title: Efficiency of cellular delivery of antisense peptide nucleic acid by electroporation depends on charge and electroporation geometry.
abstract: Electroporation is potentially a very powerful technique for both in vitro cellular and in vivo drug delivery, particularly relating to oligonucleotides and their analogs for genetic therapy. Using a sensitive and quantitative HeLa cell luciferase RNA interference mRNA splice correction assay with a functional luciferase readout, we demonstrate that parameters such as peptide nucleic acid (PNA) charge and the method of electroporation have dramatic influence on the efficiency of productive delivery. In a suspended cell electroporation system (cuvettes), a positively charged PNA (+8) was most efficiently transferred, whereas charge neutral PNA was more effective in a microtiter plate electrotransfer system for monolayer cells. Surprisingly, a negatively charged (-23) PNA did not show appreciable activity in either system. Findings from the functional assay were corroborated by pulse parameter variations, polymerase chain reaction, and confocal microscopy. In conclusion, we have found that the charge of PNA and electroporation system combination greatly influences the transfer efficiency, thereby illustrating the complexity of the electroporation mechanism.
authors.affiliation: Department of Oncology, Copenhagen University Hospital Herlev, Herlev, Denmark.
authors.forename: Mette
authors.initials: M
authors.lastname: Joergensen
authors.forename: Birgit
authors.initials: B
authors.lastname: Agerholm-Larsen
authors.forename: Peter E
authors.initials: PE
authors.lastname: Nielsen
authors.forename: Julie
authors.initials: J
authors.lastname: Gehl
mesh_terms: D016375:Antisense Elements (Genetics) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002470:Cell Survival; D016503:Drug Delivery Systems / Q000295:instrumentation / Q000379:methods*; D018274:Electroporation / Q000379:methods*; D004867:Equipment Design; D005260:Female; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000032:analysis; D018613:Microscopy, Confocal; D020135:Peptide Nucleic Acids / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012326:RNA Splicing; D055672:Static Electricity; D014162:Transfection / Q000379:methods*
chemical_list: D016375:Antisense Elements (Genetics); D020135:Peptide Nucleic Acids; D008156:Luciferases