UC San Diego

In biomedical research and diagnostics, it is a significant challenge to directly isolate and identify rare cells and potential biomarkers in blood, plasma and other clinical samples. Additionally, the advent of bio- nanotechnology is leading to numerous drug delivery approaches that involve encapsulation of drugs and imaging agents within nanoparticles, which now will also have to be identified and separated from blood and plasma. AC electrokinetic techniques such as dielectrophoresis (DEP) offer a particularly attractive mechanism for the separation of cells and nanoparticles. Unfortunately, present DEP techniques require the dilution of blood/ plasma, thus making the technology less suitable for clinical sample preparation. The current dissertation explores methods and techniques of overcoming the high conductivity barrier in AC Electrokinetics to isolate nanoparticles. It further explores the various applications of isolating nanoparticles from complex physiological solutions. Using array devices with microelectrodes over-coated with porous hydrogel layers, AC electric field conditions have been found which allow the separation of DNA nanoparticles to be achieved under high conductance (ionic strength) conditions. At AC frequencies in 3000Hz to 10,000Hz range and 10 volts peak- to-peak, the separation of 10 micron polystyrene particles into low field regions, and 60nm DNA derivatized nanoparticles and 200nm nanoparticles into high field regions was carried out in 149mM 1xPBS buffer (1.68S/m). Furthermore, SYBR-Green fluorescent-stained hmw-DNA was separated from whole undiluted blood, at a level of <260ng /ml. While this DEP process concentrates hmw-DNA (>10kb) in high-field areas and cells into the low-field areas, very low molecular weight DNA <100bp and small protein molecules are only minimally affected by the process. These results may allow AC electrokinetic systems to now be developed, which can be used as seamless sample-to- answer systems for point-of-care diagnostics based on detection of early disease biomarkers directly from whole blood