Detection of protease and protease activity using a single nanocrescent SERS probe
This invention pertains to the in vitro detection of proteases using a single peptide-conjugate nanocrescent surface enhanced Raman scattering (SERS) probes with at least nanomolar sensitivity. The probe enables detection of proteolytic activity in extremely small volume and at low concentration. In certain embodiments the probes comprise an indicator for the detection of an active protease, where the indicator comprises a nanocrescent attached to a peptide, where said peptide comprises a recognition site for the protease and a Raman tag attached to the peptide.
1. A method of detecting or quantifying the presence, amount, or activity of at least one protease in a sample, said method comprising:
contacting said sample with an indicator comprising a nanocrescent attached to a peptide, wherein said peptide comprises a recognition site for said protease; and
monitoring differences in spectral characteristics of detected surface-Raman scattering spectra, the differences being changes in a Raman spectrum over time, where the differences are indicators of the presence, amount or activity of protease present in the sample.
2. The method of claim 1 , wherein said indicator further comprises a Raman label attached to said peptide.
3. The method of claim 1 , wherein said monitoring comprises monitoring surface enhanced Ramen scattering (SERS).
4. The method of claim 1 , wherein said sample comprises material selected from the group consisting of sample selected from the group consisting of whole blood, plasma, serum, synovial fluid, cerebrospinal fluid, bronchial lavage, ascites fluid, seminal fluid, bone marrow aspirate, pleural effusion, urine, and tumor cells or tissue.
5. The method of claim 1 , wherein said peptide comprises a recognition site for a protease that is a marker for a cancer presence or progression.
6. The method of claim 5 , wherein said peptide comprises a recognition site for PSA.
7. The method of claim 5 , wherein said peptide comprises recognition site comprising the amino acid sequence HSSKLQ (SEQ ID NO:1).
8. The method of claim 1 , wherein said sample comprises a material selected from the group consisting of whole blood, a blood fraction, lymph, cerebrospinal fluid, oral fluid, mucus, urine, feces, and seminal fluid.
9. A method of detecting the presence or quantity of an analyte, said method comprising
contacting a sample comprising said analyte to an indicator, said indicator comprising a nanocrescent attached to a substrate that is specifically or preferentially bound by said analyte in the presence of a Raman-labeled moiety that competes with said analyte for binding to said substrate; and
detecting the Raman spectrum of said indicator where a change over time in the amplitude and/or wavelength of one or more peaks comprising the Raman spectrum produced by dissociation of said Raman-labeled moiety from said substrate provides a measure of the presence or quantity of said analyte in said sample.
10. The method of claim 9 , wherein said substrate is a peptide or a nucleic acid.