Purpose: The main purpose of this work is the development and characterization of carbon nanotubes (CNTs) and polymeric nanoparticles (PMMA) as intracellular delivery tools of molecular beacons (MB) for the detection and localization of Survivin mRNA. Methods: Survivin molecular beacon was designed and covalently immobilized on CNT and PMMA nanoparticles. The choice of Sulfosuccinimidyl 6-(3'-[2-pyridyldithio]-propionamido)hexanoate (Sulfo-LC-SPDP) as cross-linker, allowed the possibility to release the MB once in the intracellular reducing environment. A full optical characterization of the MB was performed measuring its fluorescence at different concentrations of the specific target sequence and in presence of a random sequence. Results: An in-vitro characterization of Survivin MB was performed, 100-nM MB was examined after incubation with different target concentrations in different buffers (Tris and DMEM) with an incubation time of 3 h. An increase of the MBs fluorescence was recorded by increasing the target concentration with an experimental detection limit of 8 nM. Calibration curves were then obtained for MB immobilized both onto CNT and PMMA nanoparticles. Conclusion: Modification and characterization of CNT and PMMA nanoparticles as potential intracellular cargos for the transport of MB for the detection and localization of a specific mRNA were performed. In particular, the attention was focused on the mRNA of survivin, a protein highly expressed in most types of cancer. Acknoledgements: This work was supported by the flagship national project NANOMAX – project ENCODER.
PMMA nanopaticles and carbon nanotubes as intracellular carriers of molecular beacons for mRNA sensig and imaging
CARPI, SARA;
2013-01-01
Abstract
Purpose: The main purpose of this work is the development and characterization of carbon nanotubes (CNTs) and polymeric nanoparticles (PMMA) as intracellular delivery tools of molecular beacons (MB) for the detection and localization of Survivin mRNA. Methods: Survivin molecular beacon was designed and covalently immobilized on CNT and PMMA nanoparticles. The choice of Sulfosuccinimidyl 6-(3'-[2-pyridyldithio]-propionamido)hexanoate (Sulfo-LC-SPDP) as cross-linker, allowed the possibility to release the MB once in the intracellular reducing environment. A full optical characterization of the MB was performed measuring its fluorescence at different concentrations of the specific target sequence and in presence of a random sequence. Results: An in-vitro characterization of Survivin MB was performed, 100-nM MB was examined after incubation with different target concentrations in different buffers (Tris and DMEM) with an incubation time of 3 h. An increase of the MBs fluorescence was recorded by increasing the target concentration with an experimental detection limit of 8 nM. Calibration curves were then obtained for MB immobilized both onto CNT and PMMA nanoparticles. Conclusion: Modification and characterization of CNT and PMMA nanoparticles as potential intracellular cargos for the transport of MB for the detection and localization of a specific mRNA were performed. In particular, the attention was focused on the mRNA of survivin, a protein highly expressed in most types of cancer. Acknoledgements: This work was supported by the flagship national project NANOMAX – project ENCODER.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
