Description_x000D_ Application_x000D_ For trypsin digestion of peptides, use a ratio of about 1:100 to 1:20 for trypsin:peptide. The typical use for this product is in removing adherent cells from a culture surface. The concentration of trypsin necessary to dislodge cells from their substrate is dependent primarily on the cell type and the age of the culture. Trypsins have also been used for the re-suspension of cells during cell culture, in proteomics research for digestion of proteins and in various in-gel digestions. Additional applications include assessing crystallization by membrane-based techniques and in a study to determine that protein folding rates and yields can be limited by the presence of kinetic traps._x000D_ Trypsin from bovine pancreas has been used for enzymatic digestion of nucleus pulposus (NP) cells during human NP cells isolation. It has also been used as a immunopanning reagent for purification of cells._x000D_ Biochem/physiol Actions_x000D_ Trypsin cleaves peptides on the C-terminal side of lysine and arginine residues. The rate of hydrolysis of this reaction is slowed if an acidic residue is on either side of the cleavage site and hydrolysis is stopped if a proline residue is on the carboxyl side of the cleavage site. The optimal pH for trypsin activity is 7-9. Trypsin can also act to cleave ester and amide linkages of synthetic derivatives of amino acids. EDTA is added to trypsin solutions as a chelating agent that neutralizes calcium and magnesium ions that obscure the peptide bonds on which trypsin acts. Removing these ions increases the enzymatic activity._x000D_ Serine protease inhibitors, including DFP, TLCK, APMSF, AEBSEF, and aprotinin, amongst others, will inhibit Trypsin._x000D_ Components_x000D_ Trypsin consists of a single chain polypeptide of 223 amino acid residues, produced by the removal of the N-terminal hexapeptide from trypsinogen which is cleaved at the Lys - lle peptide bond. The sequence of amino acids is cross-linked by 6 disulfide bridges. This is the native form of trypsin, beta-trypsin. BETA-trypsin can be autolyzed, cleaving at the Lys - Ser residue, to produce alpha-trypsin. Trypsin is a member of the serine protease family._x000D_ Caution_x000D_ Solutions in 1 mM HCl are stable for 1 year in aliquots and stored at -20°C. The presence of Ca2+ will also diminish the self-autolysis of trypsin and maintain its stability in solution. Trypsin will also retain most of its activity in 2.0 M urea, 2.0 M guanidine HCl, or 0.1% (w/v) SDS._x000D_ Unit Definition_x000D_ One BAEE unit will produce a A253 of 0.001 per minute at pH 7.6 at 25°C using BAEE as a substrate. One BTEE unit = 320 ATEE units_x000D_ Preparation Note_x000D_ This product is from pancreas sourced from New Zealand. It is soluble in 1 mM HCl at 1 mg/mL._x000D_ For applications that involve EDTA, solubilizing trypsin should be done with a buffered salt solution containing no Ca2+ or Mg2+.