In accordance with the manufacturer's instructions, we vaccinated the YY-8103 and LM3 cells line at the upper chamber, and the culture was performed on a 200 μL serum-free 1640 medium. The matrigel mix (BD Biosciences,United States) covers the transwell chamber (Corning, United States) so that the invasion test can be realized and the matrigel mix is not needed for the migration experiment. The HCC cell chemical inducers made by RPMI 1640 medium and 10% FBS were lured to the bottom of the chamber. Incubation for 24 h, we fixed the color of the upper chamber. Then crystal violet (Kaigen, China) was used for dyeing for 15 min. We photographed and counted the cells in three fields in order to implement visualization.
Matrigel mix
Manufactured by Corning
Sourced in United States
Matrigel mix is a complex extracellular matrix (ECM) preparation derived from the Engelbreth-Holm-Swarm (EHS) mouse sarcoma. It is used as a component in cell culture systems to provide a more physiologically relevant microenvironment for various cell types.
Automatically generated - may contain errors
8 protocols using matrigel mix
1
Cell Invasion and Migration Assay
2
Transwell Assay for Cell Migration and Invasion
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The transwell chambers (Corning, USA) paved with and without a matrigel mix (Corning, USA) were used to assess the migration and invasion ability. GBM-Z1 cells were inoculated to the upper chamber and conditioned macrophages were inoculated to the bottom chamber. After incubation for 48 h, the cells on the upper surface of the membrane were removed. Cells traversing the membrane were fixed and then stained by crystal violet, and images were taken by an inverted microscope.
3
Transwell Invasion and Migration Assay
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The transwell chamber covered with or without Matrigel Mix (Corning, NY, USA) were used to detect the invasion and migration ability of GC cells respectively. AGS (3×104) and HGC-27 cells (2×104) suspending in 200 μl serum-free medium were cultured in upper chambers, and 600 μl 10% FBS-medium was filled in the lower chamber. After 2 days incubation, 4% formaldehyde and 0.1% crystal violet were used to fix and stain the cells covered in the bottom chamber respectively. After erasing the cells in the upper chamber, the remaining cells adhered to the bottom chambers were photographed at 100× and counted in 10 different random fields of view.
4
Transwell Assay for Cell Migration and Invasion
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Cell migration and invasion abilities were measured by 24-well transwell chambers (Millipore, MA, USA) with or without Matrigel Mix (Corning, NY, USA). 1 × 104 transfected GC cells for migration and 2 × 105 transfected cells for invasion were resuspended in medium. The upper chamber contained 200μL serum free medium, and 600μL of complete medium was added to the bottom chamber. After incubation at 37℃ for 24–48 h, 4 % formaldehyde and 0.1 % crystal violet were used to fix and stain the cells covered in the bottom chamber. Migrative or invasive cells were photographed and counted under a light microscope.
5
Wound Healing and Cell Invasion Assays
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For the wound healing assay, transfected cells were cultured in six-well plates until confluent. After scratching the monolayer, the cells were photographed at 0, and 48 h. Images were taken from five random optical fields on each filter. For the Transwell assay, a Transwell chamber (Corning, USA) which was coated with or without Matrigel mix (Corning, USA) was used to assess cell invasion and migration, respectively. After being transfected, 2 × 104 cells were plated in the top chamber with serum-free medium, and a medium containing 10% fetal bovine serum was used in the lower chamber as a chemoattractant. After incubation for 24 h, the cells located on the bottom of the chamber were fixed with 4% paraformaldehyde for 15 min, stained with crystal violet (0.1%) for 15 min, and photographed under a microscope. The migrated or invaded cells were counted in five randomly selected fields in each well. Each sample was assayed in triplicate.
6
Transwell Invasion Assay
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Cells were digested into a single cell suspension. 1×104 cells were seeded in the upper chambers with 200 μL serum-free medium. Transwell chamber (Corning, USA) was paved with matrigel mix (Corning, USA) for invasion assay. The bottom chamber was filled with a medium containing 10% FBS. After incubation at 37℃ for 24 hours, the upper chamber was washed with cold phosphate buffer saline (PBS) and fixed with cold methanol at 4℃, then stained with crystal violet (Kaigen, China) for 20 min. Finally, the cells in different fields were photographed and counted.
7
Cell Invasion and Migration Assay
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In accordance with the manufacturer instructions, we vaccinated the YY-8103 and LM3 cells line at the upper chamber, and the culture was performed on 200 microliter serum-free 1640 medium. The matrigel mix (BD Biosciences, San Jose, CA, USA) covers transwell chamber (Corning, NY, USA) so that the invasion test can be realized and the matrigel mix is not needed for migration experiment. The HCC cell chemical inducers made by RPMI 1640 medium and 10% FBS was lured to the bottom of the chamber.
Incubation for 24 hours, we xed the colour of upper chamber. Then crystal violet (Kaigen, Nanjing, China) was uesd to dyeing for 15 minutes. We photographed and counted the cells in ve elds in order to implement visualization.
Incubation for 24 hours, we xed the colour of upper chamber. Then crystal violet (Kaigen, Nanjing, China) was uesd to dyeing for 15 minutes. We photographed and counted the cells in ve elds in order to implement visualization.
8
Transwell Invasion Assay Protocol
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The invasive capacity of the cells was determined using Transwell chambers (Corning-Costar, Corning, NY, USA; cat no. 3422). Cells were incubated with PYM (100 µg/ml), LY294001221 (0.5 nM), PYM (100 µg/ml) plus IGF-1 (100 ng/ml) or an equal volume of DMSO for the negative control in serum-free DMEM for 12 h. Cells were then harvested and added to the upper chamber of a Transwell insert (1x10 4 cells in 100 µl), which was coated with a Matrigel ® mix (Corning Incorporated, Corning, NY, USA). The lower chamber was filled with complete medium. After 48 h of incubation at 37˚C, the cells on the upper surface of the membrane were removed. The cells attached to the lower surface of the membrane were fixed with 4% paraformaldehyde (Beyotime Institute of Biotechnology, Shanghai, China) for 20 min, stained with hematoxylin (Beyotime Institute of Biotechnology) for 5-10 min and counted under a microscope (Olympus CX41; Olympus Corp., Tokyo, Japan).
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