The majority of protein samples submitted for post-translational modification (PTM) analysis are submitted as Coomassie Blue stained gel bands or spots that are then subjected by the PCTC Laboratory to in gel trypsin digestion. The service charge for in gel digestion includes carrying out a control digest on an otherwise identical section of gel (supplied by the investigator) that does not contain protein. The purpose of this control is to enable rapid identification of reagent and trypsin autolysis products, if desired. Upon request, a variety of other enzymes (e.g., endopeptidase Lys-C, Asp-N and Glu-C, as well as chymotrypsin, Proteinase K, etc.) may be used in place of trypsin.Samples that do not require SDS PAGE purification may be submitted dry (after vacuum centrifugation in a Speedvac) in 1.5 ml Eppendorf tubes that do not contain O-rings. Although these samples generally may contain up to the equivalent of 50 µl of 0.5 M non-volatile salts, the investigator should email the details of these samples (i.e., exact buffer composition and estimated protein concentration) to the PCTC prior to final sample preparation. Unless requested otherwise, we will generally reduce, carboxymethylate and then digest these samples with trypsin in a final concentration of 2 M urea.
