The migration of the endothelial cells towards the NF membranes through 8 µm diameter pores was analysed using a transmigration assay setup. The membranes were placed on 24-well plates with a low-supplemented EGM-2 medium. FluoroBlok cell culture inserts (Corning, New York, NY, USA) were placed in each well and 20,000 HSVEC were seeded in the inserts. After 4 h in the cultures, the inserts were washed with PBS and fixed with 4% paraformaldehyde in PBS for 15 min at RT. The cells were then stained with a combination of the Texas Red C2-maleimide cell membrane dye (1.7 µg/mL in PBS, ThermoFisher Scientific, Waltham, MA, USA) and the Hoechst 33,258 nuclear dye (5 µg/mL of PBS) for 1 h at RT. Images of the cells that crossed the insert membrane were captured under an Olympus IX-71 epifluorescence microscope equipped with a DP80 digital camera (both from Olympus, Tokyo, Japan).
Texas red c2 maleimide cell membrane dye
Manufactured by Thermo Fisher Scientific
Sourced in United States
Texas Red C2-maleimide is a fluorescent dye used to label cell membranes. It is a maleimide-reactive dye that covalently binds to thiol groups on the cell surface, allowing for the visualization of the cell membrane.
Automatically generated - may contain errors
Lab products found in correlation
Ix71 epifluorescence microscope, by Olympus (2 mentions)
Fluoroblok cell culture insert, by Corning (2 mentions)
Dp80 digital camera, by Olympus (2 mentions)
Hoechst 33258 nuclear dye, by Thermo Fisher Scientific (2 mentions)
Tcs spe dm2500, by Leica (1 mentions)
Hoechst 33258 cell nucleus dye, by Merck Group (1 mentions)
3 protocols using texas red c2 maleimide cell membrane dye
1
Endothelial Cell Migration Assay
2
Visualizing Cell Spreading on Nanofibrous Membranes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The spreading and morphology of the cells on uncoated or protein-coated nanofibrous membranes were visualized on days 1, 3, and 7 after seeding by staining the cells with a combination of fluorescent dyes diluted in PBS (5 μg/mL Hoechst 33258 cell nucleus dye; Sigma-Aldrich; and 20 ng/mL Texas red C2-maleimide cell membrane dye; Thermo Fisher Scientific) for 1 hour at room temperature in the dark. Instead of Texas red staining, the F-actin cytoskeleton of the cells was stained with phalloidin conjugated with tetramethylrhodamine isothiocyanate fluorescent dye (Sigma-Aldrich), diluted in PBS to a final concentration of 5 μg/mL, for 1 hour at room temperature in the dark. Before staining, the cells were rinsed with PBS and were fixed with −20°C cold ethanol for 10 minutes. Images of the cells were taken using epifluorescence microscopy (magnification 10×, IX 51; Olympus, Tokyo, Japan) equipped with a digital camera (DP 70), or using the Leica TCS SPE DM2500 upright confocal micro-scope, magnification 40×/1.15 NA oil. On day 1 after seeding of the cells, the spreading area of islands formed by human HaCaT keratinocytes was measured on images taken under fluorescence microscopy using the Atlas software.
3
Cell Migration Assay using FluoroBlok Inserts
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The migration of HSVECs through 8 µm-diameter pores was analyzed using FluoroBlok cell culture inserts (Corning, New York, NY, USA). The membranes (PCL-PVA + PL and PCL-PVA) were placed in a 24-well culture plate with a low-supplemented EGM-2 medium. Then, cell culture inserts were placed in each well and 20 000 HSVECs were seeded in the inserts. After 4 h, the inserts were washed with PBS and were fixed with 4% paraformaldehyde in PBS for 15 min at RT. The cells were then stained with a solution of Texas Red C 2 -maleimide cell membrane dye (1.7 µg mL -1 in PBS, ThermoFisher Scientific, Waltham, MA, USA) and Hoechst 33258 nuclear dye (5 µg mL -1 of PBS) for 45 min at RT. Images of the cells that migrated through the insert pores were captured under an Olympus IX-71 epifluorescence microscope equipped with a DP80 digital camera (both from Olympus, Tokyo, Japan).
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!