Overview of Trypsin Medium

Trypsin Medium is a specialized solution used in cell culture and tissue dissociation. It contains the enzyme trypsin, which is a serine protease that cleaves peptide bonds on the carboxyl side of lysine and arginine residues. This medium is used primarily to detach adherent cells from the culture surface for passaging or harvesting.

Content of Trypsin Medium

• Trypsin:
  • Concentration: Typically ranges from 0.05% to 0.25%.
  • Function: Proteolytic enzyme that breaks down proteins, facilitating the dissociation of adherent cells from the culture vessel.
• EDTA (Ethylenediaminetetraacetic Acid):
  • Concentration: Commonly 0.02% to 0.05%.
  • Function: A chelating agent that binds divalent cations (e.g., calcium and magnesium). These cations are necessary for cell adhesion; their removal helps in cell detachment.
• Saline Solution:
  • Components: Often phosphate-buffered saline (PBS) or Hank's Balanced Salt Solution (HBSS).
  • Function: Provides an isotonic environment to maintain cell viability during the dissociation process.
• pH Adjusters:
  • Components: Sodium bicarbonate or HEPES buffer.
  • Function: To maintain the pH of the medium within the physiological range (typically 7.2-7.4).

Preparation and Use

• Preparation:
  • Dissolve the required amounts of trypsin and EDTA in the saline solution.
  • Adjust the pH to the desired level using a pH adjuster.
  • Sterilize the solution through filtration to ensure it is free of contaminants.
• Usage:
  • Pre-warm the Trypsin Medium to 37°C before use to enhance its enzymatic activity.
  • Remove the culture medium from the adherent cells.
  • Rinse the cells with a balanced salt solution (e.g., PBS) to remove any residual serum that can inhibit trypsin activity.
  • Add enough Trypsin Medium to cover the cell monolayer.
  • Incubate at 37°C until cells begin to detach (typically 2-5 minutes).
  • Neutralize trypsin activity by adding a medium containing serum or a trypsin inhibitor.
  • Collect the cells by gentle pipetting and proceed with passaging or analysis