Description
The purpose of the Analytical Instrumentation Facility Core is to support Center investigators in the detection, characterization, and quantification of chemicals and biomolecules.
The specific aims of Facility Core 2 are:
- To provide sensitive protein identification and modification analyses;
- To offer services for the identification and quantification of metabolites, nutrients, xenobiotics, and various types of macromolecular damage;
- To implement novel analytical methods and improve the sensitivity and throughput of existing analyses;
- To provide consultation on the selection and implementation of analytical methods and training opportunities in usage and applications of analytical instrumentation;
- To develop tissue and biofluid analytical assays relevant to translational research in coordination with the Integrated Health Sciences Facility Core;
- To foster partnerships for improved access to high end analytical instrumentation and technical expertise in support of Center research.
The Analytical Facility Core will provide sophisticated mass spectrometry resources and expertise to support Center Investigator research efforts. It will neither unnecessarily duplicate analytical capabilities of investigators' laboratories nor become a processing center for large numbers of routine analyses. The goals of the Core have changed slightly during the past funding period to reflect developments in the fields of proteomics, metabolomics and biomarker identification, and the increased interest in these new technologies by Center Members. Acquisition of major new instrumentation has enabled significant improvements in existing services as well as expansion into novel assays applied to protein, drug and metabolite analyses. The facility has worked in close cooperation with the Molecular Biology Facility Core to integrate services required for protein expression profiling projects and analysis of serum samples. We anticipate a similar degree of cooperation with the proposed Integrative Health Science Facility Core which will utilize assays developed for analysis of tissue and biofluid samples to investigate the role of environmental factors in human disease.
Significance
Research into the mechanisms of how environmental exposures impact human diseases requires an understanding of the biomolecular pathways involved and identification of biomarkers that relate to pathophysiologic phenotypes. The impact of UV on skin cancer, the link between exogenous endocrine disruptors and neurotoxicology, and chemical toxicant exposure in migrant farm workers are examples of the diversity of problems linking the environment and human disease. Integration of scientific information on many levels is required to understand the process of disease development and aid in the identification of strategies to improve human health. Studies using transgenic mice provide information on exposure in a controlled genetic background while other animal models (e.g. Xiphophorus fish) can be used for determining exposure levels of harmful ultraviolet radiation. Human tissue and serum samples from epidemiologic studies provide resources to use for understanding pathways from initial exposure to ultimate disease. Collaboration among basic, clinical, and public health researchers fosters integration of this research. Technological integration supports research goals by bringing together sophisticated instrumentation and methodologies not easily replicated in individual laboratories. Monitoring of processes from genomics to proteomics to metabolomics facilitates meta-analysis of complex environmentally incited diseases. The genetic components that affect susceptibility to disease or translate exposure to illness are currently being studied using DNA and RNA based technologies such as microarrays, SAGE and RAGE. Epigenetic influences on gene expression are susceptible to environmental factors which can influence the induction or timecourse of diseases. Downstream products of metabolism serve as temporal markers of the progression from exposure to pathology and thus require sensitive detection when present in limited quantity in a complex milieu. These include proteins, metabolites, and DNA modified by environmental agents. Exogenous compounds ingested or absorbed also can be detected directly or via their metabolites. Analytical assays for detection and quantification of chemicals and biomolecules can be utilized at every level of scientific integration, from hypothesis driven research on the consequences of DNA damage and repair, to hypothesis generating studies of global protein expression level changes, to measurement of nutrients. Sophisticated HPLC and mass spectrometry instrumentation provide a variety of platforms for sensitive detection of many types of biomolecules. Partnership with laboratories and core facilities creates an interdisciplinary team approach to the study of environmental exposure and its impact on human health.
Understanding individual pathways as well as global changes to the cell, tissue or organism requires sensitive detection methods for identifying proteins and their modifications. Assessment of global changes in protein expression patterns is possible through utilization of 2D gel electrophoresis-mass spectrometry based analytical techniques and parallel strategies are available for quantitative proteomics using isotope labeling, HPLC chromatogroaphy and mass spectrometry. Access to state-of-the-art instrumentation and technology experts is critical for the successful design and implementation of such experiments. Improvements in mass spectrometry instrumentation and software have led to femtomole sensitivity for analyzing protein samples, while growing protein and EST databases enable protein identification for an increasing variety of species. Complete genomes for human and mouse facilitate research on human tissue and biofluids and transgenic mouse models. Protein modifications and interactions can also be studied with mass spectrometry based techniques. Sensitivity is achieved by scanning for marker ions or selective enrichment protocols. Greater diversity is exhibited in the class of biomolecules known as metabolites, with a variety of techniques needed to assess these molecules. Electrochemical detection provides an extremely sensitive assay for neurotransmitters or oxidized DNA. GC-MS enables quantification of amino acids, fatty acids and lipids.
Translating knowledge of molecular pathways to improvements for human health requires detection of specific and sensitive biomarkers in readily obtainable human material. Measuring levels of oxidative DNA damage or lipids and metabolites associates a quantitative molecular marker with phenotypic disease states. Epidemiological studies benefit from biomolecular measurements of drug delivery, oxidative macromolecular damage, and identification of biomarkers. Understanding of energy balance and effects of nutritional supplements in human populations benefits from correlation of survey data with quantification of relevant compounds present in serum. All these activities are supported by Facility Core 2 resources and services, with the primary aim of implementing the goals of the Center. Work at the Analytical Instrumentation Facility Core contributed to 52 publications and supported over 50 grants for 36 Center members and 5 Pilot Projects.
Staff
Maria D. Person, Ph.D., Facility Director
H. H. Stony Lo, Ph.D., Facility Manager
