General

Fast and accurate chemical analysis methods are required to support current biomedical, pharmaceutical, clinical, environmental, and anti-terrorism activities. The ultimate goal is to develop sophisticated and, in many cases, miniaturized instrumentation that can be used to interrogate a sample with sufficient selectivity, sensitivity, and speed to provide reliable qualitative and quantitative information in near real-time. My research group focuses on the development of new instrumentation and supporting technology in the fields of capillary and microfluidic separations, mass spectrometry, and ion mobility spectrometry. Major research areas in past years include column technology for capillary chromatography and electrophoresis, instrumentation for capillary supercritical fluid chromatography, and instrumentation for time-of-flight mass spectrometry. Current research interests are described in the following paragraphs.

Polymeric Monolithic Column Technology for Capillary and Microfluidic Separations

A monolithic column is a continuous filling in a column or channel with canal-like micrometer-size through-pores and nanometer-size pores in the skeletal structure. Monoliths can be used as stationary phases in liquid chromatography, media for isolation and concentration of target analytes from mixtures, and media for reducing laminar flow dispersion in certain microfluidic operations. Monoliths are of interest because of their ease of preparation and enhanced mass transfer characteristics. We have been developing polymer monolithic materials for capillary and microchip liquid chromatography, affinity concentration before capillary electrophoresis, and dispersion reduction in electric field gradient focusing. These monolithic materials have been based on polyethylene glycol-containing acrylate monomers and crosslinkers, which have been found to resist adsorption of peptides and proteins.

Electric Field Gradient Focusing

The protein distribution in the blood is extremely sensitive to cellular conditions in the body, and consists of proteins having abundances that are dependent on age, disease state, and environmental conditions. Disease marker proteins, proteins whose expression changes during the progression of a disease, have been associated with human diseases, such as cancer, Alzheimer's, schizophrenia, and Parkinson's. Rapid screening of the protein profile in a blood sample would be desirable to identify multiple target proteins which could be used to develop more definitive assays. Unfortunately, protein analysis is an extremely difficult task because of the sheer number of proteins in biological systems and their dynamic nature. There are vastly different concentrations of the various proteins and numerous possible interactions among proteins and other ligands. Expressed proteins are often further modified by reactions such as phosporylation, glycosylation, carbamylation, deamidation, and truncation.

In order to monitor multiple cancer marker proteins in blood samples, the specific target proteins must be separated from the numerous other proteins present at vastly different concentrations. We are exploring a new technology called electric field gradient focusing (EFGF) which promises to have higher resolving power, greater speed, and better sensitivity than other protein separation techniques. EFGF is similar to capillary electrophoresis, except that the separation occurs in an electric field gradient with a counter liquid flow. Each charged analyte migrates in the separation capillary or channel until it reaches its dynamic equilibrium position where its electrophoretic migration velocity just balances the constant opposing flow velocity. The focused bands are prevented from diffusional broadening by restoring forces imposed by the electric field gradient and opposing flow. Using EFGF, we have isolated and concentrated trace proteins by more than 10,000 times.

Ion Mobility Spectrometry Trapping and Analysis

In conventional ion mobility spectrometry, an electric field produces a linear relationship between the drift velocity and electric field. A sample is introduced into an ionization region containing an ion source. Ionized samples are then accelerated into a drift tube, often with a buffer gas introduced from the opposite direction. Ions are separated as they drift through this buffer gas. Separation of ions is based on size, shape, and charge of the ions. Separations in ion mobility spectrometry occur in the millisecond time frame, so it is considered to be a high speed separation method. It is also very sensitive. However, IMS has generally received little attention because of its relatively poor resolution, limited linear dynamic range, and low to moderate selectivity.

We have been developing two novel ion mobility spectrometry technologies that promise greatly improve resolution, dynamic range, and selectivity. The so-called flow-balanced ion mobility analyzer is based on separating ions in an electric field against a counter flow of inert gas. A constant, uniform gas flow is generated using a wind tunnel. Ions that have a migration velocity that is equal and opposite to the gas flow velocity become suspended in the gas flow. Superposition of a slight flow from one side of the wind tunnel to the other carries the levitating ions to a detector. We are also developing an ion mobility trap, similar to an ion trap mass spectrometer, based on high field asymmetric waveform ion mobility spectrometry. In this technique, a high electric field intensity is applied between two surfaces for a short time, and then a low electric field intensity is applied in the opposite direction for a longer duration, with the average applied electric field being balanced. Ions oscillate back and forth between the two surfaces and stay suspended, depending on their differential mobilities. Ions are trapped in the volume between the surfaces based on the nonlinear dependence of their mobility constants with respect to the electric field intensity. An obvious extension of this work is to perform multidimensional ion mobility and mass-to-charge analysis in the same device merely by changing the applied waveform.

Hand-Portable Gas Chromatography-Mass Spectrometry.

We are developing a novel hand-portable system for point detection of chemical warfare agents and environmental chemicals. The detection system is based on the integration of solid phase microextraction sampling, low thermal mass gas chromatography separation, and ion trap mass analysis. The heart of the detection system is a novel toroidal ion trap that increases the trapping volume compared to a conventional ion trap by approximately 400 times. This allows miniaturization of the ion trap to the size of a US half dollar and minimization of the power requirements such that the trap can be operated using a conventional 12/24 volt military battery. In addition to the development and testing of the novel toroidal ion trap, selective polymer coatings for solid phase microextraction and alternative approaches for high speed gas chromatography are being investigated. Furthermore, we are developing methods for catalytically generating volatile biomarkers from biological warfare agents which can be detected using the hand-portable GC-MS system.