HEK-293T cells (CRL-3216, ATCC) and C3H/10T1/2 cells (CCL-226, ATCC) were cultured in DMEM (11965, Gibco) supplemented with 10% Fetal Bovine Serum (F4135, Sigma) and 1× penicillin–streptomycin (15140122, Gibco). Primary human renal proximal tubule epithelial cells (RPTECs) (CC-2553, Lonza) were cultured in Renal Epithelial Cell Growth Medium (CC-3190, Lonza) with supplements provided in the kit. Primary RPTECs were used in early passage. All cells were maintained in a humidified 5% CO2 atmosphere at 37°C unless otherwise specified.
Cc 2553
Manufactured by Lonza
Sourced in Switzerland
The CC-2553 is a laboratory instrument designed for cell counting and viability analysis. It provides accurate and reliable measurements of cell number and cell health. The core function of the CC-2553 is to assist researchers and scientists in monitoring cell cultures and evaluating the state of living cells.
Automatically generated - may contain errors
Lab products found in correlation
Cc 3190, by Lonza (2 mentions)
Ccl 226, by American Type Culture Collection (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Cc 2565, by Lonza (1 mentions)
Mycoalerttm mycoplasma detection kit, by Lonza (1 mentions)
Human il 6, by Toray (1 mentions)
Human il 17a, by Thermo Fisher Scientific (1 mentions)
H 6016, by Cell Biologics (1 mentions)
Human tnf α, by Thermo Fisher Scientific (1 mentions)
γ ggt activity colorimetric assay kit, by Merck Group (1 mentions)
Keratinocyte serum free medium, by Thermo Fisher Scientific (1 mentions)
Cc 2511, by Lonza (1 mentions)
Co2 incubator, by Panasonic (1 mentions)
8 protocols using cc 2553
1
Cell Culture Conditions for HEK-293T, C3H/10T1/2, and RPTECs
2
Culturing Human Renal Epithelial Cells
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3
Primary Renal Cell Culture Protocols
4
Dedifferentiation and Re-epithelialization of hRPTECs
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hRPTECs were purchased from Lonza (CC-2553; Basel, Switzerland), and were used between the fifth and seventh passages in renal epithelial basal medium supplemented with REGM Singlequots Bulletkit (CC-3190; Lonza). These cells were seeded onto collagen I-coated six-well plates at a density of 3 × 105 cells/well and cultivated until sub-confluent, and the medium was then replaced with serum-free medium for starvation. After starvation for 24 h, the hRPTECs were stimulated to dedifferentiate by 3 ng/mL of TGF-β1 for 48 h; thereafter, re-differentiation (re-epithelialization) was induced by incubation with TGF-β1-free fresh medium for 48 h, 72 h, or 96 h.
5
Cell Culture Protocol for Multiple Cell Types
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The normal human astrocyte (CC-2565), cardiac coronary artery endothelial cells (CC-7030), and renal proximal tubule epithelial cells (CC-2553) were obtained from Lonza (Basel, Switzerland). The cells were cultured in systems containing ABMTM Basal Medium (CC-3187) and AGMTM SingleQuotsTM Supplements (CC-4123), EBMTM-2 Basal Medium (CC-3156) and EGMTM-2 MV Microvascular Endothelial Cell Growth Medium SingleQuotsTM supplements (CC-4147), and REBMTM Basal Medium (CC-3191) and REGMTM SingleQuotsTM supplements (CC-4127), respectively. The TRF2 F/- Rosa26-CreERT2 cells were of the mouse embryonic fibroblast cell line (MEF, ATCC, Manassas, VA USA) and were immortalized by SV40-LT antigen and cultured in DMEM (ATCC® 30-2002™) supplemented with 10% fetal bovine serum (FBS; ATCC® 30-2020™), 2 mM L-glutamine (ATCC® 30-2214™), and 0.1 mM nonessential amino acids (NEAA, 100X; Gibco cat 11140050, Seoul, South Korea). TRF2 knockout could be accomplished by Cre recombinase transduction or 1 μM 4-hydroxytamoxifen (4-OHT) treatment for 7 days with the MEFs. All cells were mycoplasma-free before and during the experiments, as determined by a MycoAlert TM Mycoplasma Detection Kit from Lonza.
6
Renal Cell Responses to Cytokines
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Human primary kidney cells used in the experiments include renal proximal tubule epithelial cells (RPTEC; CC-2553, Lonza, Switzerland), human renal glomerular microvascular endothelial cells (HRGEC; ACBRI128, Cell Systems, Kirkland, WA), kidney fibroblasts (HKF; H-6016, Cell Biologics, Chicago, IL), and human renal mesangial cells (HRMC; #4200, ScienCell, Carlsbad, CA). The cells were plated in 96-well plates (1 × 10 4 cells/well) and stimulated with some combination of human IL-6 (100 ng/ml; Toray Industries) plus human soluble IL-6Rα (100 ng/ml; PeproTech), human IL-17A (50 ng/ml; PeproTech), recombinant human ORM1 (1,000 ng/ml; Prospecbio, Rehovot, Israel), and human TNFα (50 ng/ml; PeproTech) for 3, 6 or 24 hours after serum starvation. Soluble IL-6Rα was added, because non-immune cells normally express only IL-6 signal-transducing receptor subunit gp130. Recombinant ORM1 was inactivated by incubating at 100°C for 10 min. The cells were harvested, and total RNA was prepared for real-time PCR. For mechanistic analysis, RPTEC were immortalized using SV40 large T antigen. Immortalized RPTEC showed a similar response to primary RPTEC following cytokine stimulation (Supplementary Fig. 1). Human hepatoma Hep3B cells were stimulated overnight with IL-6/IL-6Rα, IL-17 or their combination.
7
Renal Proximal Tubule γ-GGT Assay
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iNephLOs, primary non-neoplastic human renal proximal tubular epithelial cells (RPTEC; Lonza, #CC-2553), hESCs (Day0), and corresponding control cell organoids (shown as Control) were homogenized in 200 μL of ice-cold GGT Assay Buffer. γ-GGT activity was determined using γ-GGT activity colorimetric assay kit (Sigma, MAK089) according to the manufacturer’s instructions.
8
Culturing Human Cell Lines
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Human proximal tubular cells (HK-2) obtained from the American Type Culture Collection were cultured in keratinocyte serum-free medium (Thermo Fisher Scientific, Waltham, MA) without antibiotics. Primary human RPTECs obtained from LONZA (CC-2553, lot #530068; Walkersvillle, MD) were cultured in REBM Renal Epithelial Cell Basal Medium (LONZA) without antibiotics. Primary human dermal fibroblasts obtained from LONZA (CC-2511, lot #21TL116652) were cultured in FBM Fibroblast Basal Medium (LONZA) without antibiotics. Serial cultures were maintained in 55-cm2 tissue culture dishes and incubated at 37°C in a humidified 5% CO2 atmosphere in a CO2 incubator (CO2 incubator, Panasonic, Newark, NJ).
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