Ex-vivo detection of neural events using THz BioMEMS.

BACKGROUND: Electromagnetic frequencies up to a few terahertz (THz) can yield real-time and noninvasive measurements on biological matter. Unfortunately, strong absorption in aqueous solutions and low spatial resolution return difficult free-space investigations. A new approach based on integrated THz circuits was used. The authors designed and fabricated a BioMEMS (Biological MicroElectro-Mechanical System) compatible with microfluidic circulation and electromagnetic propagation. It is dedicated to the ex vivo detection of nitric oxide synthase (NOS) activity, which is involved in neurodegenerative phenomena. MATERIAL/METHODS: The biological model was a leech's central nervous system. After its injury, the production of NO was observed and measured in the far-THz spectral domain. The nerve cord was put inside a BioMEMS realized in polydimethylsiloxane (PDMS) sealed on a glass wafer. Glass is a good material for supporting high-frequency integrated waveguides such as coplanar waveguides (CPWs). Measurements were performed with vectorial network analyser (VNA). RESULTS: The transmission parameter in the frequency range of 0.14-0.22 THz was measured through CPWs located just below the microchannel containing the injured leech nerve cord. The lesion caused a decreased transmission coefficient due to the NOS activity. L-NAME was injected inside the microchannel and it was verified that it inhibits this activity. CONCLUSIONS: It was demonstrated that THz spectroscopy can detect a biochemical event, such as NOS activity around an injured leech's nerve cord, in real time. Future studies will be dedicated to quantitative measurements of the reaction products using the sophisticated management of several drugs allowed with microfluidic microsystems.

Cold plasma functionalized TeraHertz BioMEMS for enzyme reaction analysis.

In this paper, we describe the development, functionalization and functionality testing of a TeraHertz (THz) Bio-MicroElectroMechanical System (BioMEMS) dedicated to enzyme reaction analysis. The microdevice was fabricated by mixing clean room microfabrication with cold plasma deposition. The first is used to build the microfluidic circuits and the THz sensor, while the later serves for the polymerization of allylamine using a homemade glow discharge plasma reactor for a subsequent immobilization of enzymatic biocatalysts. Thermal stability of the deposited plasma polymer has been investigated by infrared spectroscopy. Fluorescent detection confirmed the efficiency of the immobilization and the enzyme hydrolysis into the BioMEMS microchannels. For the first time, the progression of the hydrolysis reaction over time was monitored by the THz sensor connected to a vectorial network analyzer. Preliminary results showed that sub-THz transmission measurements are able to discriminate different solid films, various aqueous media and exhibit specific transmission behavior for the enzyme hydrolysis reaction in the spectral range 0.06-0.11 THz.

Covalent attachment of trypsin on plasma polymerized allylamine.

This paper focuses on the immobilization of a proteolytic enzyme, trypsin, on plasma polymerized allylamine (ppAA) films. The later have been deposited onto silicon substrate by means of radiofrequency glow discharge. The covalent attachment of the enzyme was achieved in three steps: (i) activation of the polymer surface with glutaraldehyde (GA) as a linker, (ii) immobilization of trypsin and (iii) imino groups reduction treatment. The effects and efficiency of each step were investigated by X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM). Fluorescent spectroscopy was used to evaluate the change of the biological activity following the immobilization steps. The results showed that enzyme immobilization on GA-modified substrate increases the enzyme activity by 50% comparing to adsorbed enzymes, while the imino reduction treatment improves the enzyme retention by about 30% comparing to untreated samples. In agreement with XPS and AFM data, UV-vis absorption spectroscopy, used to quantify the amount of immobilized enzyme, showed that allylamine plasma polymer presents a high adsorption yield of trypsin. Although the adsorbed enzymes exhibit a lower activity than that measured for enzymes grafted through GA linkers, the highest catalytic activity obtained was for the enzymes that underwent the three steps of the immobilization process.

Nanoscale devices for online dielectric spectroscopy of biological cells.

Nanoscale probes have been developed for the online characterization of the electrical properties of biological cells by dielectric spectroscopy. Two types of sensors have been designed and fabricated. The first one is devoted to low (<10 MHz) frequency range analysis and consists of gold nanoelectrodes. The second one works for high (>40 Hz) frequency range analysis and consists of a gold nanowire. The patterning of the sensors is performed by electron beam lithography. These devices are integrated in a microfluidic channel network for the manipulation of the cells and for the improvement of the performances of the sensors. These devices are used for the analysis of a well-characterized biological model in the area of the ligand-receptor interaction. The purpose is to monitor the interaction between the lactoferrin (the ligand) and the nucleolin and sulfated proteoglycans (the receptors) present or not on a set of mutant Chinese hamster ovary cell lines and their following internalization into the cytoplasm. Initial measurements have been performed with this microsystem and they demonstrate its capability for label-free, real-time, analysis of a dynamic mechanism involving biological cells.