Penicillin streptomycin

Manufactured by Roche

Sourced in Switzerland

Penicillin/streptomycin is a commonly used antibiotic solution for cell culture applications. It contains a combination of the antibiotics penicillin and streptomycin, which are effective against a broad spectrum of bacteria.

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Streptomycin (41 citations) Penicillin (35 citations) Penicillin and streptomycin (5 citations)

35 protocols using penicillin streptomycin

Generating HCC Cell Lines from Distinct Mouse Models

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Myc^OE/Trp53^KO, Myc^OE/Pten^KO, Nras^G12D/Pten^KO, and Nras^G12V/Pten^KO HCC cell lines were generated and isolated from end-stage tumor-bearing mice of each distinct HCC mouse model as previously described in ref. ^{109 (link)}. Cells were grown on Collagen Type I rat tail (Corning, cat.no. 354236) pre-coated flasks/dishes and cultured using DMEM (Gibco, cat. no. 61965059) supplemented with 10% FCS, 1x penicillin/streptomycin (Roche) (referred to as complete medium).
AML12 cells were provided as a kind gift from Urszula Hibner (Institut de Genetique Moleculaire de Montpellier, France) and cultured with DMEM-F12+GlutaMax supplemented with 10% FBS (Capricorn, cat.no. FBS-12A), 1% penicillin/streptomycin (Roche) and 1% insulin-Transferrin-Selenium (ITS) (Gibco) (referred to as complete F12 medium)^{114 (link)}.
HEK-Blue^TM cells (Invivogen cat. code hkb-il1r) were cultured in complete medium supplemented with 100 µg/mL Normocin according to the manufacturer’s instructions.
All cell lines were cultured at 37 °C and 5% CO₂ and routinely tested for mycoplasma contamination using a MycoAlert® mycoplasma detection kit (Lonza, cat: LT07-218). Only mycoplasma-negative cells were used. The purchased cell lines were not authenticated.

Ramirez C.F., Taranto D., Ando-Kuri M., de Groot M.H., Tsouri E., Huang Z., de Groot D., Kluin R.J., Kloosterman D.J., Verheij J., Xu J., Vegna S, & Akkari L. (2024). Cancer cell genetics shaping of the tumor microenvironment reveals myeloid cell-centric exploitable vulnerabilities in hepatocellular carcinoma. Nature Communications, 15, 2581.

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NIH 3T3 Cell Transfection and Protein Isolation

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NIH 3T3 mouse fibroblast cells (ATCCRCRL-1658) were maintained in Dulbecco’s modification of Eagle’s medium (DMEM, with 4.5 g/l glucose, L-glutamine & sodium pyruvate, Corning) with fetal bovine serum (FBS, 10% v/v, Biowest) and Penicillin/Streptomycin (100 U/mL, Roche Diagnostics GmbH). Cells were cultured on 10 cm sterile plates in 37°C, in humidified a chamber with 5% CO_2. Lipofectamine 2000 (Invitrogen) was used to transfect cells (at 70% confluence, one day after seeding) according to manufacturer’s protocol. 10 μg of plasmid DNA (4 variants of shRNA carrying plasmids, OriGene TL501619) DNA was used to transfect one plate. The medium was replaced 6 hours after transfection with a standard one. 3 days after transfection cells were washed twice with cold PBS (137 mM NaCl, 7.97 mM Na₂HPO₄ × 12 H₂O, 2.68 mM KCl, 1.47 mM KH₂PO₄), detached in PBS and moved to a 1.5 ml tube. After centrifugation the cell pellet was lysed in 1 ml of cold RIPA buffer (50 mM Tris-HCl pH7.4, 1% NP-40, 0.5% Na-deoxycholate, 0.1% SDS, 150 mM NaCl, 2 mM EDTA, 50 mM NaF) with freshly added protease (cOmplete ULTRA tablets, EDTA-free, Roche Diagnostics GmbH) and phosphatase inhibitors (PMSF 1mM, Na₃VO₄ 0.2mM). The protein isolate was separated from cell debris by centrifugation (15 min, 14000g, 4°C) and stored at -20°C.

Olejniczak I., Ripperger J.A., Sandrelli F., Schnell A., Mansencal-Strittmatter L., Wendrich K., Hui K.Y., Brenna A., Ben Fredj N, & Albrecht U. (2021). Light affects behavioral despair involving the clock gene Period 1. PLoS Genetics, 17(7), e1009625.

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Steroidogenic Pathway Analysis in Cells

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McCoy's 5A medium and 0.4% trypan blue were purchased from Invitrogen/GIBCO (Carlsbad, CA). Penicillin-streptomycin was purchased from Roche Diagnostics (Indianapolis, IN). BSA and anti-β-tubulin antibody were purchased from Sigma Chemical Co. (St. Louis, MO). Purified hCG was purchased from Dr. A. F. Parlow (National Hormone and Peptide Program, Torrance, CA). Forskolin was obtained from BIOMOL Research Laboratories (Plymouth Meeting, PA). Rapamycin, anti-mouse or anti-rabbit IgG horseradish peroxidase conjugates, anti-p-S6K1, and anti-S6K1 antibodies were purchased from Cell Signaling Technology (Beverly, MA). Anti-CYP11A1 was obtained from Abcam (Cambridge, MA). Femto Super Signal Chemiluminescence reagent and Restore stripping buffer were purchased from Pierce (Rockford, IL). Progesterone Enzyme Immunoassay (EIA) kit was purchased from Cayman Chemical (Ann Arbor, MI). Real-time PCR reagents, as well as the primers and probes for STAR (assay id: Hs00264912_m1), CYP11A1 (assay id: Hs00167984_m1), HSD3B1 (assay id: Hs01084547_gH), and 18S rRNA (assay id: Hs99999901_s1) were purchased from Applied Biosystems (Foster City, CA). All other reagents used were conventional commercial products.

Moravek M.B., Shang M., Menon B, & Menon K. (2016). HCG-Mediated Activation of mTORC1 Signaling Plays a Crucial Role in Steroidogenesis in Human Granulosa Lutein Cells. Endocrine, 54(1), 217-224.

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Isolation of Human T Lymphocytes

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T lymphocytes were isolated from buffy-coats of a healthy volunteer, using RosetteSep^® Human T Cell Enrichment Cocktail (StemCell Technologies, Vancouver, BC, Canada) according to manufacture protocol. The enriched T cells were washed two times with phosphate buffer saline (PBS) +2% fetal bovine serum. The lymphocytes were suspended in completed Roswell Park Memorial Institute (RPMI1640) (RPMI1640 with L-glutamine (Sigma) supplemented with 10% fetal calf serum (FCS) (Gibco) and 1% penicillin/streptomycin (Roche)) and T cell suspension was adjusted at 2 × 10⁶ Cells/ml.

Alipour R., Adib M., Hashemi-Beni B, & Sadeghi F. (2014). The effect of stem cell from human exfoliated deciduous teeth on T lymphocyte proliferation. Advanced Biomedical Research, 3, 202.

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Culturing Various Cell Lines

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HAP1 cells have been described previously^{43 (link)}. A375, A431, A549, DLD1, JY, Raji, RKO and SKBR3 cells were purchased from American Type Culture Collection (ATCC). B16F10 cells were kindly provided by D. Peeper. Ba/F3 cells are bone-marrow-derived, immortalized cells which are IL-3-dependent, as described in^{44 (link)}. NKIRTIL006 cell line was generated from a patient treated at the Netherlands Cancer Institute.
HAP1 cells were cultured in IMDM (ThermoFisher Scientific) supplemented with 10% Fetal Calf Serum (FCS, Sigma), 100 U/mL penicillin (Roche) and 100 μg/mL streptomycin (Roche) (penicillin/streptomycin) and L-glutamine. NKIRTIL006 cells were cultured in IMDM supplemented with 10% FCS and penicillin/streptomycin. A375, A549 and B16F10 were cultured in DMEM supplemented with 10% FCS and penicillin/streptomycin. A431, DLD1, Raji and JY cells were cultured in RPMI supplemented with 10% FCS and penicillin/streptomycin. Ba/F3-Her2 cells were cultured in RPMI supplemented with 10% FCS, penicillin/streptomycin, 0.2 ng/mL mouse IL-3 (Immunotools) and 5.0 μg/mL puromycin. SKBR3 cells were cultured in IMDM supplemented with 20% FCS and 100 U/mL penicillin/streptomycin. Cells were cultured at 37°C and 5% CO₂.

Logtenberg M.E., Jansen J.H., Raaben M., Toebes M., Franke K., Brandsma A.M., Matlung H.L., Fauster A., Gomez-Eerland R., Bakker N.A., van der Schot S., Marijt K.A., Verdoes M., Haanen J.B., van den Berg J.H., Neefjes J., van den Berg T.K., Brummelkamp T.R., Leusen J.H., Scheeren F.A, & Schumacher T.N. (2019). Glutaminyl cyclase is an enzymatic modifier of the CD47- SIRPα axis and target for cancer immunotherapy. Nature medicine, 25(4), 612-619.

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Culturing Various Cell Lines

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Lentiviral Transduction of Podocytes with GFP-LC3

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Podocytes were kindly provided by Moin Saleem (Bristol Medical School, University of Bristol, Bristol, UK) and transduced to stably express GFP-LC3 via lentiviral expression vector (kind gift of Noboru Mizushima, Department of Biochemistry and Molecular Biology, University of Tokyo, Japan). Podocytes were cultured as previously described (see ref [92 (link)]) and seeded in RPMI 1640 (Thermo Fisher Scientific, 2187034) supplemented with 10% fetal calf serum, insulin/transferrin/selenite, penicillin/streptomycin (Roche, 11074547001), pyruvate, 100x minimal essential medium and HEPES (4-[2-hydroxyethyl]-1-piperazineethanesulfonic acid) buffer (all Life technologies, 11360070, 11140050 and 15630080, respectively). When cells had grown to 60% confluence differentiation was induced at 37°C for 10–14 d. Cultured human podocytes were fully differentiated before any treatment.

Bork T., Liang W., Yamahara K., Lee P., Tian Z., Liu S., Schell C., Thedieck K., Hartleben B., Patel K., Tharaux P.L., Lenoir O, & Huber T.B. (2019). Podocytes maintain high basal levels of autophagy independent of mtor signaling. Autophagy, 16(11), 1932-1948.

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Primary Human Keratinocyte Isolation

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Normal primary human keratinocytes were isolated from neonatal foreskin tissue as described previously (Kuhn et al., 1999 ). N-TERT (Dickson et al., 2000 (link)) and primary keratinocytes were cultured in EpiLife media supplemented with Human Keratinocyte Growth Serum (ThermoFisher Scientific, Waltham, MA) and 1000U Penicillin/Streptomycin (Roche, Indianapolis, IN). All experiments were conducted using sub-confluent, low passage primary normal human keratinocytes. All relevant procedures using human tissue have been approved by the Indiana University School of Medicine Institutional Review Board.

Loesch M.M., Collier A.E., Southern D.H., Ward R.E., Tholpady S.S., Lewis D.A., Travers J.B, & Spandau D.F. (2016). Insulin‐like growth factor‐1 receptor regulates repair of ultraviolet B‐induced DNA damage in human keratinocytes in vivo. Molecular Oncology, 10(8), 1245-1254.

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Evaluating Tumor-Initiating Potential in Breast Cancer

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All animal experiments were conducted in accordance with accepted standards of animal care and in agreement with a protocol established with our Ethics Committee. The study has been approved by the Institutional Review Board “CEINGE-Biotecnologie Avanzate”. To evaluate in vivo tumorigenicity, unsorted HCC1937 or sorted cell subpopulations were resuspended in media and Matrigel (1:1; BD Biosciences), and injected into the left and right flanks of 4-week-old NOD/SCID mice (C. River laboratories). To evaluate the tumorigenic potential of the unsorted cell line, 2 × 10⁶ and 4 × 10⁵ of HCC1937 cells were injected into the left and right flanks of mice respectively (n = 5). To compare the tumorigenic potentials of the CD24⁺ and CD24^- cell populations, 5 × 10⁴ or 5 × 10⁵ cells were injected into the left and right flanks of NOD/SCID mice respectively (n = 5). Tumor formation was assessed and measured once a week. Animals were killed when the tumor reached the size of 1.5 cm. Tumor tissues were minced into < 1 mm pieces, dissociated in an enzymatic solution consisting of collagenase type 1 (1.5 mg/ml, Sigma), penicillin/streptomycin 20%, amphotericin 1% and DNase (1 mg/ml, Roche), and incubated at 37°C for 60 min with gentle agitation. The single cell suspensions were analyzed by flow-cytometry after staining with appropriate antibodies.

Leccia F., Del Vecchio L., Mariotti E., Di Noto R., Morel A.P., Puisieux A., Salvatore F, & Ansieau S. (2014). ABCG2, a novel antigen to sort luminal progenitors of BRCA1- breast cancer cells. Molecular Cancer, 13, 213.

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Establishing Endometrial and Ovarian Tumor Cell Cultures

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11 primary endometrial and ovarian tumor cell cultures were established from tumors of patients undergoing surgery at the Division of Gynecologic Oncology, UZ Leuven (Belgium). Tissue was washed with PBS supplemented with penicillin/streptomycin and fungizone, digested with collagenases type IV (1 mg/ml; Roche, Vilvoorde, Belgium) and DNAse I (0.1 mg/ml; Roche) in RPMI+ medium. Single cell suspensions were prepared by filtration through a 70-µm filter. Red blood cells were lysed using ammonium chloride (Stem Cell Technologies, Grenoble, France). Single cells were plated into a 25-cm (Parsons et al., 2012 (link)) culture flask. After 1–3 weeks, when cells reached 60–70% confluency, fibroblasts were removed using mouse anti-human CD90 (Clone AS02; Dianova, Hamburg, Germany) bound to Mouse Pan IgG Dynabeads (Life Technologies, Erembodegem, Belgium). Cell cultures were subsequently passaged at 70–90% confluency and banked at −80°C. Primary tumor cell cultures were grown in RPMI Medium 1640 supplemented with 20% fetal bovine serum (FBS), 2 mM L-Glutamine, 100U/ml penicillin, 100 μg/ml streptomycin, 1 μg/ml fungizone, and 10 μg/ml gentamicin (Life Technologies) up to 25 passages.

Zhao H., Thienpont B., Yesilyurt B.T., Moisse M., Reumers J., Coenegrachts L., Sagaert X., Schrauwen S., Smeets D., Matthijs G., Aerts S., Cools J., Metcalf A., Spurdle A., Amant F, & Lambrechts D. (2014). Mismatch repair deficiency endows tumors with a unique mutation signature and sensitivity to DNA double-strand breaks. eLife, 3, e02725.

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