title: The establishment of an up-scaled micro-mixer method allows the standardized and reproducible preparation of well-defined plasmid/LPEI polyplexes.
abstract: Polyplexes based on linear polyethylenimine (LPEI) and plasmid DNA are known as efficient non-viral gene delivery systems. However, the requirement for freshly prepared complexes prior to administration due to their instability in aqueous suspension poses the risk of batch-to-batch variations. Therefore, the aim of the study was the establishment of a reproducible and up-scalable method for the preparation of well-defined polyplexes. Polyplexes consisting of pCMVLuc plasmid and 22 kDa linear polyethylenimine (LPEI) were prepared by classical pipetting or with a micro-mixer method using different mixing speeds and plasmid DNA concentrations (20-400 μg/mL). The z-average diameter of the polyplexes was measured by dynamic light scattering. Metabolic activity and transfection efficiency was evaluated on murine neuroblastoma cells after transfection with polyplexes. When varying mixing speeds of the micro-mixer, polyplex size (59-197 nm) and polydispersity index (0.05-0.19) could be directly controlled. The z-average diameter (65-170 nm) and polydispersity index (0.05-0.22) of the polyplexes increased with increasing plasmid DNA concentration (20-400 μg/mL). The established up-scaled micro-mixer method allows the standardized and reproducible preparation of well-defined, transfection-competent plasmid/LPEI polyplexes with high reproducibility.
authors.affiliation: Department of Pharmacy, Pharmaceutical Technology and Biopharmaceutics, Ludwig-Maximilians-University, Munich, Germany. julia.kasper@cup.uni-muenchen.de
authors.forename: Julia Christina
authors.initials: JC
authors.lastname: Kasper
authors.forename: David
authors.initials: D
authors.lastname: Schaffert
authors.forename: Manfred
authors.initials: M
authors.lastname: Ogris
authors.forename: Ernst
authors.initials: E
authors.lastname: Wagner
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Friess
mesh_terms: D000818:Animals; D020811:Biomedical Technology / Q000295:instrumentation / Q000379:methods; D045744:Cell Line, Tumor; D017930:Genes, Reporter; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors; D051379:Mice; D009447:Neuroblastoma / Q000235:genetics / Q000378:metabolism; D009994:Osmolar Concentration; D010316:Particle Size; D010957:Plasmids / Q000737:chemistry / Q000633:toxicity; D011094:Polyethyleneimine / Q000737:chemistry / Q000633:toxicity; D011786:Quality Control; D018675:Toxicity Tests; D014162:Transfection
chemical_list: D011094:Polyethyleneimine