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Analyzing phosphorylated proteins

  • US 7,041,459 B2
  • Filed: 05/21/2002
  • Issued: 05/09/2006
  • Est. Priority Date: 05/21/2001
  • Status: Expired due to Fees
First Claim
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1. A method for determining the presence or absence of one or more target polypeptides in a sample suspected of containing the target polypeptides, the method comprising the steps of:

  • a)—

    mixing the sample with a class-specific reagent and one or more electrophoretic probes, wherein the class-specific reagent comprises (i) a cleavage-inducing moiety having an effective proximity and (ii) an immobilized metal affinity chromatography (IMAC) resin that binds a phosphate group on the one or more target polypeptides, and wherein the one or more electrophoretic probes each comprises a binding moiety that specifically binds one of the target polypeptide(s) and one or more electrophoretic tags each attached thereto by a cleavable linkage, under conditions that permit the class-specific reagent and the one or more electrophoretic probes to bind to the one or more target polypeptides;

    b)—

    cleaving the cleavable linkage(s) of any of the one or more electrophoretic probes that are within the effective proximity of the cleavage-inducing moiety so that one or more electrophoretic tags are released; and

    c)—

    determining the presence or absence of the one or more target polypeptides based on the presence or absence of the released electrophoretic tag(s).

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