Identification of polystyrene nanoparticle penetration across intact skin barrier as rare event at sites of focal particle aggregations

Abstract

The question whether nanoparticles can cross the skin barrier is highly debated. Even in intact skin rare events of deeper penetration have been reported, but technical limitations and possible artifacts require careful interpretation. In this study, horizontal scanning by 2-photon microscopy (2 PM) of full-thickness human skin samples placed in a lateral position yielded highly informative images for skin penetration studies of fluorescently tagged nanoparticles. Scanning of large fields of view allowed for detailed information on interfollicular and follicular penetration in tissue blocks without damaging the sample. Images in histomorphological correlation showed that 2P-excited fluorescence signals of fluorescently tagged 20 and 200 nm polystyrene nanoparticles preferentially accumulated in the stratum corneum (SC) and in the upper part of vellus hair follicles (HFs). Rare events of deeper penetration in the SC and in the infundibulum of vellus HFs were observed at sites of high focal particle aggregations. Wide-field 2 PM allows for imaging of nanoparticle penetration in large tissue blocks, whereas total internal reflection microscopy (TIRFM) enables selective detection of individual nanoparticles as well as clusters of nanoparticles in the SC and within the epidermal layer directly beneath the SC, thus confirming barrier crossing with high sensitivity.

Keywords: 2-photon microscopy; fluorescently tagged polystyrene nanoparticles; single-particle tracking; skin penetration; total internal reflection fluorescence microscopy.