The migration ability of VSMCs was measured in the Boyden chamber assay and wound-healing assay. In the Boyden chamber assay, the VSMCs were planted into a serum-free medium in the upper 24-well transwell chamber with 8 μm pores (Merck kGaA, Darmstadt, Germany). Then, 10% FBS was injected into the lower chamber, with the cellular migration measured 24 h later. The VSMCs that had not migrated from the upper chamber were scratched using cotton swabs, and those that had migrated into the lower chamber were subjected to 1% crystal violet staining. Five fields were randomly selected from each chamber, and the cells were counted. In the wound-healing assay, the VSMCs (2 × 105 cells/mL) were seeded into 6-well plates, and cultured for 24 h until reaching an 80–90% confluence. Then, the cells were scratched with the tip of a standard 1 mL pipette, until a wound was observed in the central area. Once the cellular debris was washed using PBS, the medium was freshly replenished. Photos of the healing process were captured at 0 and 24 h, using an inverted microscope (Axio Vert. A1, Zeiss, Oberkochen, Germany). The distances among migrated cells were photographed and figured.
Upper 24 well transwell chamber
Manufactured by Merck Group
Sourced in Germany, United States
The Upper 24-well transwell chamber is a laboratory equipment used for cell culture and research applications. It consists of a 24-well plate with a porous membrane insert that allows for the study of cell migration, invasion, and permeability. The core function of this product is to facilitate the separation and analysis of cells or molecules between the upper and lower chambers of the well.
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Lab products found in correlation
2 protocols using upper 24 well transwell chamber
1
Measuring VSMC Migration Capacity
2
Transwell Assay for Migration and Invasion
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Transwell assay was performed to detect the migration and invasion abilities of MM cells. Firstly, Transwell Assay was performed to detect the migration and invasion abilities of A375 and G361 transfected with miR-146a mimics or miR-146a inhibitor. At the same time, the migration and invasion abilities of A375 cells transfected with SMAD4 and miR-146a mimics were also detected by Transwell Assays. Melanoma cells A375 and G361 were seeded into 6-well plates with the different transfection treatments. 48 h after the transfection, the cells were harvested and re-suspended with DMEM culture medium with FBS. A total of 1×105 cells/ml was prepared in DMEM, 150 µl cell suspensions was added into the upper 24-well transwell chamber (EMD Millipore) with or without Matrigel (Clontech Laboratories, Inc., Mountainview, CA, USA). Then, we added 600 µl DMEM with 10% FBS to the lower chambers. After a 24 h incubation period at 37°C, the cotton swabs were used to wipe out the non-migratory or non-invasive cells on the upper chambers. The cells on the lower chambers were fixed with 100% methanol for 20 min, after that, the cells were stained with 0.1% crystal violet (Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) for 30 min. The cells were counted under an inverted microscope (Olympus Corporation, Tokyo, Japan).
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