203. Salt Conditions during Polyplex Formation Significantly Alters Transfection

Abstract

Synthetic histidine-lysine (HK) peptides, comprised of histidine and lysine, have been shown to be effective carriers of nucleic acids. We compared the transfection ability of one these HK peptides, H2K4b, in complex with plasmids prepared under three conditions: 1) water, 2) water and then Opti-MEM (or 150 mM NaCL), or 3) Opti-MEM (or 150 mM NaCL). When H2K4b in complex with a luciferase-containing plasmid complex was first formed in water before Opti-MEM was added, luciferase activity increased significantly in most cell lines compared to when the polyplex was formed in Opti-MEM; in two cell lines, this sequential change in salt (e.g., water/ then Opti-MEM) increased transfection activity by as much as five-fold. Furthermore, H2K4b: luciferase-containing plasmid complex formed only in water resulted in a marked decrease in luciferase activity compared to when the complex was formed in Opti-MEM or water/then Opti-MEM. Similarly, sequential changes in salt conditions (e.g., water/Opti-MEM) during the preparation of the H2K4b: β-galactosidase plasmid complex augmented galactosidase expression. H2K4b polyplexes, formed first in water and then in Opti-MEM were intermediate in size (761 ± 292 nm) compared to polyplexes formed in water (81.1 ± 25.6 nm) or Opti-MEM (1729 ± 559nm); atomic force microscopy confirmed these findings. Nevertheless, the surface charge on the H2K4b/plasmid DNA complex changed minimally despite different conditions in which the polyplex was formed. Variation in the salt conditions during formation of the HK:DNA complexes may increasetransfection due to the size and DNA release from the polyplexes. Synthetic histidine-lysine (HK) peptides, comprised of histidine and lysine, have been shown to be effective carriers of nucleic acids. We compared the transfection ability of one these HK peptides, H2K4b, in complex with plasmids prepared under three conditions: 1) water, 2) water and then Opti-MEM (or 150 mM NaCL), or 3) Opti-MEM (or 150 mM NaCL). When H2K4b in complex with a luciferase-containing plasmid complex was first formed in water before Opti-MEM was added, luciferase activity increased significantly in most cell lines compared to when the polyplex was formed in Opti-MEM; in two cell lines, this sequential change in salt (e.g., water/ then Opti-MEM) increased transfection activity by as much as five-fold. Furthermore, H2K4b: luciferase-containing plasmid complex formed only in water resulted in a marked decrease in luciferase activity compared to when the complex was formed in Opti-MEM or water/then Opti-MEM. Similarly, sequential changes in salt conditions (e.g., water/Opti-MEM) during the preparation of the H2K4b: β-galactosidase plasmid complex augmented galactosidase expression. H2K4b polyplexes, formed first in water and then in Opti-MEM were intermediate in size (761 ± 292 nm) compared to polyplexes formed in water (81.1 ± 25.6 nm) or Opti-MEM (1729 ± 559nm); atomic force microscopy confirmed these findings. Nevertheless, the surface charge on the H2K4b/plasmid DNA complex changed minimally despite different conditions in which the polyplex was formed. Variation in the salt conditions during formation of the HK:DNA complexes may increasetransfection due to the size and DNA release from the polyplexes.