Measuring Melittin Uptake into Hydrogel Nanoparticles with Near-Infrared Single Nanoparticle Surface Plasmon Resonance Microscopy

This paper describes how changes in the refractive index of single hydrogel nanoparticles (HNPs) detected with near-infrared surface plasmon resonance microscopy (SPRM) can be used to monitor the uptake of therapeutic compounds for potential drug delivery applications. As a first example, SPRM is used to measure the specific uptake of the bioactive peptide melittin into N-isopropylacrylamide (NIPAm)based HNPs. Point diffraction patterns in sequential real-time SPRM differential reflectivity images are counted to create digital adsorption binding curves of single 220 nm HNPs from picomolar nanoparticle solutions onto hydrophobic alkanehiol-modified gold surfaces. For each digital adsorption binding curve, the average single nanoparticle SPRM reflectivity response, (Delta%R-NP), was measured. The value of (Delta%R-NP) increased linearly from 1.04 +/- 0.04 to 2.10 +/- 0.10% when the melittin concentration in the HNP solution varied from zero to 2.5 mu M. No change in the average HNP size in the presence of melittin is observed with dynamic light scattering measurements, and no increase in (Delta%R-NP) is observed in the presence of either FLAG octapeptide or bovine serum albumin. Additional bulk fluorescence measurements of melittin uptake into HNPs are used to estimate that a 1% increase in (Delta%R-NP) observed in SPRM corresponds to the incorporation of approximately 65000 molecules into each 220 nm HNP, corresponding to roughly 4% of its volume. The lowest detected amount of melittin loading into the 220 nm HNPs was an increase in (Delta%R-NP) of 0.15%, corresponding to the absorption of 10000 molecules.