Fresh, healthy fallopian tube fimbriae were obtained from 3 donors at UPMC Magee-Women’s Hospital. Specimens were collected in a 50 ml of DMEM F12 Ham medium (Sigma-Aldrich) supplemented with 10% FBS (Gibco) and 1% Penn/Strep (Sigma-Aldrich). Upon receipt, FT tissues were processed under sterile tissue culture conditions as described earlier [30 (link),31 (link)] and cultured in Prigrow cell culture medium (Applied Biological Materials Inc., catalog #TM004) supplemented with 2% Ultroser G serum substitute (Pall France). To immortalize FT epithelial (FTE) cells, we used lentiviral transduction with SV40 large plus small T antigen (FTE-Tag) that suppresses TP53 signal transduction pathway thus mimicking the genetic makeup of STIC lesion [31 (link)]. Unmodified lentivirus was used in control cells. Recombinant lentiviral expression vector (Catalogue #CILV01) and an empty lentiviral expression vector (Catalogue # LV000) were commercially purchased from ALSTEM and used to transduce primary FTE cells in the presence of transduction reagent TransPlus following the manufacturer’s protocol [32 (link)–34 (link)].
Penn strep
Manufactured by Merck Group
Penn/Strep is a laboratory product that contains a combination of penicillin and streptomycin. It is commonly used in cell culture applications to prevent bacterial contamination.
Automatically generated - may contain errors
Lab products found in correlation
Lipopolysaccharides (lps), by Merck Group (2 mentions)
Dmem f12 ham medium, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Rnaeasy mini kit, by Qiagen (1 mentions)
Umuc3, by American Type Culture Collection (1 mentions)
Minimum essential medium (mem), by Thermo Fisher Scientific (1 mentions)
Sw780, by American Type Culture Collection (1 mentions)
Tryple express, by Thermo Fisher Scientific (1 mentions)
Vacutainer cpt tube, by BD (1 mentions)
Cd14 microbead, by Miltenyi Biotec (1 mentions)
5 protocols using penn strep
1
Immortalization of Fallopian Tube Epithelial Cells
2
Culture of Cell Lines and Primary B Cells
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3T3 cell lines, Ntera2, HepG2, GC-1-Spg, and GC-2-Spd(ts) cells were all cultured in DMEM with 10% FBS and penn/strep. Primary B220+ sorted splenic B cells were cultured in RPMI with 15% FBS, penn/strep, beta-mercaptol, and LPS (1 ug/ml from Sigma-Aldrich).
3
Transcriptomic Analysis of BmN4 Cells
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The Bombyx mori larval ovary-derived cell line BmN4 (RRID:CVCL_Z634) [7 (link)] was kindly provided by the Ketting group (Institute of Molecular Biology, Mainz, Germany)). Cells were cultured in Insect media IPL-40 (Pan Biotech) with 10% heat-inactivated FBS (Sigma) and 1x Penn-Strep (Sigma) at 27 °C.
For RNA-sequencing total RNA was extracted from 10 million cells using the RNAeasy Mini Kit (Qiagen) according to standard protocol. RNA integrity was tested by agarose gel electrophoresis and Bioanalyzer (RNA Nano Assay). RNA was quantified using Qubit.
For RNA-sequencing total RNA was extracted from 10 million cells using the RNAeasy Mini Kit (Qiagen) according to standard protocol. RNA integrity was tested by agarose gel electrophoresis and Bioanalyzer (RNA Nano Assay). RNA was quantified using Qubit.
4
Bladder Cancer Cell Culture Protocol
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Bladder cancer cell lines were recently purchased from ATCC: UMUC3 (# CRL-1749), T24 (# HTB-4), SW780 (# CRL-2169) and J82 (# HTB-1). Cells were cultured in MEM (Gibco) supplemented with 10% FBS (HyClone™) and Penn/Strep (Sigma) antibiotic solution under 37° C, 5% CO2 conditions. When confluency was reached, cells were passaged using TrypLE Express (Gibco) trypsinization solution.
5
Differentiation of Human Monocyte-Derived Dendritic Cells
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Two hundred milliliters of whole blood was collected in BD Vacutainer CPT tubes (cat# 362753) from healthy human donors in the MSD Volunteer Research Donor Program. CD14+ cells were isolated from PBMCs by positive selection using CD14 Microbeads (Miltenyi Biotec cat# 130-050-201). CD14+ cells were plated to non-adherent 6 well plates at 3 × 10^6 cells/mL in 3 mL of in X-Vivo 15 media, 10% autologous human serum, 1% PennStrep (Sigma cat# P4333), IL-4 (1000 IU/mL, R&D cat# 204-IL-010/CF), and GM-CSF (1000 IU/mL, R&D cat# 7954-GM-050/CF). Cytokines were replenished on days 2 and 4. Antigens were added to cells on day 4, and DCs were matured with a pulse of LPS (1 μg/mL, Sigma cat# L2880). Cells were harvested on day 5 for analysis, and markers for mature DCs were validated by flow cytometry.
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