Cell invasion was performed using the Matrigel-coated film insert (8 mm pore size) fitted into 24-well invasion chambers (BD Biosciences, San Jose, CA, USA) [44 (link)]. After 24 h incubation, the filter inserts were removed from the wells and the cells on the top surface of the filter were removed using cotton swabs. The cells on the bottom surface of the filter were stained with 4′, 6-diamidino-2-phenylindole (DAPI) for 1 min, washed 3 times with PBS, and the cell number was counted under an Olympus fluorescence microscope.
24 well invasion chambers
Manufactured by BD
Sourced in United States
The 24-well invasion chambers are a laboratory equipment designed to assess the invasive potential of cells. The chambers consist of a 24-well plate with inserts that contain a layer of extracellular matrix, allowing for the study of cell migration and invasion through this barrier.
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10 protocols using 24 well invasion chambers
1
Matrigel-Based Cell Invasion Assay
2
Measuring Cell Invasion Using Matrigel
3
Cell Migration and Invasion Assay
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Cells were seeded in six-well plates, scratched with 10 µl pipet tips and cultured in 1× DMEM solution without serum. Then, the cells were washed three times with phosphate-buffered saline (PBS) and photographed by microscopy (Leica, Wetzlar, Germany) at 0 hours, 6 hours and 12 hours. The migration distance was measured by Image-Pro Plus software.
Cells were seeded in 24-well invasion chambers (BD Biosciences, New Jersey, USA) with a Matrigel-coated film insert (8 mm pore). The mixed solution was diluted to generate a 1× DMEM solution containing 10% serum. Two days later, cells on the bottom surface of the filter were subjected to staining with crystal violet for 15 min and then washed three times with PBS, and the cell number was counted under a microscope (Leica, Wetzlar, Germany).
Cells were seeded in 24-well invasion chambers (BD Biosciences, New Jersey, USA) with a Matrigel-coated film insert (8 mm pore). The mixed solution was diluted to generate a 1× DMEM solution containing 10% serum. Two days later, cells on the bottom surface of the filter were subjected to staining with crystal violet for 15 min and then washed three times with PBS, and the cell number was counted under a microscope (Leica, Wetzlar, Germany).
4
Quantifying Cell Invasion in Response to EGF
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Cells were seeded in 24-well invasion chambers (BD Biosciences, San Jose, CA, USA) with the Matrigel-coated film insert (8 mm pore). The mixed solution was diluted to give a 1× DMEM solution containing 10% serum. The cells were seeded in absence or presence of EGF (100 ng/ml) (chemokinesis). Two days later, cells on the bottom surface of the filter subjected to staining with 4′,6-diamidino-2-phenylindole for 1 min, then washed three times with PBS, and the cell number was counted under a fluorescence microscope (Olympus).
5
3D Matrigel Invasion Assay for Cell Migration
6
In vivo Invasion Screening Protocol
7
Cell Invasion Assay using Matrigel Chambers
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Cell invasion ability was detected using 24‐well invasion chambers (Coring) coated with Matrigel (BD Biosciences). The upper chamber was supplemented with serum‐free RPMI 1640 medium and 1 × 105 CRC cells, and the lower chamber was filled with full RPMI 1640 medium supplemented with 10% FBS (Hyclone). The chambers were then maintained at 37°C in 5% CO2 for 48 hours. Subsequently, the cells that were adhered to the undersurface of the filter membranes were removed, and the invaded cells were then fixed and stained with crystal violet for cell counting. An olympus microscope (CX41) was used to capture the digital photographs at five arbitrarily selected (non‐overlapped) fields. Then the average number of the invaded cells was counted.
8
Quantifying Glioma Cell Invasion Potential
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Cell invasion assays were performed using Transwell membranes coated with Matrigel (BD Biosciences, Franklin Lakes, NJ, USA). According to the manufacturer's instructions, 500 µl DMEM without serum was used to prehydrate the 24-well invasion chambers (8.0 µm; BD Biosciences) for 2 h at 37°C with 5% CO2. In DMEM without serum, the cells were seeded at a density of 5×104 cells/well in the upper chamber. The lower chamber was filled with 500 µl 20% FBS as a chemo-attractant. The non-migrating cells were removed from the top well using a cotton swab following incubation for 24 h. The cells in the bottom chamber were then fixed using 75% alcohol (37°C for 5 min) and subsequently stained the cells with 0.1% crystal violet (37°C for 1 min). The cells in each field of view migrating towards the bottom side across the filter were counted at a magnification of ×100 (light microscope) to quantify glioma cell migration. A total five fields of view were counted in this experiment. Three independent experiments were conducted.
9
Matrigel Invasion Assay for OC Cells
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Matrigel invasion assay was performed using 24-well invasion chambers purchased from BD Biosciences. Briefly, 3 × 104 OC cells were seeded to each insert in medium without FBS overnight. Medium containing 20% FBS was added to the bottom of the inserts. After incubation for 48 h, cells remaining above the insert membrane were removed by a sterile cotton swab. Cells invaded through the Matrigel to the bottom of the insert were fixed in 4% paraformaldehyde for 15 min and stained with 0.5% crystal violet. Penetrated cells were counted under a light Olympus microscope.
10
Cell Invasion Assay with EGF
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The indicated cells were seeded in 24‐well invasion chambers (BD Biosciences) with the Matrigel‐coated film insert (8 mm pore). The mixed solution was diluted to a 1 × DMEM solution containing 10% serum. The cells were cultured in the absence or presence of EGF (100 ng/mL) (chemokinesis). After 2 days, cells on the bottom surface of the filter were subjected to staining with DAPI for 1 minute, and then were washed three times with PBS, and the cell number was counted under a fluorescence microscope (Olympus).
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