFP-XPC HCT116), a kind gift from Jurgen Marteijn (Erasmus MC Rotterdam, The Netherlands; (38 (link)), were cultured in RPMI media supplemented with 20% AmnioMAX™-II Complete Media (Thermo Fisher Scientific), 10% FBS and 1% PS. HEK293T cells were obtained from the Cancer Research UK London Research Institute Cell Facility and were used for virus production, by culturing in Dulbecco's modified Eagle media (Sigma-Aldrich) and supplemented with 10% FBS and 1% PS.
3
Isolation and Expansion of Muscle-Derived Cells
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Biopsies from the vastus lateralis muscle were obtained under local anaesthesia from each participant, using the Weil-Blakesley conchotome technique. The muscle biopsies analysed in this study were isolated and cultured29 (link), as reported previously. Briefly, biopsy samples were cut in small pieces (1 mm3) and digested in 5 mL of trypsin–EDTA for 3 × 15 min on a magnetic stirring platform at 37 °C to dissociate muscle-derived mononuclear cells. Supernatant derived following each treatment was collected and pooled with hiFBS. Cell supernatant was centrifuged at 450g for 5 min. Cell pellet was resuspended in growth media [Hams F-10 nutrient mix (Lonza, Basel, Switzerland) with added l -glutamine (2.5 mM), 10% heat-inactivated FBS (Gibco, Thermo Fisher Scientific, Altincham, UK), and 1% penicillin–streptomycin (Life Technologies, Warrington, UK)] and plated on a pre-coated T25 cm2 culture flask (Corning, Life Sciences, New York, USA) for cell population expansion. The cells were expanded until passage 3 and then frozen in GM with 10% dimethyl sulfoxide (DMSO) in liquid N2 as a cryopreservant.
4
Muscle Stem Cell Biopsy Protocol
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This methodological approach has been described in detail elsewhere (Baumert et al., 2021) . Briefly, Muscle Stem Cell study participants were instructed to avoid any strenuous exercise 48 h before the biopsy procedure. Biopsies of the vastus lateralis muscle were obtained approximately halfway between the anterior superior iliac spine and the patella under local anaesthesia by using the Weil-Blakesley conchotome technique (Baczynska et al., 2016) . The biopsies were isolated and cultured (Baumert et al., 2021) with growth media (Hams F-10 nutrient mix [Lonza] with added L-glutamine [2.5 mM], 10% heat-inactivated fetal bovine serum [Gibco, Thermo Fisher Scientific], 1% penicillin-streptomycin [Life Technologies], and 1% L-glutamine [Gibco] ). After the extraction procedure, cells were washed every 48 h following two brief washes with phosphate-buffered saline (Sigma-Aldrich) and were passaged via trypsinisation until 80% confluency for cell expansion. Cells were incubated in a HERAcell 150i CO 2 Incubator (Thermo Scientific) at 37°C/5% CO 2 . All experiments were performed on cells between passages 3 and 6 of their growth kinetics to ensure consistency and to avoid potential issues of senescence.
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