Skin biopsies (∼5 × 5 × 5 mm) were taken from the upper medial arm of donor III.4 and an unrelated person who served as a control. These experiments took place at the Institute for Biomedical Ageing Research, Innsbruck, where the samples were immediately rinsed with phosphate buffered saline (PBS, pH 7.4) and incubated for 30 min in Dulbecco's modified Eagle's medium (DMEM, TFS) supplemented with penicillin (500 IU/ml), streptomycin (500 μg/ml) and Fungizone (12 μg/ml, all three Sigma-Aldrich). The dermis was separated from the epidermis after an overnight incubation at 37°C in DMEM supplemented with dispase (2.5 mg/ml, Roche), penicillin (100 IU/ml), and streptomycin (100 μg/ml). The hence separated dermis was then incubated at 37°C for 8 h in DMEM containing collagenase A (1.5 mg/ml, Roche), dispase (2.5 mg/ml), penicillin (100 IU/ml), and streptomycin (100 μg/ml). To arrive at a single cell suspension, the obtained solution was filtered twice with a 100-μm cell strainer (Sigma-Aldrich). The cells were counted and resuspended for expansion in DMEM containing 10% fetal bovine serum (FBS), 2 mM l -glutamine (both TFS), 100 IU/ml penicillin and 100 μg/ml streptomycin. Skin fibroblasts were maintained in DMEM as described before (43 (link)).
Streptomycin
Manufactured by Roche
Sourced in Germany, Switzerland, United States, Sweden
Streptomycin is a common antibiotic used in laboratory settings. It is a broad-spectrum antibiotic that is effective against a wide range of bacterial species. Streptomycin works by inhibiting protein synthesis in bacterial cells, leading to their death or inhibition of growth.
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Lab products found in correlation
Penicillin, by Roche (31 mentions)
Penicillin, by Thermo Fisher Scientific (7 mentions)
Penicillin, by Merck Group (5 mentions)
Streptomycin, by Thermo Fisher Scientific (5 mentions)
Mouse il 3, by Immunotools (4 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (4 mentions)
Streptomycin, by Merck Group (4 mentions)
Hepes, by Roche (4 mentions)
L glutamine, by Roche (4 mentions)
Transferrin, by Roche (4 mentions)
Rpmi medium, by Thermo Fisher Scientific (3 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Penicillin streptomycin, by Roche (3 mentions)
Insulin, by Roche (3 mentions)
Penicillin, by Immunotools (2 mentions)
Streptomycin, by Immunotools (2 mentions)
Imdm medium, by Thermo Fisher Scientific (2 mentions)
Roswell park memorial institute (rpmi), by Immunotools (2 mentions)
Cd8 t cell isolation kit, by Miltenyi Biotec (2 mentions)
Glutamax, by Merck Group (2 mentions)
41 protocols using streptomycin
1
Primary Dermal Fibroblast Isolation
2
Cultivation of Human Myeloid Leukemia Cell Lines
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Human myeloid leukemia cell lines (KG-1a, KG-1, HL-60, NB-4, ML-2, THP-1, MV-4-11, and U-937) were purchased from DSMZ (German Collection of Microorganisms and Cell Cultures, Braunschweig, Germany). Cells were cultured in RPMI medium (Life Technologies, Villebon-sur-Yvette, France) supplemented with 10% heat-inactivated fetal bovine serum (FBS, Life Technologies), 2 mM L-glutamine (Life Technologies), 100 U/mL penicillin (Sigma-Aldrich, Saint-Quentin-Fallavier, France), and 100 µg/mL streptomycin (Boehringer-Mannheim, Meylan, France) at 37 °C in fully humidified air and 5% CO2. For all experiments, cells were harvested from culture while in their exponential growth phase.
3
Humane Bleeding Procedure for Poultry
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All the bleeding procedures were approved by the Poultry Production Department at Kuwait Institute for Scientific Research and by the University of Reading, UK. This procedure follows the official decrees of the Ministry of Agriculture in Egypt relevant to animal welfare No. 27 (1967). These procedures are humane, respect animal rights and welfare and do not cause any suffering to animals. The practice of stunning and bleeding were used to slaughter the birds. To further elaborate, the bird was restrained and electrically stunned, it was then bled through a cut made in the neck. This procedure is aimed at minimizing the pain felt by the bird and the distress that could occur during bleeding out. It also immobilizes the bird to allow easy and accurate neck cutting (Gregory, 1995 ; Guarnieri et al., 2004 ). Approximately 15 ml of blood was collected in heparinized tubes into which additional heparin was added to prevent clotting. The thymus and spleen were separated carefully to avoid any contamination, all extra fat and tissues were removed, it was then weighed and placed in cell culture medium (CCM) on ice. Composition of CCM included RMPI-1640 supplemented with 10% fetal calf serum, 2 mmol/1 glutamine, and antibiotics (100 units/ml penicillin and 100 μg/ml streptomycin, Boehringer Mannheim, Germany).
4
Propagation of ORFV Virus in Lamb Testis Cells
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The ORFVNZ7 strain (Robinson et al., 1987 (link)) was propagated in primary lamb testis (LT) cells as described previously (Robinson et al., 1982 (link)). LT cells were maintained in minimum essential medium (MEM) (GIBCO, Invitrogen) and supplemented with FBS at 10% for growth and 5% for culture maintenance. LT cells were supplemented with PKS solution [kanamycin, (Roche Life Science); streptomycin. penicillin (Gibco)]. Cells were incubated at 37°C in a humidified 7% CO2 atmosphere.
5
Isolation and Culture of CD8+ T Cells
6
Preparation and Characterization of CAF from Transformed CD8+ T Cells
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293 T cells were obtained from the ATCC and cultured in DMEM, supplemented with 10% FBS, 2 mM Glutamine and 1 mM sodium pyruvate. TZM-bl cells were obtained through the NIH AIDS Research and Reference Reagent Program, Division of AIDS, NIAID, NIH: TZM-bl from Dr. John C. Kappes, Dr. Xiaoyun Wu and Tranzyme Inc. The CD8+ T cell line, TG, was previously established by Herpes virus saimiri (HVS)-transformation of CD8+ T cells from a chronically HIV-1 infected subjected from the MACS. TG cells were grown in RPMI with 20% FBS (100 nm filtered, Invitrogen Life Sciences, Carlsbad, CA), supplemented with 25 mM HEPES, Penicillin (100 U/mL) and Streptomycin (100 ug/mL) and rIL2 (50 U/mL, Roche Diagnostics). CAF from these transformed CD8+ T cells was prepared as described before [8 (link)]. Briefly, TG cells were cultivated for 14 days, after which, the cells were centrifuged at 300 g and the resulting supernatant was then further centrifuged at 4°C at the following speeds: 2000 g for 30 minutes, 6000 g for 20 minutes and 15000 g for 1 hour to remove other debris. This conditioned media was used for further investigations on CAF.
7
Stable cell lines expressing claudins
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HT-1080 cells stably expressing mock (empty vector), human CLDN-1 to -7 and mouse CLDN-5 were developed as described previously32 . MDCKII and P3U1 cells were purchased from ATCC (Manassas, VA). HT-1080 cells expressing human CLDN-5 mutants (D68E, T75A and S151T) and MDCKII cells expressing human or mouse CLDN-5 were prepared in a similar way as described previously32 .
HT-1080 and MDCKII cells were maintained in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% heat-inactivated fetal bovine serum (FBS) (v/v) (Nichirei Biosciences, Tokyo, Japan), 100 U/mL penicillin, and 100 µg/mL streptomycin (Nacalai Tesque, Kyoto, Japan). P3U1 cells were maintained in RPMI1640 medium supplemented with 10% heat-inactivated FBS, 100 U/mL penicillin, and 100 µg/mL streptomycin. All hybridomas were maintained in 20% heat-inactivated FBS, 10% BM Condimed H1 (Roche Diagnostics), 100 U/mL penicillin, and 100 µg/mL streptomycin. All cells were incubated under an atmosphere of 5% CO2 in air at 37 °C.
HT-1080 and MDCKII cells were maintained in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% heat-inactivated fetal bovine serum (FBS) (v/v) (Nichirei Biosciences, Tokyo, Japan), 100 U/mL penicillin, and 100 µg/mL streptomycin (Nacalai Tesque, Kyoto, Japan). P3U1 cells were maintained in RPMI1640 medium supplemented with 10% heat-inactivated FBS, 100 U/mL penicillin, and 100 µg/mL streptomycin. All hybridomas were maintained in 20% heat-inactivated FBS, 10% BM Condimed H1 (Roche Diagnostics), 100 U/mL penicillin, and 100 µg/mL streptomycin. All cells were incubated under an atmosphere of 5% CO2 in air at 37 °C.
8
Establishing 5-FU Resistant GC Cell Lines
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The GC cell lines MGC-803, HGC-27, BCG-823, MKN-28, MKN-45, and SGC-7901 were obtained from Shanghai Institute for Biological Sciences, Chinese Academy of Science. MGC-803-derived 5-FU-resistant sublines (MGC-803/5-FU) were induced by gradual exposure of 5-FU in culture medium. Briefly, MGC-803 cells were cultured in fresh medium without drugs for 24 h. Subsequently, the medium was changed and 0.01 μM 5-FU in complete medium was added. MGC-803 cells were exposed to 5-FU for 48–72 h; thereafter, the 5-FU-treatment medium was removed and cells were allowed to recover (in normal medium) for about 1 week. When cells reached 70% confluence, the treatment process was repeated for several times until they were stable (3–4 weeks). Once stable, cells were subjected 0.01, 0.1, 0.6 and finally 1.2 μM 5-FU treatment. Thereafter, the 5-FU-resistant cells were maintained in full medium supplemented with 1.2 μM 5-FU (to maintain 5-FU resistance). All GC cell lines were maintained at 37°C in a humidified atmosphere of 5% CO2 and 95% air in RPMI 1640 medium (Thermo Electron Corporation, Beijing, China) supplemented with 10% (v/v) fetal bovine serum (FBS; Life Tech, Mulgrave Vic, Australia) and penicillin/streptomycin (10,000 IU/ml penicillin and 20 mg/ml streptomycin; Roche, Swiss). The medium was changed twice per week.
9
Culturing Human Islets from Brain-Dead Donors
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Human islets from brain-dead, heart-beating donors, provided by the Nordic Network for Clinical Islet Transplantation (22–42 islets/donor, for donor data, Table 1 ), were individually cultured in CMRL 1066 supplemented with 10% (v/v) FCS, 2 mmol/L l -glutamine, 0.1 mg/mL streptomycin (Roche Diagnostics Scandinavia) and 5.5 or 20 mmol/L glucose. Experiments involving human islets were approved by the regional ethical board in Uppsala, Sweden.
Human islet donor characteristics.
| Donor 1 | M | 38 | 41.7 | 5.2 (33) |
| Donor 2 | F | 59 | 22.0 | 5.8 (40) |
| Donor 3 | F | 73 | 22.3 | 5.8 (40) |
| Donor 4 | F | 69 | 23.9 | 6.5 (47) |
Data received from the Nordic Network for Clinical Islet Transplantation.BMI, body mass index; HbA1C, glycated hemoglobin.
10
Isolating and Sorting Mouse Islets
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Mouse islets were isolated by collagenase digestion, hand-picked and incubated in groups of 100 overnight at 37°C (air/CO2, 95:5) in 5 mL RPMI-1640 medium (Sigma-Aldrich) supplemented with 2 mmol/L l -glutamine, 11.1 mmol/L glucose, 10% (v/v) fetal calf serum (FCS) and 0.1 mg/mL streptomycin (Roche Diagnostics Scandinavia) (29 (link)). The islets were thereafter dichotomously sorted in a fluorescence microscope. Microsphere-containing islets were size-matched to control islets without microspheres from the same mouse (8 (link), 9 (link)).
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