Director, Protein Microarray Laboratory, Infectious Diseases
School of Medicine
Project Scientist, Epidemiology
School of Medicine
Ph.D., Michigan State University, 1978, Neurosciences and Biochemistry
M.S., Michigan State University, 1975, Neurosciences and Biochemistry
B.S., Michigan State University, 1972
Phone: (949) 824-1407, 0460
Fax: (949) 824-5490
University of California, Irvine
3052 Hewitt Hall
Mail Code: 4068
Irvine, CA 92697
Vaccines, Gene Therapy, Drug Delivery, Liposomes, Biophysics, Protein Microarray, Epidemiology
Postdoctoral Scholar -Biophysics, University of Virginia, Charlottesville,(1978)
Our Applied Proteomics Research Laboratory has developed a high throughput protein expression system called PCR Express which can be used to rapidly generate complete proteomes from any sequenced microorganism. The technology allows different genes to be expressed from their PCR products at the rate of hundreds of proteins per week, manually or with the aid of a robotic workstation. The proteomes can be made available in a purified soluble form, on proteome microarray chips, and as arrays of expression ready plasmid clones. At the heart of this technology is an in vivo recombination cloning method that allows PCR fragments to be rapidly inserted into expression plasmids in a high throughput manner. The plasmids can be expressed in an in vitro system, the resulting proteins spotted onto microarray chips, and the chips can be used to monitor antigen specific antibody titers against the arrayed proteins. A paper from our lab describing some aspects of this technology was published in the Journal of Biological Chemistry.
One of the ways to take advantage of proteome arrays from infectious microorganisms is in the area of vaccine antigen discovery and vaccine development. Our “Agile” vaccine development pathway, is intended to accelerate the discovery and deployment of safe and effective vaccines. One of the most difficult tasks in developing a recombinant protein subunit vaccine or DNA vaccine is the identification of the antigens that will stimulate the most effective immune response against the pathogen, particularly when the genome of the organism is large. A comprehensive way to accomplish this would be to obtain each of the structural, metabolic, and regulatory antigens of the pathogen and test their protective immunity individually or as mixtures in the vaccine. Although this approach may work for small viruses encoding several antigens, it is not practical for large viruses like smallpox or bacteria which encode 100s or thousands of antigens. In response to this challenge, our laboratory has developed PCR Express to rapidly generate plasmids that can be expressed in an in vitro transcription/translation system. The resulting proteins are spotted onto microarray chips, and the chips are used to monitor serum antibody titers from vaccinated or infected, humans and animals. This analysis can comprehensively scan entire microorganism proteomes and identify those antigens that are recognized by the immune system following vaccination or infection. This empirical data set can be combined with bioinformatics approaches to select superior vaccine antigen targets. Our experience is that this empirical analysis leads to the identification of a manageable subset of antigens, and our expectation is that for each infectious agent several antigens will be administered together in the vaccine. We are currently supported to use this approach to identify the vaccine antigen sets for smallpox, F. tularensis, P. falciparum (malaria), and B. pseudomallei, and we are accumulating proof that the approach is effective.
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Liang, X., et al., Transcriptionally Active Polymerase Chain Reaction (TAP). High Throughput Gene EXpression Using Genome Sequence Data. J Biol Chem, 2002. 277(5): p. 3593-8.
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Felgner, P.L. and G. Rhodes, Gene therapeutics. Nature, 1991. 349(6307): p. 351-2.
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Felgner, P.L., Nonviral strategies for gene therapy. Sci Am, 1997. 276(6): p. 102-6.
Ulmer, J.B., et al., Heterologous protection against influenza by injection of DNA encoding a viral protein [see comments]. Science, 1993. 259(5102): p. 1745-1749.
Wolff, J.A., et al., Direct gene transfer into mouse muscle in vivo. Science, 1990. 247(4949 Pt 1): p. 1465-1468.
Felgner, P.L., et al., Lipofection: a highly efficient, lipid-mediated DNA-transfection procedure. Proc Natl Acad Sci U S A, 1987. 84(21): p. 7413-7.
Vaccina Proteome Affinity Reagents From Phage Display $1,000,000, 2 yrs
Scanning the F. tularensis Proteome for Vaccine Antigens $2,800,000 4 yrs
Scanning B. pseudomallei proteome for vaccine antigens $5,804,000, 5 yrs
Chair, NIH Study Section: 'Gene and Drug Delivery Systems'
Syntex Research 1982—1988
Chief Scientific Officer, Founder
Vical Incorporated 1988—1998
Director, Protein Microarray Laboratory, Infectious Disease