When sufficient separation is achieved, we collect the peaks and perform mass spectrometry prior to sequence analysis. This procedure ,though somewhat less efficient than digestion in free solution, has proven to be a successful approach. Adsorption A major factor in the isolation of small quantities of protein is sample loss due to surface adsorption to plastic and glass. For example, direct the effluent from ion exchange directly to reverse phase columns avoiding dialysis losses. Avoid polystyrene, and use polyethylene or polypropylene micro centrifuge tubes keeping plastic exposure as low as possible by minimizing total volume.
Losses become critical at sub microgram levels.
Avoid drying or lyophilizing samples if possible. Use reagent grade water. Otherwise, use fresh, analytical grade formic acid. If you use urea, deionize it with mixed bed Amberlite immediately prior to use. The key word here being enrichment .