Trypsin is compatible with many buffer types, with the optimal pH between 8-9. Typical reaction buffers are 50 mM Tris-HCl, pH 8.0, or 50 mM ammonium bicarbonate, pH 7.8. Calcium ions, although not necessary for its activity, can stabilize trypsin, especially in extended digestion conditions. A concentration of 1 - 10 mM is often used.
The selection of the digestion buffer also depends on how the protein sample is collected or prepared, and if the digestion is immediately followed by mass spectrometry analysis. Generally, ammonium bicarbonate buffer is considered the most mass spectrometry-friendly buffer system, as both the anions and cations are volatile.
