The invention provides a simple and quick protocol for chemical treatment,
enzymatic or chemical digestion, and subsequent identification of
proteins or polypeptides on a protein chip such as protein chip arrays.
The chemical treatment includes denaturation, reduction and alkylation
while the enzymatic digestion includes deglycosylation,
dephosphorylation, and digestion by various proteases. The proteins or
polypeptides can also be digested by using various chemicals that are
known to induce proteolysis. All reactions are carried out sequentially
on chip. Subsequent peptide mass fingerprinting or product ion searches
allow the identification of specific peptides, which can be correlated to
proteins. The methods of the present invention can be used to analyze
biological samples such as urine and plasma and to identify biomarkers in
diseased states. The methods of the present invention allow complete on
chip treatment, which can be used for rapid protein identification and
structural characterization of heavily posttranslationally modified
proteins.