The trypsin:protein ratio is often between 1:20 and 1:100, with 1:50 commonly recommended, resulting in efficient digestion without excessive trypsin autolysis. This ratio is often dependent on factors such as the nature and source of the protein sample, the use of an optimal buffer, and the incubation temperature.
When using Trypsin-ultra™, Mass Spectrometry Grade (Recombinant) (NEB #P8119) for fast or high-throughput workflows, we have used ratios of 1:5 or 1:10 (without observable artifacts), enabling the digest to be performed in 30 minutes at 37 °C.
