DNA chips: state-of-the art.
Wolpert Polymers, Inc., Richmond, VA 23225-4636, USA. email@example.com
The technology and applications of microarrays of immobilized DNA or oligonucleotides are reviewed. DNA arrays are fabricated by high-speed robotics on glass or nylon substrates, for which labeled probes are used to determine complementary binding allowing massively parallel gene expression and gene discovery studies. Oligonucleotide microarrays are fabricated either by in situ light-directed combinational synthesis or by conventional synthesis followed by immobilization on glass substrates. Sample DNA is amplified by the polymerase chain reaction (PCR), and a fluorescent label is inserted and hybridized to the microarray. This technology has been successfully applied to the simultaneous expression of many thousands of genes and to large-scale gene discovery, as well as polymorphism screening and mapping of genomic DNA clones.
Induction of primary carcinoembryonic antigen (CEA)-specific cytotoxic T lymphocytes in vitro using human dendritic cells transfected with RNA.
Nair SK. Boczkowski D. Morse M. Cumming RI. Lyerly HK. Gilboa E.
Center for Genetic and Cellular Therapies, Department of Surgery, Duke University Medical Center, Durham, NC 27710, USA.
Dendritic cells (DC) generated from the peripheral blood mononuclear cells of healthy individuals or from cancer patients transfected with carcinoembryonic antigen (CEA) mRNA stimulate a potent CD8+ cytotoxic T lymphocyte (CTL) response in vitro. DCs are effectively sensitized with RNA in the absence of reagents commonly used to facilitate mammalian cell transfection. RNA encoding a chimeric CEA/LAMP-1 lysosomal targeting signal enhances the induction of CEA-specific CD4+ T cells, providing a strategy to induce T-help that may be necessary to generate and/or maintain an optimal CD8+ CTL response in vivo. CEA RNA-transfected DCs also serve as effective targets in cytotoxicity assays, thus providing a general method for inducing, as well as measuring, CEA-specific CTL responses across a broad spectrum of HLA haplotypes.