3st Report IAEA: PREPARATION OF ALBUMIN AND GOLD/ALBUMIN NANO-PARTICLES BY RADIATION-INDUCED CROSS-LINKING

GRASSELLI, MARIANO; ALONSO, S. DEL V.; ACHILLI, ESTEFANIA; FLORES, CONSTANZA Y.

2014-04-01/2018-05-01

Nanotecnologia

Tecnologia sanitaria y curativa-Varios

During this period a further characterization of multilayer coated (core/shell) gold/Albumin-NPs have been prepared by radiation-induced crosslinking. NPs in the range of 60 to 80 nm have been prepared and characterized by using UV-Vis, TEM and AFM in liquid state. In addition FT-IR spectroscopy, Zeta potential and biorecognition experiments were performed. A dot-blot analysis onto nitrocellulose membrane shows specific antibody recognition of the nanoparticle. Nanoparticle uptake experiments were run onto PC3 cell line. Nanoparticle stability has been studied by SDS-PAGE electrophoresis. Au/Albumin-NPs show that the Albumin increase the molecular weight and the Au-NPs have a protein coating; however a huge amount of Albumin is not covalently attached to the NP structure. Decoration of gold/Albumin-NPs with DOTA-Bombesin derivative (DOTA-BBN) has been achieved by using hetero-bifunctional linkers. In a first step, DOTA-BBN-linker was prepared and characterized by ESI-Mass spectroscopy. After purification this compound was linked to the NPs covalently to the NPs. Different techniques have been studied to quantify the peptide onto the nanoconstruct, such as Dissociation-Enhanced Lanthanide Fluoro-Immunoassay, transchelation with Arsenazo III-Pb(II) and Inductively Coupled Plasma Atomic Emission Spectroscopy (ICP-AES) using Sr(II) as testing ion. Only ICP-AES technique was confident to measure the peptide content which was in the order of 400 DOTA-BBN per NP. However this technique requires a huge amount of sample to determine the ligand. DOTA-BBN-NPs has been used for cell uptake experiment under controlled conditions. It was found an important cytoplasm uptake compared to the same nanoparticle without the specific ligand using very low NP concentration. In addition NP samples were sent to Italy-Padova and Pakistan for further biological analysis.