PC9-BrM3 cells were transduced with dox-inducible shRNAs against either non-target control (Tet-shNTC), AXL (Tet-shAXL), or TAZ (Tet-shTAZ) as well as stable expression through lentiviral transduction of empty vector control or L1CAM cDNA (expressed in N174-MCS vector backbone). 72 h prior to seeding in Matrigel transwell invasion chambers (Corning), cells were treated with 500 ng/mL dox to induce shRNA expression. On the day of the assay, transwell filters were equilibrated for 2 h at 37°C with serum-free media prior to seeding 1x105 cells in 500 uL serum-free media containing 500 ng/mL dox (upper chamber). Complete culture media containing serum was added to the lower chamber to serve as a chemoattractant, and cells were allowed to invade for 24 h prior to fixation and staining with 0.5% crystal violet solution (0.5% w/v crystal violet, 25% v/v methanol, 75% v/v dH2O). The total number of invaded cells from eight non-overlapping brightfield images (10X) per condition from three independent experiments were counted and the means from each condition were analyzed in GraphPad (n=3).
Matrigel transwell invasion chambers
Manufactured by Corning
Matrigel transwell invasion chambers are laboratory equipment used to assess the invasive potential of cells. The chambers consist of a porous membrane coated with a layer of Matrigel, a reconstituted basement membrane matrix. Cells are seeded onto the Matrigel, and their ability to migrate through the membrane is measured, providing insights into the cells' invasive characteristics.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using matrigel transwell invasion chambers
1
Matrigel Transwell Invasion Assay
2
Matrigel Transwell Invasion Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
100,000 cells/well were seeded into Matrigel Transwell Invasion chambers (Corning #354480) in serum-free media, with serum-containing media in bottom chambers. Cells were incubated at 37°C for 24 hours, then the top chambers were scrubbed to remove cells that had not invaded. Chambers were then fixed in 70% ethanol for 10 minutes, stained in 0.2% Crystal Violet for 10 minutes, and left to dry overnight. Brightfield images of each insert were acquired at 2X magnification on an Olympus IX51, and the ImageJ Color Inspector 3D plugin was used to quantify the percent coverage of purple pixels per insert image. Assays were performed in technical triplicate and biological triplicate. Multiple comparisons ANOVA was conducted on the data in GraphPad Prism 9. For assays with siRNA-treated cells, cells were seeded into chambers 48 hours post-transfection. For assays with IFN-α-treated cells, cells were seeded into chambers either with no prior IFN-α treatment or with 48 hours pre-treatment with IFN-α, and were seeded in serum-free media containing 100 IU/ml of IFN-α.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!