T. gondii strains ME49 and Prugniaud were maintained in Vero cells in RPMI (Nacalai Tesque) supplemented with 2% heat-inactivated fetal bovine serum (FBS; JRH Bioscience), 100 U/mL penicillin, and 0.1 mg/mL streptomycin (Nacalai Tesque), as previously described (Ma et al., 2014 (link)). HFFs were maintained in RPMI (Nacalai Tesque) supplemented with 2% heat-inactivated FBS (JRH Bioscience), 100 U/mL penicillin, and 0.1 mg/mL streptomycin (Nacalai Tesque). IMR-32 cells were maintained in MEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 1% non-essential amino acids (Nacalai Tesque), 100 U/mL penicillin, and 0.1 mg/mL streptomycin. A172 cells were maintained in DMEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 100 U/mL penicillin, and 0.1 mg/mL streptomycin. U251-MG cells were maintained in EMEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 100 U/mL penicillin, and 0.1 mg/mL streptomycin.
Roswell park memorial institute (rpmi)
Manufactured by Nacalai Tesque
Sourced in Japan
RPMI is a common cell culture medium used for the in vitro cultivation of various cell types, including primary cells, established cell lines, and hybridomas. It provides a balanced mixture of nutrients, vitamins, and other essential components to support the growth and maintenance of cells.
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Lab products found in correlation
Penicillin, by Nacalai Tesque (5 mentions)
Streptomycin, by Nacalai Tesque (5 mentions)
Dulbecco modified eagle medium (dmem), by Nacalai Tesque (5 mentions)
Fetal bovine serum (fbs), by Merck Group (4 mentions)
Fetal calf serum (fcs), by Nacalai Tesque (2 mentions)
Penicillin, by Meiji Seika Pharma (2 mentions)
Streptomycin, by Meiji Seika Pharma (2 mentions)
Penicillin, by R&D Systems (2 mentions)
Minimum essential medium (mem), by Nacalai Tesque (1 mentions)
Non essential amino acids (neaa), by Nacalai Tesque (1 mentions)
E mem, by Nacalai Tesque (1 mentions)
Streptomycin sulfate, by Nacalai Tesque (1 mentions)
Penicillin g, by Nacalai Tesque (1 mentions)
Iscove modified dulbecco media (imdm), by Nacalai Tesque (1 mentions)
Fetal bovine serum (fbs), by Nacalai Tesque (1 mentions)
Penicillin streptomycin solution, by Fujifilm (1 mentions)
Fetal bovine serum (fbs), by Biosera (1 mentions)
Neon system, by Thermo Fisher Scientific (1 mentions)
Streptomycin penicillin streptomycin mixed solution, by Nacalai Tesque (1 mentions)
B16f10 murine melanoma cells, by American Type Culture Collection (1 mentions)
17 protocols using roswell park memorial institute (rpmi)
1
Culturing and Maintaining T. gondii Strains and Human Cell Lines
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T. gondii strains ME49 and Prugniaud were maintained in Vero cells in RPMI (Nacalai Tesque) supplemented with 2% heat-inactivated fetal bovine serum (FBS; JRH Bioscience), 100 U/mL penicillin, and 0.1 mg/mL streptomycin (Nacalai Tesque), as previously described (Ma et al., 2014 (link)). HFFs were maintained in RPMI (Nacalai Tesque) supplemented with 2% heat-inactivated FBS (JRH Bioscience), 100 U/mL penicillin, and 0.1 mg/mL streptomycin (Nacalai Tesque). IMR-32 cells were maintained in MEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 1% non-essential amino acids (Nacalai Tesque), 100 U/mL penicillin, and 0.1 mg/mL streptomycin. A172 cells were maintained in DMEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 100 U/mL penicillin, and 0.1 mg/mL streptomycin. U251-MG cells were maintained in EMEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 100 U/mL penicillin, and 0.1 mg/mL streptomycin.
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T. gondii strains ME49 and Prugniaud were maintained in Vero cells in RPMI (Nacalai Tesque) supplemented with 2% heat-inactivated fetal bovine serum (FBS; JRH Bioscience), 100 U/mL penicillin, and 0.1 mg/mL streptomycin (Nacalai Tesque), as previously described (Ma et al., 2014 (link)). HFFs were maintained in RPMI (Nacalai Tesque) supplemented with 2% heat-inactivated FBS (JRH Bioscience), 100 U/mL penicillin, and 0.1 mg/mL streptomycin (Nacalai Tesque). IMR-32 cells were maintained in MEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 1% non-essential amino acids (Nacalai Tesque), 100 U/mL penicillin, and 0.1 mg/mL streptomycin. A172 cells were maintained in DMEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 100 U/mL penicillin, and 0.1 mg/mL streptomycin. U251-MG cells were maintained in EMEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 100 U/mL penicillin, and 0.1 mg/mL streptomycin.
2
MCF-7 Breast Cancer Cell Culture
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The human breast adenocarcinoma cell line MCF-7 was cultured in RPMI-1640 medium (RPMI, Nakalai Tesque; #30264) supplemented with 10% fetal calf serum (FCS), 100 units/ ml penicillin G (Nakalai Tesque; #26239-42), and 100 µg/ml streptomycin sulfate (Nakalai Tesque; #33204-92). Cells were maintained at 37 °C in humidified atmosphere containing 5% CO2.
In the experiments for culturing in glutamine-free medium (−Gln), MCF-7 cells were cultured in RPMI-1640 medium without L-glutamine (RPMI 1640 without L−Gln, Nakalai Tesque; # 05176-25) supplemented with 10% fetal calf serum (FCS), 100 units/ ml penicillin G, and 100 µg/ml streptomycin sulfate. Glutamine (Nakalai Tesque; #16948-04; 200mM-L-Glutamine Stock Solution) was added to glutamine-free medium with the same concentration to that in the standard RPMI-1640 medium (RPMI, Nakalai Tesque; # 30264) to produce glutamine-free medium with added glutamine (−Gln/+Gln).
In the experiments for culturing in glutamine-free medium (−Gln), MCF-7 cells were cultured in RPMI-1640 medium without L-glutamine (RPMI 1640 without L−Gln, Nakalai Tesque; # 05176-25) supplemented with 10% fetal calf serum (FCS), 100 units/ ml penicillin G, and 100 µg/ml streptomycin sulfate. Glutamine (Nakalai Tesque; #16948-04; 200mM-L-Glutamine Stock Solution) was added to glutamine-free medium with the same concentration to that in the standard RPMI-1640 medium (RPMI, Nakalai Tesque; # 30264) to produce glutamine-free medium with added glutamine (−Gln/+Gln).
3
Culturing Toxoplasma gondii and Cell Lines
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All T. gondii strains were maintained in Vero cells in RPMI (Nacalai Tesque) supplemented with 2% heat-inactivated FBS (JRH Bioscience), 100 U/ml penicillin, and 0.1 mg/ml streptomycin (Nacalai Tesque), as previously described (55 (link)). HAP1 cells were maintained in IMDM (Nacalai Tesque) containing 10% heat-inactivated FBS, and 100 U/ml penicillin, and 0.1 mg/ml streptomycin. HFFs, Huh7 cells were maintained in RPMI (Nacalai Tesque) containing 10% heat-inactivated FBS, and 100 U/ml penicillin, and 0.1 mg/ml streptomycin. HeLa cells and MEF cells were maintained in DMEM (Nacalai Tesque) containing 10% heat-inactivated FBS, 100 U/ml penicillin, and 0.1 mg/ml streptomycin.
4
Cell Culture Maintenance Protocol
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hNF, mNF, BOSC, WM, MDA, PC9, B16F10, 4T1, and Vn1919 were grown and maintained as monolayer cultures at 37 °C in 5% CO2 humidified atmosphere, using Dulbecco’s modified Eagle’s medium (high glucose with l -glutamine and phenol red, Fujifilm Wako Pure Chemical Corporation) supplemented with 1 vol% penicillin–streptomycin solution (×100) (Fujifilm Wako Pure Chemical Corporation) and 10 vol% FBS (Sigma-Aldrich Co., Llc.). HL-60 and K562 were grown and maintained as floating cultures at 37 °C in 5% CO2 humidified atmosphere, using Roswell Park Memorial Institute medium (RPMI, Nacalai Tesque, Inc.) supplemented with 1 vol% penicillin–streptomycin solution and 10 vol% FBS. OVMANA was grown and maintained as monolayer cultures at 37 °C in 5% CO2 humidified atmosphere, using RPMI, Nacalai Tesque, Inc. supplemented with 1 vol% penicillin–streptomycin solution and 10 vol% FBS. The cells were sub-cultured every 4–6 days with or without trypsin solution (0.5 w/v% trypsin-5.3 mmol/L EDTA・4Na solution without phenol red (×10), Fujifilm Wako Pure Chemical Corporation).
5
Human OCCC Cell Lines Cultivation
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The human OCCC cell lines OVTOKO, RMGV, and OVISE were obtained from the Health Science Research Resources Bank of Osaka, Japan. OVTOKO and OVISE were cultured in RPMI (Nacalai Tesque) supplemented with 10% fetal bovine serum (FBS) (Biosera). RMGV was cultured in Ham‐F (Nacalai Tesque) supplemented with 10% FBS (Biosera, Nuaille, France). All cell lines were certified to be free of fungal, bacterial, and mycoplasma contaminations by the cell bank.
6
Culturing Toxoplasma gondii in Mice
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6–8-wk-old BALB/c mice were obtained from SLC. All animal experiments were conducted with the approval of the Animal Research Committee of Research Institute for Microbial Diseases in Osaka University. ME49, RHΔhxgprtΔku80 and its derivatives of T. gondii were maintained in Vero cells by biweekly passage in RPMI (Nacalai Tesque) supplemented with 2% heat-inactivated FCS (JRH Bioscience), 100 U/ml penicillin, and 0.1 mg/ml streptomycin (Nacalai Tesque). 293T cells and fibroblasts were maintained in DMEM (Nacalai Tesque) containing 10% heat-inactivated FCS and antibiotics.
7
Murine Melanoma and Ovarian Cancer Cell Culture with NHEJ Inhibition
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B16F10 murine melanoma cells were purchased from the American Type Culture Collection. B16F10 cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) (Nacalai Tesque) supplemented with heat inactivated 10% fetal bovine serum (FBS) (Biowest), 100 U/mL penicillin, and 100 µg/mL streptomycin (penicillin–streptomycin Mixed Solution) (Nacalai Tesque). ID8-luc2 murine ovarian cancer cells were a gift from the Department of Obstetrics and Gynecology, Osaka University Graduate School of Medicine. ID8 luc2 cells were cultured in Roswell Park Memorial Institute (RPMI) (Nacalai Tesque) supplemented with heat inactivated 10% FBS, 100 U/mL penicillin and 100 µg/mL streptomycin, at 37 °C and 5% CO2 in the CO2 incubator. NHEJ was inhibited by 1 µM NHEJ inhibitor-SCR7 following manufacture’s instructions (Xcess Biosciences Inc.) after electroporation using the NEON system (Thermo Fisher Scientific).
8
Establishing LVRN-Transfected Swan71 Cells
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The human acute monocytic leukemia cell line THP-1 (RRID: CVCL_0006) was purchased from ATCC. To activate THP-1 cells, we added Phorbol12-myristate13-acetate (PMA, Sigma-Aldrich) and cultured them for 48h. Swan71 (RRID: CVCL_D855) was gifted by Prof. Mor G, which is human telomerase reverse transcriptase-transfected immortalized first-trimester trophoblast cell line.18 (link) Both cell lines were authenticated by STR profiling.
To establish LVRN-transfected Swan71 cells (Swan71_LVRN), hLVRN-His cDNA was inserted into pTargeT vector (Promega). Then, r-LVRN-His/pTargeT (500 ng) was cleaved and linearized with Psp1406I and transfected into Swan71 cells using FuGeneHD (Promega). After 48 h of gene transfer, a stable expression strain was established and isolated using a medium containing 500 μg/mL G418. For controls, the non-inserted pTargeT vector was transfected into Swan71 cells (Swan71_NEO). Cell lines were maintained at 37°C under 5% CO2 in RPMI (Nacalai Tesque, Kyoto, Japan) supplemented with 10% FBS (Sigma-Aldrich), 100 g/mL streptomycin, and 100 IU/mL penicillin. Mycoplasma infections were detected regularly, and all experiments were performed under mycoplasma-free conditions.
To establish LVRN-transfected Swan71 cells (Swan71_LVRN), hLVRN-His cDNA was inserted into pTargeT vector (Promega). Then, r-LVRN-His/pTargeT (500 ng) was cleaved and linearized with Psp1406I and transfected into Swan71 cells using FuGeneHD (Promega). After 48 h of gene transfer, a stable expression strain was established and isolated using a medium containing 500 μg/mL G418. For controls, the non-inserted pTargeT vector was transfected into Swan71 cells (Swan71_NEO). Cell lines were maintained at 37°C under 5% CO2 in RPMI (Nacalai Tesque, Kyoto, Japan) supplemented with 10% FBS (Sigma-Aldrich), 100 g/mL streptomycin, and 100 IU/mL penicillin. Mycoplasma infections were detected regularly, and all experiments were performed under mycoplasma-free conditions.
9
Mouse model for Toxoplasma gondii study
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C57BL/6NCrSlc (C57BL/6N) mice were purchased from SLC. Mice lacking IFN-γR have been previously described (30 (link)). All animal experiments were conducted with the approval of the Animal Research Committee of Research Institute for Microbial Diseases at Osaka University. RHΔhxgprtΔku80 and its derivatives of T. gondii were maintained in Vero cells by bi-weekly passage in RPMI (Nacalai Tesque) supplemented with 2% heat-inactivated fetal calf serum (FCS; JRH Bioscience), 100 U/mL penicillin and 0.1 mg/mL streptomycin (Nacalai Tesque). Mouse embryonic fibroblasts were maintained in Dulbecco’s modified Eagle medium (DMEM; Nacalai Tesque) supplemented with 10% heat-inactivated FCS, 100 U/mL penicillin, and 0.1 mg/mL streptomycin (Nacalai Tesque).
10
Inducing Epithelial-Mesenchymal Transition in Colon Cancer Cells
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Human colon cancer cell lines HT29 and DLD‐1 were obtained from the American Type Culture Collection (ATCC) and were maintained in DMEM (Nacalai Tesque) and RPMI (Nacalai Tesque) supplemented with 10% FBS (Thermo Fisher Scientific), respectively. EMT was induced as previously described using epidermal growth factor (Sigma‐Aldrich) at 20 ng/mL and basic fibroblast growth factor (Sigma‐Aldrich) at 10 ng/mL in the absence of FBS.8 (link) In some experiments, mithramycin A (Sigma‐Aldrich) was added into the culture medium at a final concentration of 200 nmol/L 1 hour before transfection of reporter constructs in luciferase reporter assays or 24 hours before extraction of total RNA.
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