Sterile: Ultrapure, Filtered 0.1-micron membrane TWICE and 0.04-micron membrane ONCE.
World’s cleanest buffer for all cell and molecular biology experiments
Description:
FastDetach Trypsin Medium (1X) is a ready-to-use medium which provides a balance between rapid cell detachment and maintaining cell viability, making it suitable for routine cell culture applications where speed is prioritized.
The high concentration of trypsin (0.5%) combined with EDTA is particularly effective for detaching strongly adherent cells, transformed fibroblasts, multilayer cultures, and certain tumor cell lines. Transformed fibroblasts and some strongly adherent cell lines produce more extracellular matrix proteins, creating a dense network that anchors them more firmly to the substrate. Additionally, the well-developed cytoskeleton with numerous focal adhesions makes them more resistant to gentle dissociation methods. These cell types also often have increased expression of adhesion molecules and form strong attachments to the culture surface compared to other cell types. Fast and Quick detachment process is highly desireable for such type of cell culture settings.
The trypsin-EDTA in DPBS combination is effective for dissociating dense, multilayered cultures, which are common with strongly adherent cell lines and transformed fibroblasts. EDTA in DPBS acts as a chelating agent that weakens cell-matrix interactions by binding calcium ions, which are crucial for cell adhesion. This enhances the proteolytic action of trypsin, making it more effective for strongly adherent cells. DPBS provides a balanced salt solution that helps maintain cell viability during the detachment process. DPBS provides an isotonic environment, preventing osmotic stress on the cells during detachment. Moreover, DPBS is compatible with both trypsin and EDTA, allowing for effective cell detachment without interfering with the action of these components. Trypsin and EDTA can quickly dissociate strongly adherent cells, often within 5 minutes at 37°C, compared to other methods like collagenase which may require up to 60 minutes. It is recommended to observe cells under a microscope and stop trypsinization once cells round up and detach.
* This medium provides efficient and fast cell detachment but should be used judiciously, considering the specific needs of the cell type and experimental requirements. It's particularly useful when rapid detachment is necessary, but researchers should be aware of its potential effects on cell surface proteins and subsequent cellular behavior. This medium may not be optimal for primary cells where cell surface protein integrity is critical, easily detachable cells like some epithelial cells, stem cells and sensitive cell types. FBS containing media can be use to immediately neutralize and stop the trypsinization process.
* If an addition of D-glucose (or Dextrose) is required, please ask us. We can customize the trypsinization medium.
Usage considerations:
Should be used with caution for sensitive cell types or applications where preserving cell surface proteins is critical.
Requires careful timing to minimize exposure and potential cell damage.
Neutralization with a serum-containing medium or trypsin inhibitor is important after detachment.
Cells may require longer recovery times (8-24 hours) for surface proteins to regenerate after exposure to this higher trypsin concentration.
Pre-warm trypsin and neutralizing medium to 37°C to optimize enzyme activity and minimize exposure time.
Advantages:
Faster detachment compared to lower trypsin concentrations
Convenient ready-to-use formulation
Maintains high cell viability for most cell lines
Effective for dissociating dense, multilayered cultures
Applications:
Routine passaging of robust cell lines
Situations requiring quick cell harvesting
High-throughput cell culture applications
Strongly adherent cell lines in early passages
Transformed fibroblasts
Highly confluent, multilayer cultures
Some tumor cell lines
Protocol:
Remove the spent medium from the culture vessel by aspiration.
Wash the monolayer by adding a balanced salt solution without calcium and magnesium to the side of the flask opposite the cells.
Rinse the cell sheet by rocking the flask for 1 to 2 minutes and discard the wash solution.
Add FastDetach Trypsin Medium (1X) to the side of the flask opposite the cells. The volume should be sufficient enough to completely cover the monolayer of the cells.
Rock the flask to ensure that the dissociation solution covers the cell sheet.
Incubate the flask at 37°C for 2 to 3 minutes. Monitor the process by observing the flask under an inverted microscope and one can gently tap the plate/flask on the benchtop to help detachment. When dissociation is complete, the cells will be in suspension and appear rounded. In addition to rocking gently, flasks of cell lines that are characteristically difficult to remove from substratum may be tapped to expedite removal. Note: The exact time needed to dissociate cells will round-up in shape and vary in shapes according to the cell line. The dissociation process should be monitored closely to avoid cell damage.
Once the cell dissociation is complete add serum (i.e., FBS or FCS) containing complete medium to the flask to inhibit the tryptic activity, thus it stops further damage the cells.
Disperse the cells into a single cell suspension by pipetting gently and repeatedly.
Count and subculture the cells.
Quality Control:
Appearance: Red-Pink Liquid.
pH: 7.4
Sterility: Filtered 0.1 micron Twice and 0.04 micron once in a sterile environment.
DNase activity: None was detected after incubation of plasmid DNA, and this product was used for 18 hours at room temperature.
RNase activity: No RNase activity was detected after incubating ribosomal RNA with this product for 18 hr at room temperature.
Storage and Shelf Life:
Storage: -20℃ for long term
Shelf life: 2 years
Shipping condition: Dry Ice
CUSTOMIZATION: Please INQUIRE if other concentrations, additions of chemicals/compounds/proteins/supplements, different pH, and modifications are needed.
All DiagnoCine Precision Sterile buffers are filtered-sterilized with a 0.1-micron filtration two times and 0.04-micron filtration one time. Thus, mycoplasma contamination is prevented. The smallest size mycoplasma type can be about 0.2 microns.
* This product is manufactured under ISO 13485-certified and CE-approved facilities (Suppliers of DiagnoCine Precision). All final packaging, quality assurance, and testing are done at the DiagnoCine R&D and Quality Testing Center. All specific customization requests and assembly were accomplished at DiagnoCine Precision in Totowa, New Jersey, USA
