Migration and invasion assays were performed as described previously (17 (link)). For Transwell migration assays, between 2.5×104 and 5.3×104 cells were plated in the upper chamber with the non-coated membrane (24-well insert; pore size, 8 µm; BD Biosciences). For invasion assays, 1.25×105 cells were plated in the upper chamber with a Matrigel-coated membrane (24-well insert; pore size, 8 µm; BD Biosciences). In the two assays, cells were plated in medium without serum or growth factors in the upper chamber, and medium supplemented with 10% FBS (Gibco; Thermo Fisher Scientific, Inc.) was used as a chemoattractant in the lower chamber. The cells were incubated at 37°C in a 5% CO2 incubator for 24 h and cells that did not migrate or invade through the pores were removed using a cotton swab. Cells on the lower surface of the membrane were stained with the Diff-Quick Staining set at 37°C for 30 min (Dade Behring; Siemens AG) and counted by a phase contrast microscope (Leica Microsystems, Inc.).
Non coated membrane 24 well insert
Manufactured by BD
Non-coated membrane (24-well insert) is a laboratory product that consists of a membrane insert with 24 wells. The membrane is uncoated, providing a neutral surface for various cell culture applications.
Automatically generated - may contain errors
2 protocols using non coated membrane 24 well insert
1
Cell Migration and Invasion Assays
2
Transwell migration and invasion assays
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For the transwell migration assays, 1×104 cells were plated in the top chamber containing a non-coated membrane (24-well insert; 8 mm pore size; BD Biosciences). For the invasion assays, 2×105 cells were plated in the top chamber containing a Matrigel-coated membrane (24-well insert; 8 mm pore size; BD Biosciences). For both assays, the cells were plated in the serum-free medium, and medium supplemented with 10% (v/v) serum was used as a chemoattractant in the lower chamber. The cells were incubated for 16 h at 37°C in a tissue culture incubator with 5% (v/v) CO2. After 16 h, the non-migrated/non-invading cells were removed from the upper sides of the transwell membrane filter inserts using cotton-tipped swabs. The migrated/invaded cells on the lower sides of the inserts were stained with Giemsa, and the cells were counted.
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