To evaluate cell viability, 1.5 × 105 cells per well were seeded on 6-well plates (Corning Incorporated Costar). After 48 h in culture, cells were trypsinized (Biowest), resuspended on the medium they were cultured in, and centrifuged at 300g for 5 min. The supernatant was discarded and the cell pellet was resuspended again in fresh medium. Cells were washed twice in PBS, followed by another two washes with Annexin V binding buffer (BioLegend). The resulting cell pellet was resuspended and incubated with Annexin V-FITC (BioLegend) diluted with a 1:40 ratio in Annexin V binding buffer for 15 min at RT. Cells were filtered and data were acquired using a BD FACSCanto II (Becton, Dickinson and Company) and analyzed with the FlowJo Software (v10). Two independent biological replicates were analyzed.
Trypsinized
Manufactured by Biowest
Sourced in France
Trypsinized is a laboratory reagent used to detach adherent cells from cell culture surfaces. It contains the enzyme trypsin, which cleaves the proteins that anchor cells to the substrate, allowing them to be harvested for further analysis or experimentation.
Automatically generated - may contain errors
2 protocols using trypsinized
1
Annexin V-FITC Assay for Cell Viability
2
Cell Culture and Secretome Preparation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
CL1-0 and CL1-5 cells were cultured at 37°C with 5% carbon dioxide in RPMI-1640 medium supplemented with 10% FBS. The cells were grown to 70% confluence and washed four times with serum-free medium before they were incubated in serum-free medium for 24 hours. The collected conditioned medium (CM) was concentrated by centrifugation in Amicon Ultra-15 tubes (3-kDa cutoff; Millipore, Billerica, MA). The cell extracts (CEs) were also collected in parallel as follows. The cultured cells were washed with phosphatebuffered saline and trypsinized (Biowest, Nuaillé, France). The collected cells were resuspended in distilled water and lysed by microsonication at 4°C. The suspension was centrifuged to remove any cell debris. The Bradford assay (Bio-Rad, Hercules, CA) was used to determine the total protein concentration in the prepared CM and CE samples.
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!