The human metastatic BC cells, e.g., MDA-MB-231 (purchased from ATCC), were maintained in the Dulbecco's Modified Eagle's Medium (Gibco-BRL) supplemented with 10% fetal bovine serum (FBS) (Euroclone), 2 mM L-glutamine (Euroclone), 100 U/ml penicillin, and 100 μg/ml streptomycin (Euroclone), at 37°C, 5% CO2. The human BC cell line BT549 (purchased from ATCC) was grown in Roswell Park Memorial Institute (RPMI) 1640 supplemented with 10% FBS, 2 mM L-glutamine, 100 U/ml penicillin and 100 μg/ml streptomycin, and 0.023 U/ml insulin at 37°C, 5% CO2. The human cardiomyocytes (HCMs, purchased by PromoCell) cells were maintained in the Myocyte Growth Medium (PromoCell) plus Myocyte supplements mix (PromoCell), in addition to 10% FBS, 2 mM L-glutamine, 100 U/ml penicillin, and 100 μg/ml streptomycin, at 37°C, 5% CO2. The rat cardiomyocytes cell line H9C2 (purchased by PromoCell) was maintained in DMEM-F12, supplemented with 10% FBS, 2 mM L-glutamine, 100 U/ml penicillin, and 100 μg/ml streptomycin. All the cell lines used in the study were routinely checked for eventual mycoplasma contamination, before being used.
Penicillin
Manufactured by PromoCell
Sourced in Germany
Penicillin is a type of laboratory equipment used for the production and purification of the antibiotic compound penicillin. It is a crucial tool in the field of microbiology and pharmaceutical research.
Automatically generated - may contain errors
Lab products found in correlation
Streptomycin, by PromoCell (18 mentions)
Fetal calf serum (fcs), by PromoCell (5 mentions)
Penicillin, by Thermo Fisher Scientific (5 mentions)
Streptomycin, by Thermo Fisher Scientific (5 mentions)
Streptomycin, by Merck Group (4 mentions)
Supplementmix, by PromoCell (3 mentions)
Penicillin, by Merck Group (3 mentions)
L glutamine, by PromoCell (2 mentions)
Biocoll separating solution, by Harvard Bioscience (2 mentions)
Cd4 t cell isolation kit, by Miltenyi Biotec (2 mentions)
Rpmi 1640, by Merck Group (2 mentions)
Mesenchymal stem cell growth medium, by PromoCell (2 mentions)
Penicillin, by Euroclone (1 mentions)
Streptomycin, by Euroclone (1 mentions)
Penicillin, by American Type Culture Collection (1 mentions)
Streptomycin, by American Type Culture Collection (1 mentions)
Myocyte supplements mix, by PromoCell (1 mentions)
Myocyte growth medium, by PromoCell (1 mentions)
Mda mb 231, by American Type Culture Collection (1 mentions)
L glutamine, by Euroclone (1 mentions)
20 protocols using penicillin
1
Cell Culture Protocols for Breast Cancer and Cardiomyocytes
2
Cell Culture Protocols for Various Cell Lines
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L929 mouse fibroblasts were obtained from the American Type Culture Collection (Manassas, VA, USA). The cells were grown in complete Roswell Park Memorial Institute (RPMI) 1640 medium (with l -glutamine) supplemented with 10% fetal bovine serum (FBS), 100 IU/mL penicillin, and 100 µg/mL streptomycin.
MG-63 human osteoblast-like cells were purchased from the European Collection of Authenticated Cell Cultures (Salisbury, UK). The MG-63 cells were cultured in Eagle’s Minimum Essential medium (EMEM) supplemented with 10% FBS, 2 mMl -glutamine, 1 mM sodium pyruvate, nonessential amino acids, and antibiotics (100 U/mL penicillin, 100 μg/mL streptomycin).
Adipose-derived human mesenchymal stem cells (ADSCs) were purchased from PromoCell and cultured in Mesenchymal Stem Cell Growth Medium® supplemented with 10% Supplement Mix® (PromoCell GmbH, Germany), 100 U/mL penicillin, and 100 µg/mL streptomycin.
All cell lines were cultured at 37 °C in a humidified atmosphere with 5% CO2 and passaged at 70–80% confluency using a 0.04% trypsin ethylenediaminetetraacetic acid (EDTA) solution (ADSC cells), 0.25% trypsin-EDTA solution (MG-63 cells), or cell scraper (L929 fibroblasts). All compounds used for cell culture were purchased from Sigma-Aldrich (Darmstadt, Germany).
MG-63 human osteoblast-like cells were purchased from the European Collection of Authenticated Cell Cultures (Salisbury, UK). The MG-63 cells were cultured in Eagle’s Minimum Essential medium (EMEM) supplemented with 10% FBS, 2 mM
Adipose-derived human mesenchymal stem cells (ADSCs) were purchased from PromoCell and cultured in Mesenchymal Stem Cell Growth Medium® supplemented with 10% Supplement Mix® (PromoCell GmbH, Germany), 100 U/mL penicillin, and 100 µg/mL streptomycin.
All cell lines were cultured at 37 °C in a humidified atmosphere with 5% CO2 and passaged at 70–80% confluency using a 0.04% trypsin ethylenediaminetetraacetic acid (EDTA) solution (ADSC cells), 0.25% trypsin-EDTA solution (MG-63 cells), or cell scraper (L929 fibroblasts). All compounds used for cell culture were purchased from Sigma-Aldrich (Darmstadt, Germany).
3
Isolation of CD4+ T Cells from PBMCs
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PBMCs were isolated from fresh blood or buffy coat from healthy donors by means of density gradient centrifugation (Biocoll separating solution, 1.077 g/ml, Biochrom AG, Berlin, Germany). PBMCs were washed three times with PBS and CD14+ magnetically labeled cells were positively selected via the autoMACS separator (autoMACS, program: possel, Miltenyi Biotec, Bergisch Gladbach, Germany) twice. Untouched CD4+ T cells were purified via the CD4+ T Cell Isolation Kit (MiltenyiBiotec) and autoMACS program: depletes. Cells were cultured in RPMI 1640 (Sigma-Aldrich, Taufkirchen, Germany) supplemented with 100 IU/ml of penicillin, 100 µg/ml streptomycin containing 10% heat inactivated fetal calf serum (Promocell, Heidelberg, Germany) at 37°C in a humidified atmosphere in the presence of 5% CO2.
4
HERV-K dUTPase effects on PAECs
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Commercial human PAECs (PromoCell) were cultured in EC media supplemented with 5% FBS, EC growth supplement, and penicillin (50 U/mL)/streptomycin (50 U/mL) and used at passage 3–8. All tissue culture reagents were purchased from ScienCell. For experiments assessing the effect of recombinant HERV-K dUTPase, human PAECs were treated with 10 μg/mL of recombinant HERV-K dUTPase once a day for 3 days, followed by every 3 days up to 20 days. PAECs were evaluated for alterations in cell phenotype, gene, and protein expression at 1, 3, 10, and 20 days after initial treatment using quantitative real-time PCR (qPCR), immunoblotting, and immunofluorescence staining using a confocal microscope (FV1000, Olympus). Apoptosis was assessed on day 1, 10, and 20 following overnight (16 hours) serum withdrawal. Cells were then incubated for 1 hour in 30 μL of Caspase 3/7 Luciferase Reagent Mix (Promega), and total luminescence was measured in a plate reader (BioTek Synergy H1 Hybrid Reader).
5
Xenograft Model of NSCLC Adenocarcinoma
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The human NSCLC adenocarcinoma cell line NCI-H460-LNM3512 (link) was maintained in RPMI-1640 medium supplemented with 2 mM glutamine, penicillin (100 U/mL), streptomycin (100 μg/mL) and 10% fetal calf serum (Promocell, Heidelberg, Germany). Tumour cells (5×106 cells per mouse) were implanted subcutaneously into nu/nu BALB/c mice (Harlan Laboratories, Venray, The Netherlands) anaesthetised with ketamine (Ketalar; Pfizer, New York, New York, USA) and xylazine (Rompun vet; Bayer Healthcare, Leverkusen, Germany). At 4 days after tumour implantation, the mice were randomly assigned to receive an intraperitoneal injection of the antiangiogenic compounds, or control human immunoglobulin G. Once the largest tumour diameter reached 19 mm in length, the mice were sacrificed and the primary tumours were excised, cut in halves and fixed with 4% paraformaldehyde. The paraffin-embedded tumour tissues were cut into 5–7 μm sections, and then stained with H&E.
6
Maintenance of MARC-145 and BUVEC Cells
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MARC‐145 cell layers were maintained in Dulbecco's Modified Eagle Medium (Sigma‐Aldrich) cell culture medium supplemented with 1% penicillin (500 U/ml; Sigma‐Aldrich), streptomycin (500 mg/ml; Sigma‐Aldrich) and 5% foetal calf serum (FCS; Gibco, part of Thermo Fisher Scientific, Dreieich, Germany) and incubated at 37°C and 5% CO2 until confluency. Primary BUVEC were maintained in modified ECGM [endothelial cell growth medium (PromoCell, Heidelberg, Germany); 30% (v/v) ECMG and 70% (v/v) M199, supplemented with 1% penicillin and streptomycin and 5% FCS] at 37°C in 5% CO2 atmosphere until confluency.
34 (link)
34 (link)
7
Cultivation of Human Oral Epithelial Cells
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Human oral epithelial cells (EC) (TERT-2 OKF-6, BWH Cell Culture and Microscopy Core, Boston, MA, USA) were cultured in Keratinocyte-SFM medium (Life Technologies, Saint-Aubin, France). To reduce the risk of contamination, 100 units/mL of penicillin and 100 μg/mL of streptomycin (PromoCell, Heildelberg, Germany) were added to all cell media. Cells were grown at 37 °C in a humidified atmosphere with 5% CO2 and media were changed each 3 days.
8
Cell Culture Protocols for Cancer and Vascular Research
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The HT29 human colorectal adenocarcinoma cell line and the HCT116 human colorectal carcinoma cell lines were obtained from the European Collection of Cell Cultures (Sigma Aldrich, Dorset, UK). Human umbilical vein endothelial cells (HUVECs) were purchased from Promocell (Heidelberg, Germany) and Human dermal fibroblasts (HDFs) from Invitrogen (Paisley, UK). HT29 and HCT116 cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 1 g/L glucose, 10% FBS, 100 units/ml penicillin and 100 μg/ml streptomycin (all from Invitrogen, Paisley, UK). HUVECs were cultured in complete endothelial growth medium (EGM) (Promocell, Heidelberg, Germany) supplemented with 10% FBS and 100 units/ml penicillin and 100 μg/ml streptomycin. HDFs were cultured in high glucose (5 g/L) DMEM supplemented with 10% FBS and 100 units/ml penicillin and 100 μg/ml streptomycin. All cell types were routinely maintained as 2D monolayers at 37 °C in standard cell culture conditions (5% CO2/air and 95% humidity).
9
Isolation and Expansion of Mesenchymal Stem Cells
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We used several lines obtained from different donors (2 BMSCs and 5 TMSCs). The donor information was provided in Supplementary Table S3 . The BMSCs were purchased from PromoCell (Heidelberg, Germany). Tonsil-derived MSCs were thawed from a cell stock obtained from the patients using a study protocol approved by the Institutional Review Board (IRB) of Mokdong Hospital, Ewha Womans University (ECT 11-53-02) [29 (link)]. Written informed consent was obtained from all donors.
The TMSCs were cultured in Dulbecco’s modified Eagle’s medium containing high glucose (DMEM-HG; Welgene Inc., Gyeongsan, Republic of Korea) supplemented with 10% fetal bovine serum (FBS; Corning, NY, USA), 100 U/mL penicillin, 100 μg/mL streptomycin, and 0.25 μg/mL amphotericin B (Thermo Fisher Scientific, Waltham, MA, USA) at a density of 7 × 103 cells per cm2. The BMSCs were maintained in Mesenchymal stem cell growth medium (PromoCell) supplemented with 100 U/mL penicillin, 100 μg/mL streptomycin, and 0.25 μg/mL amphotericin B at 37 °C in a humidified atmosphere containing 5% CO2. Cells were passaged when they reached 80% of confluence. The MSCs from passages 3, 6, and 7 were used for the experiments described herein.
The TMSCs were cultured in Dulbecco’s modified Eagle’s medium containing high glucose (DMEM-HG; Welgene Inc., Gyeongsan, Republic of Korea) supplemented with 10% fetal bovine serum (FBS; Corning, NY, USA), 100 U/mL penicillin, 100 μg/mL streptomycin, and 0.25 μg/mL amphotericin B (Thermo Fisher Scientific, Waltham, MA, USA) at a density of 7 × 103 cells per cm2. The BMSCs were maintained in Mesenchymal stem cell growth medium (PromoCell) supplemented with 100 U/mL penicillin, 100 μg/mL streptomycin, and 0.25 μg/mL amphotericin B at 37 °C in a humidified atmosphere containing 5% CO2. Cells were passaged when they reached 80% of confluence. The MSCs from passages 3, 6, and 7 were used for the experiments described herein.
10
Isolation and Characterization of Human Mesenchymal Stem Cells
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HMADS cells used as human MSC model were isolated from adipose tissues obtained from young donors after informed parental consent as previously reported.44 (link) The human fetal ventricular RL14 cell line and primary HUVEC were purchased from the American Type Cell Culture (ATCC, LGC Standards S.a.r.l. Molsheim France) and PromoCell (Heidelberg, Germany), respectively.
HMADS cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM), 1 g/l glucose, containing 10% heat-inactivated fetal bovine serum (FBS; Dominique Dutscher, Issy Les moulineaux, France), 100 U/ml penicillin, 100 μg/ml streptomycin and 10 mM HEPES (Invitrogen, Carlsbad, CA, USA). As described earlier,44 (link) HMADS cells exhibited the following phenotype: CD44+, CD49b+, CD105+, CD90+, CD13+, Stro-1−, CD34−, CD15−, CD117−, Flk-1−, Gly-A−, CD133−, HLA-DR− and HLA-Ilow.
Human RL14 cells were grown in DMEM/F-12 (Life Technologies, Carlsbard, CA, USA) supplemented with 12.5% heat-inactivated FBS, 100 U/ml penicillin, 100 μg/ml streptomycin and 10 mM HEPES.45 (link) HUVEC cells were expanded on gelatin (2%)-coated dishes with the growth medium recommended and commercialized by the manufacturer (PromoCell). All cell types were maintained in a 5% CO2 atmosphere at 37 °C.
HMADS cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM), 1 g/l glucose, containing 10% heat-inactivated fetal bovine serum (FBS; Dominique Dutscher, Issy Les moulineaux, France), 100 U/ml penicillin, 100 μg/ml streptomycin and 10 mM HEPES (Invitrogen, Carlsbad, CA, USA). As described earlier,44 (link) HMADS cells exhibited the following phenotype: CD44+, CD49b+, CD105+, CD90+, CD13+, Stro-1−, CD34−, CD15−, CD117−, Flk-1−, Gly-A−, CD133−, HLA-DR− and HLA-Ilow.
Human RL14 cells were grown in DMEM/F-12 (Life Technologies, Carlsbard, CA, USA) supplemented with 12.5% heat-inactivated FBS, 100 U/ml penicillin, 100 μg/ml streptomycin and 10 mM HEPES.45 (link) HUVEC cells were expanded on gelatin (2%)-coated dishes with the growth medium recommended and commercialized by the manufacturer (PromoCell). All cell types were maintained in a 5% CO2 atmosphere at 37 °C.
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