ABSTRACT
Figure 4.1 (a) Procedure for SERS protein detection based on a Western blot; (b) SERRS spectra of Mb with different amounts on a nitrocellulose membrane — a: solid Mb, b: 4 ng, c: 6 ng, d: 8 ng, and e: 10 ng. The inset shows an intensity of a SERRS peak at 1370 cm-1 versus the amount of Mb on a nitrocellulose membrane. (Reprinted with permission from ref. 33. Copyright 2008 American Chemical Society.)
In another approach, the proteins were detected after a Western blot [33]. Western blotting is an analytical method used to detect proteins in a mixture. First, the protein mixture is separated with gel electrophoresis. The separated proteins are then transferred onto a membrane and targeted with an antibody or recently have been analyzed using mass spectroscopy [34-36]. In that proposed technique, the bands on a slab gel were transferred onto a nitrocellulose membrane and then stained with a silver colloidal suspension. The SERS experiment was performed with a Raman instrument equipped with a 514.5 nm laser, where its use contributes to further enhancement of Raman scattering (surfaceenhanced resonance Raman scattering [SERRS]). Figure 4.1 shows the procedure for SERS protein detection based on a Western blot. Myoglobin (Mb) and bovine serum albumin
(BSA) were used as model proteins, and the SERRS detection limit of Mb is found to be 4 ng/band. In a more recent study, Culha and coworkers demonstrated a novel approach for label-free protein detection [37]. In that study, proteins and colloidal AgNPs were convectively assembled on glass surfaces and the SERS spectra of proteins regardless of the protein size and surface charge properties were successfully acquired. The convective assembly process, which is assembly of particles from a moving and drying droplet, allows the proteins and AgNPs to assemble into ordered structures in a reproducible manner. Figure 4.2 shows the assembly process using “convective assembly” and the thin film formed on glass surface (a) and SERS spectra of avidin with decreasing concentration (b). The detection limit was improved more than one order of magnitude (0.5 µg/mL) compared to previous reports.
