Co-culture experiments used a transwell system (3.0 μm pore size, Polyester (PET) Membrane; Corning Life Sciences, Lowell, MA, USA). For experiments that assessed 3T3-L1 adipocyte biology, 5 × 104 MCF-7 or MDA-MB-231 cells were seeded in the upper chamber with mature adipocytes in the bottom for the indicated times. Conversely, for experiments assessing cancer cell biology, 3T3-L1 adipocytes were grown then differentiated in the upper chamber with 5 × 104 breast cancer cells in the bottom. Adipocytes or cancer cells cultured alone served as controls.
Polyester pet membrane
Manufactured by Corning
Sourced in United States
The Polyester (PET) membrane is a lab equipment product manufactured by Corning. It is a microporous membrane made of polyester material that is used for various filtration and separation applications in laboratory settings. The core function of the Polyester (PET) membrane is to provide a physical barrier for the selective passage of certain substances, while retaining others, based on their size and/or other properties.
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Lab products found in correlation
4 protocols using polyester pet membrane
1
Transwell Co-culture of Adipocytes and Breast Cancer Cells
2
Transwell Co-culture Assay for Cell Proliferation
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A Transwell system (0.4 µm pore size, Polyester (PET) membrane; Corning, Corning, NY, USA) was employed. Either 5 × 104 MCF-7 or MDA-MB231 or BT-474 were cultured in the bottom surface of a 6-well plate. 3 × 105 Human ASCs were cultured in the upper transwell basket. The co-culture was performed for 5 days followed by a cell count using a Neubauer chamber. Wells without ASCs in the transwell were counted as control and compared with co-cultured cell counts.
3
Transwell Co-culture for Profibrotic Gene Expression
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A Transwell system (0.4 µm pore size, Polyester (PET) membrane; Corning, USA) was employed. Mouse or human adipose-derived stem cells were cultured in the top transwell basket compartment, while mouse (L929 cells, ATCC, USA) or human foreskin fibroblasts (ATCC, USA) were cultured in the lower compartment. Fibroblasts were grown to confluence and irradiated using a 10Gy radiation dose. Twenty-four hours post-irradiation 3 × 105 ASCs or 20 µM metformin or both were added to the upper transwell basket. Co-culture was maintained for 72 h. Cells were lysed, and total RNA was isolated using TRIzol (Sigma, USA) reagent following the manufacturer’s protocol. cDNA was synthesized and the expression of profibrotic gene battery (IL-6, CTGF, Collagens 1 and 3) was analyzed using gene-specific primers employing quantitative real-time PCR.
4
Transwell Co-culture for Fibrosis Mitigation
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A Transwell system (0.4 μm pore size, Polyester (PET) membrane; Corning, USA) was employed. Mouse adipose-derived stem cells or mouse bone marrow cells were cultured in the top transwell basket compartment, while mouse fibroblast or Ad-ECM was cultured in the lower compartment. A schematic of the co-culture approach is shown in Figure 5 C. For studying the role of HGF in fibrosis mitigation, mouse fibroblasts were grown to confluence and irradiated using a 10Gy radiation dose. Twenty-four hours post-irradiation 3 ×10 5 ASCs were added to the upper transwell basket. Co-culture was maintained for 48 hours. Cells were lysed, and total RNA was isolated using TRIzol (Sigma, USA) reagent following the manufacturer’s protocol. cDNA was synthesized and expression of pro-fibrotic gene battery (TGF β1, CTGF, NF-κB, IL-1, Collagens 1-6) was analyzed using gene-specific primers employing quantitative real-time PCR.
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