Ultra-high specificity fluorescent labeling

A method for detecting a target material in a sample, comprising the steps of:

• (a) tagging said target material with at least a first peptide tag and a second peptide tag at a distance of about 0.1 R0 to about 2 times R0 from said first peptide tag to form a tagged target material;
• (b) exposing said sample to a first fluorescent conjugate including a first fluorochrome, said first conjugate being capable of selectively associating with said first peptide tag;
• (c) exposing said sample to a second fluorescent conjugate including a second fluorochrome having a distinct excitation and emission maxima from said first fluorochrome, said second conjugate being capable of selectively associating with said second peptide tag;
• (d) allowing said first and second fluorescent conjugates to associate with said tagged target material, wherein said first and second fluorescent conjugates independently assemble on said target material by associating with said first and second peptide tags, respectively, so as to form a self assembled relay probe (SARP), such that upon excitation of said first fluorescent conjugate, fluorescence energy transfer results in excitation of said second fluorescent conjugate;
• (e) exposing said sample containing said SARP to light of a suitable wavelength for said excitation of said first fluorescent conjugate to occur;
• and
• (f) detecting said energy transfer.

A method of monitoring a reaction between analytes, comprising the steps of:

• (a) providing a first analyte in a sample, said first analyte having bound thereto at least a first and second peptide tag, wherein a distance between said tags is from about 0.1 to about 2 times Ro;
• (b) exposing said sample to a first fluorescent conjugate which selectively associates with said first tag;
• (c) allowing said first analyte to react with a second analyte labeled with a second fluorescent conjugate which selectively associates with said second tag, wherein the second conjugate is capable of participating in fluorescence energy transer with said first fluorescent conjugate;
• (d) exposing said sample to light of a suitable wavelength to allow said energy transfer to occur between said fluorescent conjugates;
• (e) monitoring the reaction between said analytes by monitoring a detectable signal generated as a result of said energy transfer between said fluorescent conjugates.

A composition comprising a first peptide tag/spacer/second peptide tag module, wherein:

• (i) said first peptide tag is of the form C(Xi)C, wherein X is any amino acid, C is Cysteine and i is 0-6;
• (ii) said second peptide tag is of the form:
• C(X)iC(X)jC(X)kC, wherein C is Cysteine, X is an amino acid, and i, j, and k are each independently 0 or an integer of from 1 to 8;
• and (iii) said spacer, which separates said first and second peptide tags is of the form (X)n, where X is any amino acid and n is an integer from about 6 to about 26.

A recombinant nucleic acid sequence encoding a first peptide tag/spacer/second peptide tag module, wherein:

• (i) said first peptide tag is of the form C(Xi)C, wherein X is any amino acid, C is Cysteine and i is 0-6;
• (ii) said second peptide tag is of the form:
• C(X)iC(X)jC(X)kC, wherein C is Cysteine, X is an amino acid, and i, j, and k are each independently 0 or an integer of from 1 to 8;
• and (iii) said spacer, which separates said first and second peptide tags, is of the form (X)n, where X is any amino acid and n is an integer from about 6 to about 26.

A composition comprising an isolated or recombinant nucleic acid sequence encoding:

• (a) at least one target material;
• and
• (b) a first peptide tag/spacer/second peptide tag module, wherein (i) said first peptide tag is of the form C(Xi)C, wherein X is any amino acid, C is Cysteine and i is 0-6;
• (ii) said second peptide tag is of the form C(X)iC(X)jC(X)kC, wherein C is Cysteine, X is an amino acid, and i, j, and k are each independently 0 or an integer of from 1 to 8;
• and (iii) said spacer, which separates said first and second peptide tags, is of the form (X)n, where X is any amino acid and n is an integer from about 6 to about 26.