Control of Oligonucleotide Distribution on the Shell of Thermo-responsive Polymer Nanoparticles

We used Forster resonance energy transfer (FRET) to characterized the adsorption of single strand oligonucleotides (ODN) onto the thermo-responsive shell of polymer nanoparticles with a glassy poly(methyl methacrylate) (PMMA) core and a positively charged shell of poly(N-isopropyl acrylamide) (PNIPAM). The ODNs have 25 thymine units (dT(25)) and are labeled at the 5'-terminus with either rhodamine X (dT(25)-ROX, energy donor) or malachite green (dT(25)-MG, energy acceptor). The kinetics of FRET was analyzed with a distribution model for energy transfer in restricted geometry, which allowed us to obtain the distribution of the ODN strands below and above the volume phase transition temperature of the PNIPAM shell (T-VPT similar to 30 degrees C). Below the T-VPT the ODN molecules adsorb to the expanded particle shell without changing the particle hydrodynamic radius. However, the distribution of ODN is not homogeneous: at low ODN concentrations, we observed preferential adsorption close to the core, while with the increase in ODN content, the distribution extends toward the shell/water interface. At these temperatures the mobility of the adsorbed ODN is very high and their environment is strongly hydrophilic. Above the T-VPT, the shell is collapsed onto the core, with the particle hydrodynamic radius determined by dynamic light scattering (DLS) changing from 109 nm (at 11 degrees C) to 72 nm (at 45 degrees C). The distribution of adsorbed ODNs also decreases in thickness, but to a value (14.7 +/- 0.8 nm) that is larger than the shell thickness obtained by DLS (L-shell = 5 nm). This indicates that the ODN chains are fixed in the collapsed shell but protruding into the water phase, perpendicularly to the core. Time resolved fluorescence anisotropy results further show that the ODN rotation is strongly hindered above T-VPT.