Patent ID: 8877516

Claim:
A method of sequentially detecting three epitopes of a biomarker in a biological sample, said method comprising: (a) contacting said biological sample with a device having an immobilized binding agent on the surface thereof in a pattern that generates a signal so that said biomarker in said sample specifically binds to said immobilized binding agent, (b) contacting said surface with a first detecting binding agent, wherein said first detecting binding agent specifically binds to a first epitope of the biomarker and not to a second or third epitope of the biomarker; (c) detecting said first epitope by a change in signal produced by said pattern upon binding of said first detecting binding agent to said first epitope; (d) contacting said surface with a second detecting binding agent, wherein said second detecting binding agent specifically binds to the second epitope of said biomarker and does not specifically bind to said first or third epitope; (e) detecting said second epitope by a change in signal produced by said pattern upon binding of said second detecting binding agent to said second epitope; (f) contacting said surface with a third detecting binding agent, wherein said third detecting binding agent specifically binds to the third epitope of said biomarker and does not specifically bind to said first or second epitope; and (g) detecting said third epitope by a change in signal produced by said pattern upon binding of said third detecting binding agent to said third epitope.