Melanoma cell invasion was determined using the transwell assay. Transfected A375 cells were placed on the top of the Matrigel-coated invasion chambers (BD Biosciences, USA) with serum-free DMEM. In the lower chamber, we added 500 μl of DMEM containing 10% foetal bovine serum as a chemoattractant. After 24 h, cotton swabs were used to remove the non invasive cells. The invading cells were fixed with 95% ethanol, stained with 0.1% crystal violet, and counted and photographed under an inverted microscope (×100). The scratch wound assay was used to assess cell migration ability. First, transfected A375 cells were seeded into 6-well plates. After 24 h, cell layers were scratched using a 200 μl pipette tip to form wound gaps, and then the cells were maintained in 10% FBS-supplemented DMEM. The cells were photographed (0 and 24 h) under an inverted microscope to record the wound width. The experiments were independently repeated in triplicate.
Matrigel coated invasion chamber
Manufactured by BD
Sourced in United States
Matrigel-coated invasion chambers are laboratory equipment designed to assess the invasive capabilities of cells. They consist of a membrane coated with Matrigel, a reconstituted basement membrane extract, which serves as a barrier to mimic the extracellular matrix. Cells are seeded on the chamber, and their ability to migrate through the Matrigel-coated membrane is measured, providing an indication of their invasive potential.
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Lab products found in correlation
Non coated migration chambers, by BD (4 mentions)
Ckx31 41, by Olympus (4 mentions)
Dmi4000b, by Leica (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Fluoroblock cell culture insert, by BD (2 mentions)
Nucblue live cell stain readyprobes reagent, by Thermo Fisher Scientific (1 mentions)
Sp5 confocal microscope, by Leica (1 mentions)
Fetal calf serum (fcs), by Thermo Fisher Scientific (1 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions)
24 transwell plate, by Cell Biolabs (1 mentions)
Matrigel basement membrane matrix, by BD (1 mentions)
Dmil inverted light microscope, by Leica (1 mentions)
Matrigel, by BD (1 mentions)
Inverted microscope, by Leica (1 mentions)
Mitomycin c, by Roche (1 mentions)
Upper transwell chamber, by Corning (1 mentions)
Transwell insert, by Corning (1 mentions)
Eclipse ti microscope, by Nikon (1 mentions)
Eclipse ts100 microscope, by Nikon (1 mentions)
Fluorescence mounting medium, by Agilent Technologies (1 mentions)
77 protocols using matrigel coated invasion chamber
1
Evaluating Melanoma Cell Invasion and Migration
2
Matrigel-based Invasion Assay for GBM Cells
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Matrigel-coated invasion chambers (8 μm, BD Biosciences, San Jose, CA, U.S.A.) were prepared according the manufacturer’s instructions. Transfected GBM cells were seeded in the upper compartment and allowed to migrate for 24 h in an incubator. Cells were seeded in 1% FBS DMEM medium in the upper chamber and 10% FBS medium in the well below the insert. The cells that had invaded the lower surface were fixed with 4% paraformaldehyde and stained with 0.1% crystal violet. The number of cells that had invaded through the Matrigel was counted in six random fields for triplicate membranes under the microscope.
3
Quantifying Cell Invasion Potential
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In vitro invasion assays were performed as previously described (Bravo-Cordero et al., 2011 (link)). In brief, 5 × 105 cells were suspended in low (0.25%)-serum culture medium and plated in duplicate in the top well of growth factor–reduced Matrigel-coated invasion chambers (8-µm pore size; BD BioCoat; 354483; BD). Normal serum-containing media (5% serum) were placed in the lower chamber, and cells were allowed to invade for 24 h at 37°C. (For MDA–MB-231 cells, the low-serum culture media contained 0.5% serum, whereas normal growth media contained 10% serum.) The assay was fixed with 3.7% PFA for 20 min and then stained with DAPI or NucBlue Live Cell Stain ReadyProbes reagent (R37605; Molecular Probes) to visualize nuclei. Four to six random fields were acquired at 20× magnification on an SP5 confocal microscope (Leica Microsystems). The number of invaded cells was counted manually with ImageJ software and normalized to the total number of cells present in the insert. Data are means of three independent experiments for each condition.
4
TGF-β Induced Migration and Invasion Assay
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Prior to plating into migration chambers, cells were treated with either shSLK or a sh-scrambled adenovirus for 48 hours, as described above. The cells were treated with 2ng/mL of TGF-β for 48 hours and 5.0 × 104 cells were plated into the top part of each fibronectin-coated chamber (8 μm pores; Fisher Scientific, USA). Haptotaxis assays were run in 0.2% FBS DMEM media. Chemotactic assays were run with the bottom chamber containing TGF-β. The migration chambers were placed at 37°C at 5% CO2 for six hours. Each well was washed and fixed in 4% PFA for ten minutes. The washed membranes were removed and placed cell-side up onto a microscope slide. Each membrane was covered in ProLong Gold antifade reagent with DAPI and the migrated cells were enumerated using fluorescence microscopy. The assays were done in triplicate wells the cells were counted from five random fields per membrane. Invasion assays were performed as above but for 24 hours using Matrigel-coated invasion chambers (BD Biosciences, Canada).
5
Quantifying Cell Invasion via Transwell Assay
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Cell invasion was examined using Transwell assays. Following incubation for 48 h, 3×104 cells were transferred to the top of the Matrigel-coated invasion chambers (BD Biosciences) in serum-free DMEM. DMEM containing 10% FBS was added to the lower chamber. After 24 h, the non-invading cells were removed and the invading cells were fixed using 95% ethanol, stained with 0.1% crystal violet, and photographed at a magnification of ×100 under an inverted phase contrast microscope (Olympus CKX31/41; Olympus, Tokyo, Japan). The experiments were repeated three times.
6
Matrigel-Based Invasion Assay
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Invasion assay was carried out using Matrigel-coated invasion chambers (BD Biosciences). Briefly, cells were resuspended in culture medium without FBS at the concentration of 6 × 104 cells/ml and 500 μl of the cell suspension was seeded on the upper chamber. The lower chamber was filled with 700 μl of the culture medium without cells and 10% FBS was added as a chemoattractant. Cells were incubated with or without rhBMP4 for 18 h in a humidified tissue culture incubator, at 37°C, 5% CO2 atmosphere. After removing non-invasive cells with a cotton swab. Invasive cells adhering to membrane of the upper chamber were fixed with methanol 100% for 1 min and then stained with a Borax1% toluidine 1% solution for 1 min. Number of cells on the membrane was counted under a light microscope at 40X magnification.
7
Quantifying Cell Invasion Potential
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Cell invasion was determined using Transwell assays. Briefly, TPX2-overexpressing plasmid and siRNA to knockdown TPX2 expression were transfected into cells, and 48 h later, 3×104 cells were transferred to the top of Matrigel-coated invasion chambers (BD Biosciences) in serum-free DMEM. Next, DMEM containing 10% FBS was added to the lower chamber, and 24 h later, the non-invading cells were removed, while the invading cells were fixed with 95% ethanol and stained with 0.1% crystal violet. Images were captured in a light microscope under ×100 magnification. Experiments were performed three independent times.
8
Cell Invasion and Migration Assay
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Cell invasion and migration was evaluated by using transwell assay as previously described.31 (link) And, 3×105 Ec-9706 and TE-10 cells transfected with si-GIHCG, miR-29p-3p mimics, miR-29p-3p inhibitor and corresponding negative controls were transferred to the upper Matrigel-coated or not Matrigel-coated invasion chambers (BD Biosciences, San Jose, CA, USA) in a serum-free DMEM medium, and DMEM medium containing 10% FBS was added to the lower chambers. After 24 h, non-migrated or non-invaded cells on the upper surface were removed, and the migrating or invading cells on the underside surface were fixed with 4% paraformaldehyde, stained with 0.1% crystal violet, and then imaged in 10 random fields under a microscope.
9
Tetracycline-Induced Cell Invasion Assay
10
Evaluating Breast Cancer Cell Invasion
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In vitro invasion potential was evaluated using 24-well Matrigel-coated invasion chambers with a 8 μm pore size (BD Biosciences). For MCF-7 cells, 2.5 x 104 cells were allowed to migrate for 96 h using 10 % foetal bovine serum as a chemoattractant. Cells that reached the lower surface were stained with crystal violet. At least three independent experiments were performed with triplicates for each condition. Cells were counted in eight randomly selected microscopic fields. In the case of MDA-MB-231 cells, invasion was evaluated after 24 h.
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