Epigenetics, the study of inheritable mechanisms that regulate gene expression, has clinical ramifications from cancer to autoimmune disorders to psychiatric pathologies. The main tool to study epigenetics is chromatin immunoprecipitation (ChIP), which probes the relationship between DNA and its structural nucleosome-forming histone proteins. Standard benchtop ChIP has three major drawbacks: (1) it requires a large input volume of cells, (2) it is very time consuming and work intensive, and (3) it is low throughput. Digital microfluidic biochips (DMFB) have proven to be successful at utilizing small volumes of reagents and samples to perform high throughput bioanalyses and assays of macromolecules. Their ease of configurability, automation, and high sensitivity make them an ideal platform for ChIP adaptation, addressing the three biggest issues facing epigenetic study and workflow. Herein, we demonstrate the first step towards ChIP implementation on a DMFB by detecting specifically modified nucleosomes, the building blocks of chromatin, in a nucleosome immunoprecipitation assay. Using magnetic beads to capture the nucleosomes with magnetic fields generated by embedded current wires and fluorescent conjugated antibodies for detection, this DMFB system allows complete on-chip isolation and detection without the need for external magnets or specialized fluoroscopy equipment. This assay design can be adapted to probe for multiple specific nucleosome modifications, thus establishing a rapid screening method for antibody specificity and sensitivity. Most importantly, this novel confirmatory checkpoint, currently unavailable when running ChIP, ensures that the target analyte has been isolated prior to intensive downstream analyses such as PCR and sequencing.
MALDI-MS (matrix-assisted laser desorption/ionization mass spectrometry) is one of the most commonly used techniques for protein analysis. In conventional systems sample preparation is typically done in well-plates and transferred onto a MALDI target by robotic systems, which are complex, huge, expensive and slow. In this paper, we present a droplet-based microfluidic interface to transfer protein samples from a well-plate format onto a MALDI target for MS analysis. The droplets are actuated using the electrowetting phenomenon, and are immersed in silicone oil which prevents non-specific adsorption and enables the manipulation of high concentrations of proteins. Droplet transport and droplet formation were evaluated as a function of protein concentration using bovine serum albumin (BSA) as a test system. Droplet transport was possible for BSA concentrations up to 10mg/mL which is three orders of magnitude higher than previously reported results on handling proteins by electrowetting. Droplet formation from on-chip reservoirs, using only electrowetting forces and no external pressure assistance, was possible up to concentrations of 0.01mg/mL. An interface between a well-plate format and the electrowetting chip, and a scheme to passively stamp droplets onto a target substrate was then designed and tested by stamping BSA solutions. In two separate experiments 3.6fmoles and 16fmoles of BSA were stamped onto a glass slide using 0.001mg/mL and 0.01mg/mL samples respectively. A protein mixture with known constituents (ABI 4700 proteomics analyzer calibration solution) was stamped onto a MALDI plate and the individual proteins were correctly identified in the mass spectrum obtained using MALDI-TOF MS. The preliminary results establish the feasibility of using an electrowetting-based microfluidic system to handle proteins especially for protein stamping applications. The proposed system has a small footprint, is easy to control, and is very fast compared to conventional robotic systems. In addition, there are no moving parts and the associated mechanical reliability issues. Future work involves scaling to a larger number of samples and integration of sample preparation steps on-chip.
An ideal on-site chemical/biochemical analysis system must be inexpensive, sensitive, fully automated and integrated, reliable, and compatible with a broad range of samples. The advent of digital microfluidic lab-on-a-chip (LoC) technology offers such a detection system due to the advantages in portability, reduction of the volumes of the sample and reagents, faster analysis times, increased automation, low power consumption, compatibility with mass manufacturing, and high throughput. We describe progress towards integrating sample collection onto a digital microfluidic LoC that is a component of a cascade impactor device. The sample collection is performed by impacting airborne particles directly onto the surface of the chip. After the collection phase, the surface of the chip is washed with a micro-droplet of solvent. The droplet will be digitally directed across the impaction surface, dissolving sample constituents. Because of the very small droplet volume used for extraction of the sample from a wide colection area, the resulting solution is realatively concentrated and the analytes can be detected after a very short sampling time (1 min) due to such pre-concentration. After the washing phase, the droplet is mixed with specific reagents that produce colored reaction products. The concentration of the analyte is quantitatively determined by measuring absorption at target wavelengths using a simple light emitting diode and photodiode setup. Specific applications include automatic measurements of major inorganic ions in aerosols, such as sulfate, nitrate and ammonium, with a time resolution of 1 min and a detection limit of 30 nm/m3. We have already demonstrated the detection and quantification of nitroaromatic explosives without integrating the sample collection. Other applications being developed include airborne bioagent detection.
MEMS technology was used to fabricate bimetallic cantilever sensor for detecting the TNT and DNT residue found in mien fields. A number of experiments yielded reproducible result for the detection of pure 2,4-Dinitrotoluene nanogram particles. A few experiments were performed unsuccessfully to detect explosives directly from soil by placing it on the cantilevers. Alternatively, DNT and TNT were extracted from the soil using acetone and subsequently letting acetone to leave DNT and TNT as a residue. This residue has been placed on the cantilever for detection that yielded very uncertain result. This residua contains a number of other materials, which changes the physical properties of the residue considerable making it unfit for detection using microcantilevers. T
MEMS technology was used to fabricate arrays of sensor for detecting the explosive micro particulate residue found in mine fields. MEMS devices were fabricated by a surface micromachining process provided by MCNC. The sensor consists of a bimorph structure of polysilicon/gold cantilevers. An optical detection system was designed to detect the deflection of the cantilevers. The sensors were heated by either radiation or conduction using an UV lamp and a small heater under the sensor chip respectively. The deflection of the cantilevers with increasing temperature is presented. Experiments have been performed to detect the response of the cantilevers in the presence of an explosive particle. The cantilevers show a response due to the presence due to the presence of nanograms of TNT and RDX in the vicinity of the cantilevers. Currently it is understood that the response shown by the cantilevers is due to the vaporization of the micro particles, which pulls significant heat out of the temperature sensitive beams causing detectable beam motion. The chemical selectivity of the sensor is provided by the unique melting temperatures of TNT and RDX.
We report on the development of MEMS devices for detecting explosive particles associated with anti-personnel mines. Because of the affinity of explosive substances for surfaces and owing to the high partition coefficients of explosives in soils relative to water and air, we employ remote stimulation of the soil's surface with a high intensity, focused air ultrasonic beam whose energy can megasonically clean the target area of particles above a designed-for size. We have fabricated a MEMS electrostatic transducer to test the concept. Nanogram particle detection will occur by collecting particles on an array of temperature sensitive MEMS sensors and irradiating the particles with 3 - 5 micrometer wavelength infrared light. Explosive particles will selectively absorb the infrared energy at approximately 1600 cm-1, decompose, and give off heat which can be detected. Prototype explosive detectors have been fabricated which do not absorb energy in the peak absorption bands of the explosives, thus allowing for selective particle heating without heating the sensor device itself.
We report on a new methodology for chemical detection of explosive particles associated with anti-personnel mines. Trace particle detection can be used to complement vapor detection of explosives with an electronic noise. Our approach is to remotely stimulate a target area with a high intensity, focused air ultrasonic beam whose energy can megasonically clean the target area of particles above a designed for size. We have designed a MEMS electrostatic transducer to test the concept. Nanogram particle detection will occur by collecting particles on an array of temperature sensitive MEMS sensors and irradiating the particles with 3-5 micrometers wavelength IR light. Explosive particles will selectively absorb the IR energy at approximately 1600 cm-1, decompose, and give off heat which can be detected.
Integrated Force Arrays (IFAs) are thin film linear actuators which operate with substantial displacement and force. The methods of attachment of these devices to external systems are under development. Our current methods to incorporate IFAs in an scanning ultrasound imaging systems as well as a new material and method for attachment will be described.
Integrated Force Arrays (IFAs) are thin film membrane actuators that act as transfer devices for electrostatic force. They are capable of large amplitude motion and evidence significant energies per unit volume (eg. 8.2 erg/mm3). Devices which use IFAs as drivers to scan PZT acoustic imaging transducers are under development and will be discussed here.