Thomas J. Roussel

Cardiac Cell Culture Model As a Left Ventricle Mimic for Cardiac Tissue Generation

A major challenge in cardiac tissue engineering is the delivery of hemodynamic mechanical cues that play a critical role in the early development and maturation of cardiomyocytes. Generation of functional cardiac tissue capable of replacing or augmenting cardiac function therefore requires physiologically relevant environments that can deliver complex mechanical cues for cardiomyocyte functional maturation. The goal of this work is the development and validation of a cardiac cell culture model (CCCM) microenvironment that accurately mimics pressure-volume changes seen in the left ventricle and to use this system to achieve cardiac cell maturation under conditions where mechanical loads such as pressure and stretch are gradually increased from the unloaded state to conditions seen in vivo. The CCCM platform, consisting of a cell culture chamber integrated within a flow loop was created to accomplish culture of 10 day chick embryonic ventricular cardiomyocytes subject to 4 days of stimulation (10 mmHg, ∼13% stretch at a frequency of 2 Hz). Results clearly show that CCCM conditioned cardiomyocytes accelerate cardiomyocyte structural and functional maturation in comparison to static unloaded controls as evidenced by increased proliferation, alignment of actin cytoskeleton, bundle-like sarcomeric α-actinin expression, higher pacing beat rate at lower threshold voltages, and increased shortening. These results confirm the CCCM microenvironment can accelerate immature cardiac cell structural and functional maturation for potential cardiac regenerative applications.

Electrochemical and microfabrication strategies for remotely operated smart chemical sensors: Application of anodic stripping coulometry to calibration-free measurements of copper and mercury

Remote unattended sensor networks are increasingly sought after to monitor the drinking water distribution grid, industrial wastewater effluents, and even rivers and lakes. One of the biggest challenges for application of such sensors is the issue of in-field device calibration. With this challenge in mind, we report here the use of anodic stripping coulometry (ASC) as the basis of a calibration-free micro-fabricated electrochemical sensor (CF-MES) for heavy metal determinations. The sensor platform consisted of a photo-lithographically patterned gold working electrode on SiO2 substrate, which was housed within a custom stopped-flow thin-layer cell, with a total volume of 2-4 μL. The behavior of this platform was characterized by fluorescent particle microscopy and electrochemical studies utilizing Fe(CN)6(3-/4-) as a model analyte. The average charge obtained for oxidation of 500 μM ferrocyanide after 60s over a 10 month period was 176 μC, corresponding to a volume of 3.65 μL (RSD = 2.4%). The response of the platform to copper concentrations ranging from 50 to 7500 ppb was evaluated, and the ASC results showed a linear dependence of charge on copper concentrations with excellent reproducibility (RSD ≤ 2.5%) and accuracy for most concentrations (≤ 5-10% error). The platform was also used to determine copper and mercury mixtures, where the total metallic content was measurable with excellent reproducibility (RSD ≤ 4%) and accuracy (≤ 6% error).

Single element 3-terminal pressure sensors: A new approach to pressure sensing and its comparison to the half bridge sensors

We report the development of a novel 3-terminal single element piezoresistor for ultra-miniature pressure sensor applications and compare its performance to that of a traditional half Wheatstone bridge design. The pressure sensors reported here are 0.69-French in size (1F= 333µm) and are designed and batch-fabricated using SOI (silicon on insulator) and DRIE (deep reactive ion etching) technologies. One of the major applications of this device is for blood pressure monitoring using ultra-miniature 1F catheters. The combination of SOI and DRIE technologies results in uniform diaphragm thickness and complete elimination of the post-processing dicing step by micromachining “die separation streets” during the DRIE process. The novel 3-terminal single element design and half Wheatstone bridge sensors were optimized using finite element analysis (FEA). Performance characteristics of the half bridge and 3-terminal sensors, i.e. sensitivity, nonlinearity (NL%), temperature coefficient offset (TCO) and drift were measured and compared. It was determined that the 3-terminal pressure sensors (3-TPS) had greater sensitivity, better non-linearity and lower drift compared to half bridge design sensors. The 3-TPS devices were also less sensitive to alignment errors.

Alternative fabrication methods for capillary electrophoretic device manufacturing

This work represents research that explores the development of novel manufacturing methods to create microcapillary electrophoretic (CE) devices. Nontraditional substrates that were investigated include polymers such as SU-8, poly dimethylsiloxane (PDMS), acetate, Riston, Kapton, polyimide, and polyester. Hot embossing, chemical etching, micro-molding, wafer level bonding, chemical treatment, and lamination techniques were developed for these substrates. The purpose of this paper is to explore the feasibility of micromachining a select group of alternative materials.

Microfabrication Process and Characteristic Testing of a MEMS-Based Preconcentrator

Due to terroristic activities it has become of significant importance to accurately detect vapor or trace particles of explosive materials. This task is difficult due to the limited number of particles available to be gathered for analysis. In order to combat this problem a preconcentrator has been developed to increase collection efficiency and the signal to noise ratio. Described is the fabrication process and characteristic testing prior to use with a portable ion mobility spectrometer (IMS).

Fabrication of a Glass Capillary Electrophoresis Microchip With Integrated Electrodes

In this chapter, a detailed outline delineating the processing steps for microfabricating capillary electrophoresis (CE) with integrated electrochemical detection (ECD) platforms for performing analyte separation and detection is presented to enable persons familiar with microfabrication to enter a cleanroom and fabricate a fully functional Lab-on-a-Chip (LOC) microdevice. The processing steps outlined are appropriate for the production of LOC prototypes using easily obtained glass substrates and common microfabrication techniques. Microfabrication provides a major advantage over existing macro-scale systems by enabling precise control over electrode placement, and integration of all required CE and ECD electrodes directly onto a single substrate with a small footprint. In the processing sequences presented, top and bottom glass substrates are photolithographically patterned and etched using wet chemical processing techniques. The bottom substrate contains seven electrodes required for CE/ECD operation, whereas the top substrate contains the microchannel network. The flush planar electrodes are created using sputter deposition and lift-off processing techniques. Finally, the two glass substrates are thermally bonded to create the final LOC device.

Dual capillary electrophoresis devices with electrochemical detection on a single platform

The purpose of this paper is to demonstrate the feasibility of developing a single lab-on-a-chip (LOC) platform capable of performing dual, simultaneous separation and detection of multiple analytes. Computational modeling was performed to determine optimum device geometry and performance. The soda-lime glass-based device was fabricated using traditional microtechnology processes, including UV photolithography, buffered oxide etch (BOE), electrode deposition and compression thermal bonding. The device was characterized with a mixture of dopamine (2mM) and catechol (2mM) in a phosphate buffer (20mM, 6.5 pH). Modeling results yielded migration velocities of 0.6 mm/s and 0.42 mm/s for dopamine (electrokinetic (EK) mobility=60,000 /spl mu/m/sup 2//V/spl middot/s) and catechol (EK mobility=42,000 /spl mu/m/sup 2//V/spl middot/s), respectively. Experimental results obtained from microchips exhibiting the same EK mobilities demonstrated identical electropherograms in both detection channels with migration velocities of 0.58 mm/s for dopamine and 0.41 mm/s for catechol.

Electrochemical Dissolved Oxygen Removal from Microfluidic Streams for LOC Sample Pretreatment

Current water quality monitoring for heavy metal contaminants largely results in analytical snapshots at a particular time and place. Therefore, we have been interested in miniaturized and inexpensive sensors suitable for long-term, real-time monitoring of the drinking water distribution grid, industrial wastewater effluents, and even rivers and lakes. Among the biggest challenges for such sensors are the issues of in-field device calibration and sample pretreatment. Previously, we have demonstrated use of coulometric stripping analysis for calibration-free determination of copper and mercury. For more negatively reduced metals, O2 reduction interferes with stripping analysis; hence, most electroanalysis techniques rely on pretreatments to remove dissolved oxygen (DO). Current strategies for portable DO removal offer limited practicality, because of their complexity, and often cause inadvertent sample alterations. Therefore, we have designed an indirect in-line electrochemical DO removal device (EDOR), utilizing a silver cathode to reduce DO in a chamber that is fluidically isolated from the sample stream by an O2-permeable membrane. The resulting concentration gradient supports passive DO diffusion from the sample stream into the deoxygenation chamber. The DO levels in the sample stream were determined by cyclic voltammetry (CV) and amperometry at a custom thin-layer cell (TLC) detector. Results show removal of 98% of the DO in a test sample at flow rates approaching 50 μL/min and power consumption as low as 165 mW h L(-1) at steady state. Besides our specific stripping application, this device is well-suited for LOC applications where miniaturized DO removal and/or regulation are desirable.

Design and Characterization of a Signal Conditioning Microchip and Thin-Film Microelectrode Array for High Spatial Resolution Cardiac Mapping

A Thin-Film Microelectrode Array (TFMEA) capable of recording electrical activities of the heart with high spatial resolution, on the order of individual myocytes, was developed and characterized to study the mechanisms of arrhythmias. A customized multichannel signal conditioning microchip (SCM) consisting of a pre-amplification stage and band-pass filter was interfaced directly to the electrodes on the TFMEA to improve the signal-to-noise ratios (SNRs). The results suggest that the TFMEA-SCM system will greatly enhance the spatial resolution and improve signal quality for high resolution cardiac mapping.

Characterization of thin-film microelectrode array and signal conditioning microchip for high spatial resolution Surface Laplacian measurement

Surface Laplacian (SL) electrograms have been found to be superior to unipolar electrograms in detecting local activation by providing local activation time, activation block and wavefront initiation information. Traditional electrode configurations do not provide adequate spatial resolution or signal-to-noise ratios (SNRs) to measure SLs accurately in vivo. The purpose of this work is to develop a customized thin-film microelectrode array (TFMEA) with a custom multichannel signal conditioning microchip (SCM) that significantly improves the SNRs to make high-quality SL measurements feasible. In vitro and in vivo testing and characterization results clearly demonstrate the ability of the TFMEA-SCM system to obtain detailed SL measurements.