RESUMEN
A new approach for microsphere-mediated delivery of plasmid DNA has been developed and successfully evaluated. Basic molecular biology techniques were used to linearize and functionalize plasmid DNA by aminomodification, enabling efficient conjugation to carboxy-functionalized microspheres. A T cell hybridoma line was successfully transfected as determined by the efficient expression of a biologically relevant YFP fusion protein. Moreover, our data identified microsphere-mediated delivery of plasmid DNA as a noninvasive, nontoxic, and efficient gene delivery method with the potential to be applied to transfection-resistant, nondividing primary cells, including naïve T cells.
Asunto(s)
ADN/administración & dosificación , Microesferas , Plásmidos/administración & dosificación , Transfección , Animales , Humanos , Hibridomas , Linfocitos T/metabolismoRESUMEN
A novel multifunctional probe to monitor intracellular enzymatic activity in living cells is successfully developed. Their use as accurate intracellular sensors by conjugation of an internal control (that gives an extra feature to both evaluate cellular-uptake efficiency and track probes over time) is reported. In particular, a specific application of these multifunctional microspheres as sensors of caspase-3/7 to monitor apoptosis by flow cytometry is described. The preparation of these devices together with a kinetic study towards caspase-3 and caspase-7 and their evaluation as flow cytometry probe in apoptotic living cells are reported.