Proteomics Methodologies: The Search of Protein Biomarkers Using Microfluidic Systems Coupled to Mass Spectrometry.

Protein biomarkers have been the subject of intensive studies as a target for disease diagnostics and monitoring. Indeed, biomarkers have been extensively used for personalized medicine. In biological samples, these biomarkers are most often present in low concentrations masked by a biologically complex proteome (e.g., blood) making their detection difficult. This complexity is further increased by the needs to detect proteoforms and proteome complexity such as the dynamic range of compound concentrations. The development of techniques that simultaneously pre-concentrate and identify low-abundance biomarkers in these proteomes constitutes an avant-garde approach to the early detection of pathologies. Chromatographic-based methods are widely used for protein separation, but these methods are not adapted for biomarker discovery, as they require complex sample handling due to the low biomarker concentration. Therefore, microfluidics devices have emerged as a technology to overcome these shortcomings. In terms of detection, mass spectrometry (MS) is the standard analytical tool given its high sensitivity and specificity. However, for MS, the biomarker must be introduced as pure as possible in order to avoid chemical noise and improve sensitivity. As a result, microfluidics coupled with MS has become increasingly popular in the field of biomarker discovery. This review will show the different approaches to protein enrichment using miniaturized devices and the importance of their coupling with MS.

Directional surface enhanced Raman scattering on gold nano-gratings.

Directional plasmon excitation and surface enhanced Raman scattering (SERS) emission were demonstrated for 1D and 2D gold nanostructure arrays deposited on a flat gold layer. The extinction spectrum of both arrays exhibits intense resonance bands that are redshifted when the incident angle is increased. Systematic extinction analysis of different grating periods revealed that this band can be assigned to a propagated surface plasmon of the flat gold surface that fulfills the Bragg condition of the arrays (Bragg mode). Directional SERS measurements demonstrated that the SERS intensity can be improved by one order of magnitude when the Bragg mode positions are matched with either the excitation or the Raman wavelengths. Hybridized numerical calculations with the finite element method and Fourier modal method also proved the presence of the Bragg mode plasmon and illustrated that the enhanced electric field of the Bragg mode is particularly localized on the nanostructures regardless of their size.

Generalized analytical model based on harmonic coupling for hybrid plasmonic modes: comparison with numerical and experimental results.

Metal nanoparticle arrays have proved useful for different applications due to their ability to enhance electromagnetic fields within a few tens of nanometers. This field enhancement results from the excitation of various plasmonic modes at certain resonance frequencies. In this article, we have studied an array of metallic nanocylinders placed on a thin metallic film. A simple analytical model is proposed to explain the existence of the different types of modes that can be excited in such a structure. Owing to the cylinder array, the structure can support localized surface plasmon (LSP) modes. The LSP mode couples to the propagating surface plasmon (PSP) mode of the thin film to give rise to the hybrid lattice plasmon (HLP) mode and anti-crossing phenomenon. Due to the periodicity of the array, the Bragg modes (BM) are also excited in the structure. We have calculated analytically the resonance frequencies of the BM, LSP and the corresponding HLP, and have verified the calculations by rigorous numerical methods. Experimental results obtained in the Kretschmann configuration also validate the proposed analytical model. The dependency of the resonance frequencies of these modes on the structural parameters such as cylinder diameter, height and the periodicity of the array is shown. Such a detailed study can offer insights on the physical phenomenon that governs the excitation of various plasmonic modes in the system. It is also useful to optimize the structure as per required for the different applications, where such types of structures are used.

Soft UV nanoimprint lithography-designed highly sensitive substrates for SERS detection.

We report on the use of soft UV nanoimprint lithography (UV-NIL) for the development of reproducible, millimeter-sized, and sensitive substrates for SERS detection. The used geometry for plasmonic nanostructures is the cylinder. Gold nanocylinders (GNCs) showed to be very sensitive and specific sensing surfaces. Indeed, we demonstrated that less than 4 ×10(6) avidin molecules were detected and contributed to the surface-enhanced Raman scattering (SERS) signal. Thus, the soft UV-NIL technique allows to obtain quickly very sensitive substrates for SERS biosensing on surfaces of 1 mm (2).

Integration of short gold nanoparticles chain on SOI waveguide toward compact integrated bio-sensors.

We demonstrate the integration of short metal nanoparticle chains (L ≈700 nm) supporting localized surface plasmons in Silicon On Insulator (SOI) waveguides at telecom wavelengths. Nanoparticles are deposited on the waveguide top and excited through the evanescent field of the TE waveguide modes. Finite difference time domain calculations and waveguide transmission measurements reveal that almost all the TE mode energy can be transferred to nanoparticle chains at resonance. It is also shown that the transmission spectrum is very sensitive to the molecular environment of nanoparticles, thus opening the way towards ultra-compact sensors in guided plasmonics on SOI. An experimental demonstration is reported with octadecanthiol molecules for a detection volume as small as 0.26 attoliter.

Two-chamber AFM: probing membrane proteins separating two aqueous compartments.

Biological membranes compartmentalize and define physical borders of cells. They are crowded with membrane proteins that fulfill diverse crucial functions. About one-third of all genes in organisms code for, and the majority of drugs target, membrane proteins. To combine structure and function analysis of membrane proteins, we designed a two-chamber atomic force microscopy (AFM) setup that allows investigation of membranes spanned over nanowells, therefore separating two aqueous chambers. We imaged nonsupported surface layers (S layers) of Corynebacterium glutamicum at sufficient resolution to delineate a 15 A-wide protein pore. We probed the elastic and yield moduli of nonsupported membranes, giving access to the lateral interaction energy between proteins. We combined AFM and fluorescence microscopy to demonstrate the functionality of proteins in the setup by documenting proton pumping by Halobacterium salinarium purple membranes.