All cellular models were incubated at 37 °C and maintained in an atmosphere containing 5% CO2 and in accordance with sterile cell culture practices. Cell lines were tested for Mycoplasma (MycoAlert™, cat# LT07-318, Lonza) every 2 weeks by using the manufacturer’s conditions. Cell lines were purchased from American Type Culture Collection (ATCC®). HEPG2 (ATCC, HB-8065, human liver carcinoma), BJ (ATCC, CRL-2522, normal human foreskin fibroblast), HEK-293 (ATCC, CRL-1573, transformed human embryonic kidney cell line); and murine models 3T3-L1 (ATCC, CL-173), J774.1 (ATCC# TIB-67) and MEF (ATCC# CRL-2907) cells. Cells were grown in medium (DMEM, EMEM, IMDM or Basal medium) containing 10% fetal bovine serum (FBS, Hyclone), 2 mmol L-glutamine (GlutaMAX™), and 1% penicillin/streptomycin (for murine models) or gentamicin sulfate (50 μg/mL for human preadipocytes) to densities recommended by ATCC. The 3T3-L1 murine fibroblast is an accepted cellular adipogenesis model since it competently undergoes adipogenesis after treatment with induction medium and experiments were conducted with passages 1–16. For The human subcutaneous preadipocytes (Poietics™, PT-5020, Lonza) and visceral preadipocytes (PT-5005, Lonza) were cultured according to the manufacturer’s instructions up to passage 3 (Figure S12 ).
Pt 5005
Manufactured by Lonza
Sourced in United States
The PT-5005 is a laboratory instrument designed for the processing and preparation of biological samples. It is a compact and versatile device that can be used for a variety of applications in the life sciences and biotechnology industries. The PT-5005 is capable of performing tasks such as homogenization, cell disruption, and sample mixing, among others. Its core function is to assist in the efficient processing and preparation of biological materials for subsequent analysis or experimentation.
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Lab products found in correlation
3 protocols using pt 5005
1
Cellular Model Maintenance and Characterization
2
Cryopreserved Human Preadipocyte Differentiation
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Cryopreserved human visceral preadipocytes (Lonza #PT-5005) were cultured according to the manufacturer's protocol. On reaching confluency, cells were harvested and seeded into 96-well opaque plates at 1 × 104 cells/well in 100 μL preadipocyte growth medium (Lonza PGM-2 with 10% FBS, 1% l -glutamine, and 1% gentamycin). Cells were incubated overnight at 37°C and 5% CO2. Adipogenic differentiation was induced by the addition of 100 μL/well of differentiation medium (PGM-2 plus 2X concentration adipogenic differentiation cocktail, Lonza PT-8002), for a final culture volume of 200 μL/well. Cells were incubated at 37°C and 5% CO2 for 5 days before the addition of compounds. Adipogenic differentiation was verified before starting the cell viability assay by the presence of lipid vacuoles. Immediately before compound addition, the culture medium in plates was reduced to a volume of 95 μL/well. RZL-012 was added and tested in triplicate at final assay concentrations of 0, 0.01, 0.03, 0.1, 1, 3, 10, 30, 100, and 300 μM, as described above for Wi38 cells.
3
Indirect Co-culture of Adipocytes and Macrophages
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Mouse preadipocytes (3T3-L1) and macrophages (RAW264.7) were obtained from American Type Culture Collection (ATCC) (Manassas, VA, USA), cultured, and induced to differentiate in a growth medium as previously reported [17 (link)]. For indirect co-culture of 3T3L1 and RAW264.7 cells, media from RAW264.7 cells were collected and sterile filtered (0.2 μm). The differentiation of 3T3-L1 cells was induced using MaCM supplemented with a differentiation cocktail (1% PS, 10% bovine calf serum (BCS), 0.5 mM 3-isobutyl1-methylxanthine, 1μM dexamethasone, and 1μg/mL insulin) for 2 days. The medium was then changed to MaCM with 10% BCS and insulin (1μg/mL) and was replaced every 2 days through day 8. At least 3 independent experiments were performed for each condition. Human primary preadipocyte cells from hSAT and hVAT (#PT5001 and #PT5005, Lonza, Walkersville, MD, USA) were processed and differentiated according to the supplier’s instructions in 2 independent experiments.
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