Sensitive surface enhanced Raman scattering multiplexed detection of matrix metalloproteinase 2 and 7 cancer markers

Abstract

A surface enhanced Raman spectroscopy (SERS) based platform was developed for sensitive multiplexed detection of matrix metalloproteinases (MMP) (MMP-2 and MMP-7) with low limit of detection and high specificity. Detection is based on the virtue of enzymatic reaction where a peptide can be cleaved only by its corresponding enzyme. The platform comprises two components, a specialized SERS-based bimetallic-film-over-nanosphere (BMFON) substrate and gold nanoparticles (AuNPs). The two components were functionalized such that binding between the two would occur through biotin-avidin-biotin complexation. Binding is hindered by MMP peptide chains conjugated onto the surfaces of the substrate and AuNPs, and can be removed only by cleaving the peptide chains with corresponding enzymes. Since AuNP binding sites become free after the peptides are cleaved, the number of binding sites for AuNPs onto the substrate would increase. By tagging the AuNPs, concentrations of MMP-specific enzymes can be quantified through examining intensities of signature SERS peaks of the tags. This cleave-and-bind mechanism was first validated by individual detection and quantification of MMP-2 and MMP-7. The platform was demonstrated to be able to sensitively detect concentrations of specific enzymes ranging from 1 ng/mL to 40 µg/mL, with close correlation between SERS intensity and concentrations. Finally, the multiplexed detection of MMP-2 and MMP-7 was demonstrated. The multiplexity of this platform provides a robust way to analyze diseases associated with MMP-2 and MMP-7 enzymes. Our work can be further developed as a clinical diagnostic tool to detect other MMP proteinase in bio-fluids samples, widening the number of biomarkers needed to characterize diseases better.

Superhydrophobic silver film as a SERS substrate for the detection of uric acid and creatinine

“…Since the first report on SERS in 1977, many efforts have been made to achieve higher sensitivity in chemical and biological detection [8,9]. It is generally regarded that SERS enhancement stems from two mechanism [10,11]: One is chemical enhancement, which is realized by charge transfer between the analytes and the SERS substrate; the other is electromagnetic enhancement, which plays the dominant role in SERS enhancement [12,13]. It is well known that metal nanoparticles such as silver and gold can lead to strong electromagnetic fields, which is termed "hot spot" in SERS measurements [14], thus nanostructured metal material with different morphologies (nanorod, nanosphere, nanocubic, nanowire, etc) were fabricated to create more hot spots [15].…”

Detection of Matrilysin Activity Using Polypeptide Functionalized Reduced Graphene Oxide Field-Effect Transistor Sensor

“…Sensitivities of the proposed assay for detection of MMP-7 in both buffer and human plasma are higher than that of the colorimetric approach reported previously 19 and comparable to ELISA and optical assays. [36][37][38] Additionally, LODs achieved by this assay are within clinically relevant ranges (e.g. 0.1~5 µg/mL for salivary gland cancer 19 and 1~10 ng/mL for bladder cancer patients 40 ).…”

“…19 Indeed, the sensitivity of the proposed assay is comparable to ELISA assay 36 and optical assays (e.g. surface enhanced Raman scatteringbased assay 37 and the fluorescence-based assay 38 ). Also, a LOD of 40 ng/mL was achieved upon detection of MMP-7 in human plasma.…”

Recent Progress on Liquid Biopsy Analysis using Surface-Enhanced Raman Spectroscopy

“…In this platform, avidin-modified bimetallic-film-over-nanosphere (BMFON) substrate and biotin-modified SERS tags are shielding by MMP peptides; in the presence of corresponding enzymes, the peptides are cleaved, leading to the binding of BMFON with SERS tags and thus strong SERS signals are generated ( Figure 21B ). This method enabled detection of individual enzymes ranging from 1 ng/mL to 40 µg/mL, and multiplexed detection of MMP-2 and MMP-7 was realized through simultaneous incubation with 4-aminothiophenol (4ATP)- and 2-naphthalenethiol (NT)-labeled peptide-modified SERS tags 144.…”

“…b, SEM images of (i) clean BMFON substrate (ii) peptide-shielded avidin-conjugated BMFON and (iii) SERS tags bound to BMFON after enzyme cleavage. Adapted with permission from 144, copyright 2015 Optical Society of America.…”