Characterization of protein mixtures using Surface-Enhanced Raman Scattering (SERS)

The analysis of complex protein mixtures has high significance for clinical diagnostics, particularly in the field of early diagnosis of diseases and for the identification of biomarkers. Currently gel electrophoresis in combination with mass spectrometry is used for this purpose. Unfortunately this procedure is very time consuming, needs extensive sample preparation and does not yield any structural information of the bio-molecules investigated.

Therefore, the aim of this project is the development of an alternative procedure based on the combination of the separation technique “Free-Flow-Electrophoresis (FFE)” with sensitive “Surface-Enhanced Raman Scattering (SERS)” - detection. The technique is characterized by its extremely small interference to water, negligible sample preparation and its high information content on the molecular structure of the examined samples.

The realization of such a highly sensitive Raman scattering detection system is challenging. In order to eliminate interfering fluorescence emissions, near-infrared excitation at 785nm is used. For increasing the usually very bad Raman detection limit, which in general is not sufficient for the identification and quantification of the usually low concentrated protein species, two different techniques are applied:

On one hand low-loss liquid-core optical waveguides are used as detection-cells in order to substantially increase the interaction path between the laserbeam and the sample, and therefore increase the Raman signal.

: Figure 1: Electron micrograph of a SERS-substrate

On the other hand SERS promises huge increase of the Raman scattering intensity up to 10¹⁴ orders of magnitude. This strong enhancement is generated by plasmonic resonances on metallic nano-structured surfaces which induces huge local electric field enhancements. In figure 1 a micrograph of such a nano-structured SERS-surface is shown.

Figure 2 shows the efficiency of SERS enhancement in comparison to a usual Raman scattering. In figure 2a a Raman spectrum of pure crystalline bovine insulin is displayed. In comparison the Raman spectrum of a 300 mmol/l aqueous insulin solution on a smooth gold surface is depicted in figure 2b. Essentially only the Raman spectrum of the substrate can be detected. In contrast figure 2c shows the SERS spectrum of the 300 mmol/l insulin solution on a SERS-substrate. The extreme signal enhancement gained by means of SERS is easily recognizable. This diagram clarifies the enormous potential of this technique for the detection and identification of smallest protein quantities.

: Figure 2: a) Pure crystalline Bovine insulin, b) 0,5µl of 300 µmol/l insulin-solution on a smooth gold surface (dried), c) 0,5µl of 300 µmol/l insulin-solution on a SERS-surface (dried)

Sponsorship:

Landesinitiative Biophotonik des Niedersächsischen Ministeriums für Wirtschaft, Arbeit und Verkehr

Co-operation partners:

Laser-Laboratorium Göttingen GmbH, Prof. Dr. G. Marowsky
Microliquids GmbH, Prof. Dr. B. Abel
Universität Göttingen, Prof. Dr. W. Lauterborn