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Transwell polycarbonate membrane filters

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Transwell polycarbonate membrane filters are laboratory equipment used for cell culture applications. They feature a porous polycarbonate membrane that allows for the study of cell migration, permeability, and transport between different compartments.

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Lab products found in correlation

Matrigel, by BD (1 mentions) Paraformaldehyde (pfa), by Merck Group (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)

Close spelling variants of this product

Transwell filters (307 citations) Transwell membranes (159 citations) Polycarbonate membrane (137 citations) Polycarbonate filters (105 citations) Polycarbonate transwell filters (71 citations) Polycarbonate membrane filters (34 citations) Transwell polycarbonate membranes (31 citations) Transwell membrane filter (21 citations) Polycarbonate transwell membrane filter (2 citations) Membrane filter (2 citations)

7 protocols using transwell polycarbonate membrane filters

1

Matrigel Invasion Assay for Cell Migration

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A Matrigel invasion assay was performed by using Transwell polycarbonate membrane filters with a pore size of 8.0 μm (Costar, Cambridge, MA). Briefly, Matrigel was thawed, diluted and placed into the upper chamber of a 24-well Transwell. After 4 h of incubation for Matrigel gelling, the cells were harvested, resuspended in 1% FBS media at a density of 106 cells/ml and placed onto the Matrigel. The lower chamber was filled with 600 μl of 10% FBS media. After a 6-h incubation at 37 °C, the nonmigrated cells on the upper surface of the membranes were eliminated by cotton swabs. The membranes were fixed with pure methanol and stained with a solution of crystal violet. Cells migrating to the substratum of the membrane were counted.
Cai Y.C., Yang H., Wang K.F., Chen T.H., Jiang W.Q, & Shi Y.X. (2020). ANGPTL4 overexpression inhibits tumor cell adhesion and migration and predicts favorable prognosis of triple-negative breast cancer. BMC Cancer, 20, 878.
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2

Isolation and Culture of Primary Proximal Tubule Cells

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The MOVAS-1 cells were purchased from Lonza (ATCC, Manassas, VA) and cultured in DMEM supplemented with 10% FBS, 2 mM glutamine, and 1% penicillin and streptomycin at 37°C in 5% CO2. Primary PT cells were isolated from C57/BL6 mice as previously described [39 (link)]. Briefly, kidneys were removed aseptically and dissected to obtain the cortical part of the kidney which were minced and incubated with agitation in digestion buffer containing 1% Worthington collagenase Type II and 0.25% soybean trypsin inhibitor. Cell suspensions were then passed through a series of mesh filters, resuspended in 45% Percoll and centrifuged at ~27,000× g for 15 min at 4°C. PTCs were grown onto Transwell polycarbonate membrane filters (Costar, Corning, NY) for 3 days until confluence as described previously [49 ].
Boadi E.A., Shin S, & Bandyopadhyay B.C. (2021). Tannic acid attenuates vascular calcification-induced proximal tubular cells damage through paracrine signaling. Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie, 140, 111762.
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3

Isolation and Culture of Human Endothelial Cells

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Human endothelial cells were isolated from umbilical cords and cultured as described previously (Gündüz et al., 2003 (link); Aslam et al., 2010 (link)). Briefly, the cells were cultured in PromoCell™ endothelial cell basal medium (PromoCell, Heidelberg, Germany) supplemented with 10% (vol/vol) fetal calf serum, 0.4% (vol/vol) endothelial growth supplement with heparin, 0.1 ng/ml human endothelial growth factor, 1.0 μg/ml hydrocortisone, 1 ng/ml bovine fetal growth factor, and 2% (vol/vol) penicillin/streptomycin in humidified atmosphere at 37°C, 5% CO2. Confluent monolayers were trypsinized in phosphate-buffered saline [PBS; composition in mM: 137 NaCl, 2.7 KCl, 1.5 KH2PO4, and 8.0 Na2HPO4, at pH 7.4, supplemented with 0.05% (wt/vol) trypsin, and 0.02% (wt/vol) EDTA] and seeded at a density of 7 × 104 cells/cm2 on 24 mm round Corning Transwell™ polycarbonate membrane filters (0.4 μm). Four days after seeding, the experiments were performed with confluent monolayers of passage #1.
Bosche B., Molcanyi M., Rej S., Doeppner T.R., Obermann M., Müller D.J., Das A., Hescheler J., Macdonald R.L., Noll T, & Härtel F.V. (2016). Low-Dose Lithium Stabilizes Human Endothelial Barrier by Decreasing MLC Phosphorylation and Universally Augments Cholinergic Vasorelaxation Capacity in a Direct Manner. Frontiers in Physiology, 7, 593.
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4

Caco-2 Cell Permeability Assay

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Caco-2 cells were seeded on Transwell polycarbonate membrane filters (0.4 μm pore size, Corning, Germany) at a density of 2.6 × 105 cells/cm2 and permeability tests performed as described by Hubatsch et al. [34 (link)]. Cells were submerged with Hank’s balanced salt solution (HBSS), and transepithelial electrical resistance (TEER) was measured before and after the experiment. Peptide solution (1 μM) was added to the apical chamber, and 300 μL aliquots were taken after 30, 60, 90, and 120 min of incubation. Samples were analyzed using LC1-MS1. Linear curve fitting was used to calculate the apparent permeability coefficient (Papp).
Wynendaele E., Debunne N., Janssens Y., De Spiegeleer A., Verbeke F., Tack L., Van Welden S., Goossens E., Knappe D., Hoffmann R., Van De Wiele C., Laukens D., Van Eenoo P., Vereecke L., Van Immerseel F., De Wever O, & De Spiegeleer B. (2022). The quorum sensing peptide EntF* promotes colorectal cancer metastasis in mice: a new factor in the host-microbiome interaction. BMC Biology, 20, 151.
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5

Cell Proliferation, Colony Formation, and Invasion Assays

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For cell proliferation assays, cells were seeded on 24-well plates. Cell numbers from triplicate wells were counted. For colony formation assays, 200 viable cells were seeded in 6 well plates and cultured for 10–14 days. The cell colonies were stained with crystal violet and then counted. To determine the invasive ability of PCa cells, the upper sides of the transwell polycarbonate membrane filters, with 8 mm pore size (Corning Inc, Corning, New York USA), were coated with diluted Matrigel (BD biosciences). 50,000 cells were seeded in the upper chamber with serum free media, and the bottom chamber of the apparatus contained media with 10% FBS. Cells were incubated for 48 hrs at 37°C. Following incubation, the cells that had invaded and attached to the lower surface of the membrane were fixed with 100% methanol and stained with 0.5% crystal violet. All experiments were repeated 3 times with cells grown at 37°C with 5% CO2. Cell numbers were counted and quantified in 5 randomly chosen macroscopic fields per membrane using an inverted microscope. WNT3A (cat no MAB1324) and IL-6 (cat no MAB-206) neutralizing antibodies were obtained from R&D Systems.
Nandana S., Tripathi M., Duan P., Chu C.Y., Mishra R., Liu C., Jin R., Yamashita H., Zayzafoon M., Bhowmick N.A., Zhau H.E., Matusik R.J, & Chung L.W. (2017). Bone metastasis of prostate cancer can be therapeutically targeted at the TBX2-WNT signaling axis. Cancer research, 77(6), 1331-1344.
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6

Quantifying A7r5 Cell Invasion

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To determine the invasive ability of A7r5 cells, the upper sides of the transwell polycarbonate membrane filters, with 8-μm pore size (Corning Inc.), were coated with 40 μg/ml collagen type I or 5 μg/ml fibronectin. Cells (20,000/well) in serum-free medium were seeded in the upper chamber, and the bottom chamber contained medium with 10% FBS. Cells were incubated for 16 h at 37 °C. Following incubation, the cells that had invaded and attached to the lower surface of the membrane were fixed with 4% paraformaldehyde and stained with 4′,6-diamidino-2-phenylindole (DAPI, diluted 1000× in PBS) (Sigma-Aldrich). Cell number was counted in five random fields per membrane under an inverted microscope.
Pai V.C., Lo I.C., Huang Y.W., Tsai I.C., Cheng H.P., Shi G.Y., Wu H.L, & Jiang M.J. (2018). The chondroitin sulfate moiety mediates thrombomodulin-enhanced adhesion and migration of vascular smooth muscle cells. Journal of Biomedical Science, 25, 14.
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7

Caco-2 Permeability Assay for Peptides

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Caco-2 cells were seeded on Transwell polycarbonate membrane filters (0.4 µm pore size) (Corning, Germany) at a density of 2.6 x 10 5 cells/cm 2 and the permeability study performed as described by Hubatsch et al. 31 . Cells were filled with Hank's Balanced Salt Solution (HBSS) and the TER values measured before and after the experiment. Peptide solution (1 µM) was added to the apical chamber and 300 µL aliquots taken after 30, 60, 90 and 120 min of incubation. Samples were analysed using LC1-MS1. Linear curve fitting was used to calculate the apparent permeability coefficient (Papp).
Debunne N., Wynendaele E., Janssens Y., Spiegeleer A.D., Verbeke F., Tack L., Welden S.V., Goossens E., Knappe D., Hoffmann R., Wiele C.V.d., Laukens D., Eenoo P.V., Immerseel F.V., Wever O.D., & Spiegeleer B.D. (2020). The Quorum Sensing Peptide EntF* Promotes Colorectal Cancer Metastasis in Mice: A New Factor in the Microbiome-Host Interaction.
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