Dietmar Puchberger-Enengl

Microfluidic concentration of bacteria by on-chip electrophoresis

In this contribution, we present a system for efficient preconcentration of pathogens without affecting their viability. Development of miniaturized molecular diagnostic kits requires concentration of the sample, molecule extraction, amplification, and detection. In consequence of low analyte concentrations in real-world samples, preconcentration is a critical step within this workflow. Bacteria and viruses exhibit a negative surface charge and thus can be electrophoretically captured from a continuous flow. The concept of phaseguides was applied to define gel membranes, which enable effective and reversible collection of the target species. E. coli of the strains XL1-blue and K12 were used to evaluate the performance of the device. By suppression of the electroosmotic flow both strains were captured with efficiencies of up to 99%. At a continuous flow of 15 μl/min concentration factors of 50.17 ± 2.23 and 47.36 ± 1.72 were achieved in less than 27 min for XL1-blue and K12, respectively. These results indicate that free flow electrophoresis enables efficient concentration of bacteria and the presented device can contribute to rapid analyses of swab-derived samples.

Microfluidic Vortex Enhancement for on-Chip Sample Preparation

In the past decade a large amount of analysis techniques have been scaled down to the microfluidic level. However, in many cases the necessary sample preparation, such as separation, mixing and concentration, remains to be performed off-chip. This represents a major hurdle for the introduction of miniaturized sample-in/answer-out systems, preventing the exploitation of microfluidic’s potential for small, rapid and accurate diagnostic products. New flow engineering methods are required to address this hitherto insufficiently studied aspect. One microfluidic tool that can be used to miniaturize and integrate sample preparation procedures are microvortices. They have been successfully applied as microcentrifuges, mixers, particle separators, to name but a few. In this work, we utilize a novel corner structure at a sudden channel expansion of a microfluidic chip to enhance the formation of a microvortex. For a maximum area of the microvortex, both chip geometry and corner structure were optimized with a computational fluid dynamic (CFD) model. Fluorescent particle trace measurements with the optimized design prove that the corner structure increases the size of the vortex. Furthermore, vortices are induced by the corner structure at low flow rates while no recirculation is observed without a corner structure. Finally, successful separation of plasma from human blood was accomplished, demonstrating a potential application for clinical sample preparation. The extracted plasma was characterized by a flow cytometer and compared to plasma obtained from a standard benchtop centrifuge and from chips without a corner structure.

Hydrogel-based microfluidic incubator for microorganism cultivation and analyses

This work presents an array of microfluidic chambers for on-chip culturing of microorganisms in static and continuous shear-free operation modes. The unique design comprises an in-situ polymerized hydrogel that forms gas and reagent permeable culture wells in a glass chip. Utilizing a hydrophilic substrate increases usability by autonomous capillary priming. The thin gel barrier enables efficient oxygen supply and facilitates on-chip analysis by chemical access through the gel without introducing a disturbing flow to the culture. Trapping the suspended microorganisms inside a gel well allows for a much simpler fabrication than in conventional trapping devices as the minimal feature size does not depend on cell size. Nutrients and drugs are provided on-chip in the gel for a self-contained and user-friendly handling. Rapid antibiotic testing in static cultures with strains of Enterococcus faecalis and Escherichia coli is presented. Cell seeding and diffusive medium supply is provided by phaseguide technology, enabling simple operation of continuous culturing with a great flexibility. Cells of Saccharomyces cerevisiae are utilized as a model to demonstrate continuous on-chip culturing.

A mobile lab-on-a-chip device for on-site soil nutrient analysis

Abstract In this paper, a mobile sensor for on-site analysis of soil sample extracts is presented. As a versatile tool for scanning ion concentrations in liquid samples, it especially allows the analysis of NO3, NH4, K and PO4. The sensor mainly consists of a microfluidic chip in which the sample ions are separated in an electric field (capillary electrophoresis) and the individual ion concentrations are detected by a conductivity measurement. For the adaption of the device to field conditions, two major concerns were addressed. Firstly, nano-porous material was used as a barrier between the sample container and the analysis channel of the microfluidic chip. This prevents pressure driven leakage of the sample into the chip due to non-horizontal orientation of the device. Secondly, a new method for the injection of the sample into the chip was used. It reduces the number of fluidic connections between chip and operation device to three instead of the commonly used four connections. The sensor performance was tested on multi-ion solutions with calibration series for NO3, NH4, K and PO4. For the first on-site test, a quick soil nutrient extraction procedure with water was used. The sensor data was compared to standard laboratory results. The potential of the sensor for soil nutrient analysis is discussed together with required improvements of the sensor performance and of the nutrient extraction procedure.

Organically modified silicate film pH sensor for continuous wound monitoring

In this contribution we present a miniaturized optical reflectance pH sensor based on organically modified silicate for continuous measurements of wound pH. Colorimetric indicators (bromocresol green pH 3.8–5.4 and bromocresol purple pH 5.2–6.8) have been immobilized in tetraethoxysilane (TEOS) thin films. Characteristics of the thin films have been optimized by addition of (3-Glycidyloxypropyl)trimethoxysilane (GLYMO). The pH sensitive layer of the sensor is illuminated through the substrate by a chip-LED. The reflected light at a phototransistor varies in dependence of the color of the thin film. The films have been characterized by visible absorption spectroscopy and the performance of the sensor has been evaluated by measurements on artificial wounds.

In-Line Characterization of Micro-Droplets Based on Partial Light Reflection at the Solid-Liquid Interface

In this paper a novel optofluidic setup, fabricated on a single layer device for in line droplet characterization yielding droplet size, droplet frequency, and optical proper ties with compatibility for full on chip integration is presented. Chips were fabricated using a simple, fast, and cost effective technology. A T junction arrangement on the device is used for droplet generation. The optical part of the setup consists of an external light source, external silicon photodetectors, integrated air micro lenses, and an integrated waveguide. The design makes use of partial light reflection/transmission at the solid liquid interface to count, size, and discriminate droplets based on their optical properties. When passing the interrogation point, droplets having a lower refractive index as the continuous phase result in light deflections. Both, reflected and transmitted light, are detected simultaneously. A relation of those two signals is then used for the analysis resulting in a continuously stable signal. The generated pattern is unique for different droplets and can be exploited for droplet characterization. Using this arrangement, droplets of de ionized water (DI) were counted at frequencies of up to 320 droplets per second. In addition, information about the droplet sizes and their variations could be obtained. Finally, 5 mol/L CaCl 2 and DI droplets, having different indices of refraction were examined and could clearly be discriminated based on their unique reflected and transmitted light signals. This principle can be applied for the detection of dissolved molecules in droplets as long as they influence the index of refraction. Examples could be the determination of DNA or protein content in the droplet.

In-line characterization and identification of micro-droplets on-chip

Abstract We present an integrated optofluidic sensor system for in-line characterization of micro-droplets. The device provides information about the droplet generation frequency, the droplet volume, and the content of the droplet. Due to its simplicity this principle can easily be implemented with other microfluidic components on one and the same device. The sensor is based on total internal reflection phenomena. Droplets are pushed through a microfluidic channel which is hit by slightly diverging monochromatic light. At the solid-liquid interface parts of the rays experience total internal reflection while another part is transmitted. The ratio of reflected to transmitted light depends on the refractive index of the solution. Both signals are recorded simultaneously and provide a very stable output signal for the droplet characterization. With the proposed system passing droplets were counted up to 320 droplets per second and droplets with different volumes could be discriminated. In a final experiment droplets with different amounts of dissolved CaCl2 were distinguished based on their reflected and transmitted light pattern. This principle can be applied for the detection of any molecules in microdroplets which significantly influence the refractive index of the buffer solution.

A new injection method for soil nutrient analysis in capillary electrophoresis

We present a new method for the direct injection of liquid sample into a capillary electrophoresis (CE) device. Instead of a double-T injection mechanism, a single inlet provided with a membrane filter is used. From a reservoir on top of this inlet, the liquid directly enters the separation channel through the membrane. The driving force is a short electrical pulse. This avoids an additional sample channel, so that the chip needs only three microfluidic connects and no mechanical sample pumping is demanded. The high injection reproducibility and the comparatively simple setup open up the way for mobile application of soil analysis.

Hydrogel plug for independent sample and buffer handling in continuous microchip capillary electrophoresis

In microchip capillary electrophoresis most frequently electrokinetic sample injection is utilized, which does not allow pressure driven sample handling and is sensitive for pressure drops due to different reservoir levels. For efficient field tests a multitude of samples have to be processed with the least amount of external equipment. We present the use of a hydrogel plug to separate the sample from clean buffer to enable independent sample change and buffer refreshment. In-situ polymerization of the gel does away with complex membrane fabrication techniques. The sample is electrokinetically injected through the gel and subsequently separated by a voltage between the second gel inlet and the buffer outlet. By blocking of disturbing flows by the gel barrier a well-defined ion plug is obtained. After each experiment, the sample and the separation channel can be flushed independently, allowing for a continuous operation mode in order to process multiple samples.