Emil J. Geiger

High frequency dielectrophoretic response of microalgae over time.

The high frequency dielectrophoresis (>20 MHz) response of microalgae cells with different lipid content was monitored over time. Chlamydomonas reinhardtii was cultured in regular medium and under nitrogen‐depleted conditions in order to produce populations of cells with low and high lipid content, respectively. The electrical conductivity of the culture media was also monitored over the same time. The upper crossover frequency decreased for high‐lipid cells over time. The single‐shell model predicts that the upper crossover frequency is dictated primarily by the dielectric properties of the cytoplasm. The high frequency DEP response of the high‐lipid cells’ cytoplasm was changed by lipid accumulation. DEP response of the low‐lipid cells also varied with the conductivity of the culture media due to nutrient consumption. Relative lipid content was estimated with BODIPY 505/515 dye by calculating the area‐weighted intensity average of fluorescent images. Finally, microalgae cells were successfully separated based on lipid content at 41 MHz and DEP media conductivity 106 ± 1 μS/cm.

Liposomes as a model for the study of high frequency dielectrophoresis

Liposomes were used as a physical model to study the dielectrophoretic response of single‐shelled particles at high frequencies. For a typical particle, the single‐shelled theoretical model predicts a lower cross‐over frequency that depends upon the dielectric properties of the shell and an upper crossover frequency that depends upon the dielectric properties of the interior. Dried liposomes were rehydrated in media with conductivity ranging from 100 to 2000 μS/cm. The high frequency dielectrophoresis response of the liposomes was observed in the range of 1–80 MHz at 30 volts peak‐to‐peak, and the upper cross‐over frequency was recorded. The experimental results closely matched the theoretical expectations. In particular, the upper cross‐over frequency ranged from 9 to 60 MHz and was found to depend linearly on the interior conductivity of the liposome. These results further confirm the single‐shell model at high‐frequencies. Moreover, they suggest liposomes may be a useful model particle for use during the development of dielectrophoresis‐based devices.

Wireless ISFET pH Sensor Network for Offshore Microalgae Cultivation

Microalgae technology continues to show tremendous promise for becoming a major source of renewable transportation fuel in the coming decades. However, for microalgae to provide a significant fraction of the current US demand for fuel, their cultivation will be required on an enormous scale. One of the many formidable challenges that must be met to achieve this scale is the development of appropriate sensor networks to provide information about the growth conditions and the algae themselves. These sensors would monitor the heterogeneity of a) environmental parameters, such as pH, oxygen, and nutrient levels and b) algal characteristics such as size, oil content, and viability. Here we present a wireless sensor network to measure the local pH in NASA OMEGA project (Offshore Membrane Enclosures for Growing Algae). The pH is measured using Ion Sensitive Field Effect Transistor (ISFET) technology, which is more robust and has a faster response than traditional glass pH electrodes. A custom circuit drives the ISFET sensor and interfaces with an ANT wireless network system. The wireless network consists of a network hub which can service up to 8 sensor nodes and a series of relays to transmit the data to a PC. The data is logged with a custom LabVIEW program. In this work, we demonstrate operation of this network using a single ISFET pH sensor, one hub, and two relay units. The performance of the pH sensor network is evaluated and compared in parallel with an existing wired glass electrode based pH monitoring system at the NASA OMEGA project.Copyright

Epifluorescent direct-write photolithography for microfluidic applications

Abstract. We present a technique for fabricating soft-lithography molds created using an epifluorescent microscope. By focusing the UV light emitted from a Hg arc lamp, we demonstrate the ability to direct-write photoresist features with a minimum resolution of 45  μm. This resolution is satisfactory for many microfluidic applications. A major advantage of this technique is its low cost, both in terms of capital investment and on-going expenditures. Furthermore, by using a motorized stage, we can quickly fabricate a design on demand, eliminating the need, cost, and lead-time required for a photomask. With the addition of an electronic shutter, complicated separate structures can be imaged and utilized to make a wide range of microfluidic devices. We demonstrate this technique using dry-film resist due to its low cost, ease of application, and less stringent safety protocols.

Conducting Engineering Research Experiments in a Senior Mechanical Engineering Instructional Laboratory

Engineering teaching laboratory experiments often suffer from being too deterministic. While this can allow students to predictably observe and measure various engineering phenomena, the students may doubt the real world application or significance of the experiment. On the other hand, engineering research experiments can often be tedious and repetitive. Research assistants, both graduate and undergraduate, can grow bored while collecting data, leading to mistakes and reduced quality of the data. This paper will present preliminary results from using a laboratory section of a senior mechanical engineering elective course, “Introduction to Microtechnology,”, to conduct a series of experiments for a research problem in the instructor’s research laboratory.Copyright

Use of a Separability Parameter for the Design of a High Frequency Dielectrophoresis Cell Sorter Device

We present a method to quantify and enhance separation of binary cells mixture in the microfluidic device using high frequency dielectrophoresis (>20 MHz). At these frequencies, the DEP response depends primarily on the dielectric properties of the cytoplasm. In order to achieve efficient separation, there must be a difference in the intrinsic dielectric properties of the populations to be sorted. For algae cells, the shift in high frequency dielectrophoresis response during lipid accumulation can be used as a basis of separation. We defined a separability parameter based on the expected difference in the dielectrophoresis responses of the algae cells.Chlamydomonas reinhardtii cells were cultured in regular media and then the same cells were cultured under nitrogenfree conditions to accumulate neutral (non-polar) lipids. Separability of microalgae cells with different lipid content via high frequency dielectrophoresis were investigated by a thin needle shaped electrodes patterned by standard photolithographic and wet etching procedures. Experimental separability factors were measured by estimation of relative lipid content with BODIPY 505/515 fluorescence dye and calculating the area-weighted intensity average of fluorescent images. Theoretical separability parameter was calculated using analytical analysis of single shell model by MATLAB.Theoretical and experimental separability parameters, as tools to determine the optimal separation method, were calculated for microalgae cells with different lipid content. This objective function was maximized in the range of 35–45 MHz for C. reinhardtii cells after 21 days of lipid accumulation in a static separation. In order to design a continuous cell sorter device, the theoretical separation factor was maximized based on differences in the magnitude or the direction of the DEP force.Copyright

Microfluidic MEMS Device in the Cultivation of Microalgae With Positive Dielectrophoretic Cell Trapping for Media Exchange

Through COMSOL modeling and electrode design, positive dielectrophoretic (pDEP) cell trapping for media exchange has been demonstrated on live Chlamydomnas reinhardtii in regular growth medium in a PDMS-glass microfluidic MEMS device. Dielectrophoresis (DEP) is the force applied to dielectric particles in an alternating current (AC) non-uniform electric field. A DEP force toward the increasing electric field gradient is called positive (pDEP). There are several published DEP structures for various applications such as: simple interdigitated structures for particle sorting in flow, DEP tweezers for single cell manipulation, and spiral structures for general cell manipulation. pDEP trapping over large areas (area pDEP) has been demonstrated with the use of low conductivity suspending media, but for higher conductivity suspending media, such as growth media, the pDEP force is reduced, and less likely to trap and hold microalgae against the hydrodynamic forces during media exchange. Multiphysics software, COMSOL, was used to model repeating structures suited for trapping of cells over the bottom area of a microfluidic device, which is useful and necessary for media exchange of a cell culture in a simple microfluidic device. The theoretical model of dielectrophoretic (DEP) force on a homogenous sphere in a homogenous medium in an electric field is a function of the sphere radius and conductivity, medium permittivity, and the gradient of the electric field. By assuming the conductivities, permittivities, and the particle geometry remains constant, the gradient of the electric field is the determining factor for the strength of the pDEP force. Modeling the electric fields and the resulting electric field gradient of various interdigitated electrode configurations allowed for the optimization of an electrode structure’s area of higher electric field gradients. The completed microfluidic device consisted of a single channel and a wide growth chamber overlaid over patterned gold-chrome electrodes. The MEMS device was fabricated using soft lithography and photolithography on the etched chrome-gold glass slides. The pDEP trapping was successful in trapping C. reinhardtii for media exchange. Media exchange allows for nutrient replenishment and waste removal, allowing for control of the growth conditions.Copyright

Impedance Effects During High-Frequency Dielectrophoresis

Dielectrophoresis refers to the motion of electrically neutral particles in a spatially non-uniform electric field as a function of frequency. Analytical models predict a second or upper crossover frequency, but there is little experimental evidence to supporting this prediction. One reason for this is that standard bench top function generators are typically limited to maximum frequency of 20 MHz, and those with higher bandwidths are often unable to generate higher frequency electric fields with sufficient amplitude to induce DEP motion. Our experimental set-up seeks to sort cells on the basis of the upper crossover frequency in the range of 10–160 MHz. At these frequencies, the DEP response is expected to depend primarily on the dielectric properties of the cytoplasm.A needle pattern was used due to the ease of making spatially non-uniform electric fields. Sine waves were generated with a signal generator capable of generating sine waves with frequencies up to 160 MHz which was amplified with an RF amplifier. In order to help match the load to the amplifier and reduce the amount of reflected power an attenuator was placed between the amplifier and the electrodes. Two different amplifiers and attenuators were tested. Conductive copper tape and alligator clips were used to connect the glass slide to the electronic equipment. These systems were capable of 30 Volts peak-to-peak (Vpp) amplitudes in the given frequency range as monitored by an oscilloscope probing the electrodes.With this set-up, the amplitude of the signal was observed by oscilloscope to vary as much as 40 Vpp across the range of frequencies which is supplied by signal generator. We suspect that the impedance of the experimental set-up was highly frequency dependent at the high frequencies used in this study. Furthermore, the oscilloscope may also be enough of a load on the system to affect the impedance. To maintain the 30 Vpp output, the input amplitude to the RF amplifier was adjusted at each individual frequency across all frequencies of the DEP experiment.Copyright

Effect of Media Conductivity on High Frequency Dielectrophoretic Response

A method was developed for analyzing the high frequency dielectrophoresis (>20 MHz) response of microalgal cells with different lipid content on the basis of media conductivities. At these frequencies, the DEP response is expected to depend primarily on the dielectric properties of the cytoplasm. The medium used to perform DEP experiments has an important influence on their outcome since all properties are measured in relation to the medium, therefore its properties must be tightly controlled. This method is label free and only uses electric fields, and thus it is suitable for many other applications in the medical field.Chlamydomonas reinhardtii cells were cultured in regular medium and then the same cells were cultured under nitrogen-free conditions to accumulate neutral (non-polar) lipids. To measure the upper crossover frequency, two thin, needle shaped electrodes (100 nm thick) were patterned onto a glass slide using standard micro-technology processes and the motion of the cells was observed as an AC signal was swept from 20–80 MHz at ∼30Vpp (Volt peak to peak). Cells were injected to the microelectrode array in different medium conductivities that were adjusted by sodium chloride.The upper crossover frequencies of N-free cells in different media conductivities were measured. It was found that when the solution conductivity was between 4 ± 2 μs/cm and 106 ± 1 μS/cm and the operating frequency was between 20 and 80 MHz, the upper crossover frequency of the microalgae increased as the medium conductivity increased. The lipid content was verified via fluorescence microscopy and MATLAB image processing.Copyright

Maskless Photolithography Using an Epifluorescent Microscope for Microfluidic Applications

We present a technique for fabricating molds suitable for casting PDMS based microfluidic devices using equipment readily available to most microfluidic researchers, namely an epi-fluorescent microscope. By focusing the UV light emitted from the Hg arc lamp, we demonstrate the ability to direct-write photoresist features with a minimum resolution of 45 μm. A major advantage of this technique is its low-cost, both in terms of capital investment and on-going expenditures. Furthermore, by using a motorized stage, we can quickly fabricate a design on demand, eliminating the need, cost, and time required for a photomask. We demonstrate this technique using dry-film resist, due to its low-cost, ease of application, and less stringent safety protocols. In the future, we plan to expand our efforts to higher resolution photoresists such as SU-8.Copyright