Cell Dissociation and Trypsin for Cell Culture

Trypsin and cell trypsinization

Trypsin, a proteolytic enzyme, is the standard way to detach adherent cell cultures and monolayers. This globular, pancreatic protease cleaves at the C-terminal side of lysine and arginine, breaking down vessel-adhering proteins and allowing easy resuspension during cell harvesting.

Thermo Fisher Scientific offers porcine-derived trypsin, EDTA-enhanced trypsin, as well as formulations of the TrypLE enzyme, a gentle, trypsin-like enzyme. To avoid unplanned protein degradation, organic trypsin must be inactivated with trypsin inhibitors. In contrast, synthetic trypsin, such as Gibco TrypLE products, are deactivated via dilution, eliminating the need for additional solutions.

Trypsin EDTA  

EDTA, a chelating agent, improves trypsin’s ability to detach adherent cells. Binding calcium and magnesium into a hexadentate, EDTA weakens cell-cell adhesion and enhances trypsin’s access to hydrolysis-targeted peptide bonds.

Gibco Trypsin-EDTA is made from irradiated porcine-derived protease with e-beam verification as well as tested for PPV, mycoplasma, and PCV 1/2 contamination.

Help enable gentle cell dissociation with TrypLE reagents

Gibco TrypLE reagents are highly purified, recombinant cell-dissociation enzymes that replace porcine trypsin. These reagents are ideal for dissociating attachment-dependent cell lines in both serum-containing and serum-free conditions, and can be directly substituted for trypsin without protocol changes. TrypLE reagents are gentle on cells, stable at room temperature, and animal origin–free.

Learn more about TrypLE reagents

Collagenase

Relatively gentle, collagenase works by digesting connective tissue’s native triple-helical collagen fibrils. These tissues are often found in skin, tendon, blood vessels, and bone—making collagenase disaggregation suitable for human tumor, mouse kidney, human brain, and epithelium cultures.

Visit the Technical Reference Library to view protocols for collagenase dissociation of cells from primary tissue

Dispase

Dispase, or neutral protease, is a Bacillus polymyxa–produced metalloenzyme. Rapid and gentle, dispase can be used to harvest and transfer normal diploid cells and cell lines. Dispase effectively separates cell clumps and splits cells from intact tissue with little impact to membrane integrity or viability.

Versene solution

Versene solution, an EDTA solution in PBS, dissociates cells gently and non-enzymatically. This chelating agent detaches some cell types (such as epithelial cells) from vessel surfaces for suspension. It can also be used as a wash prior to trypsinization.

For further guidance on Versene solution applications, please see the cell dissociation selection guide by culture type.