Imaging the uptake of gold nanoshells in live cells using plasmon resonance enhanced four wave mixing microscopy

This paper was published in Optics Express and is made available as an electronic reprint with the permission of OSA. The paper can be found at the following URL on the OSA website: http://dx.doi.org/10.1364/OE.19.017563. Systematic or multiple reproduction or distribution to multiple locations via electronic or other means is prohibited and is subject to penalties under law.Gold nanoshells (GNS) are novel metal nanoparticles exhibiting attractive optical properties which make them highly suitable for biophotonics applications. We present a novel investigation using plasmon-enhanced four wave mixing microscopy combined with coherent anti-Stokes Raman scattering (CARS) microscopy to visualize the distribution of 75 nm radius GNS within live cells. During a laser tolerance study we found that cells containing nanoshells could be exposed to < 2.5 mJ each with no photo-thermally induced necrosis detected, while cell death was linearly proportional to the power over this threshold. The majority of the GNS signal detected was from plasmon-enhanced four wave mixing (FWM) that we detected in the epi-direction with the incident lasers tuned to the silent region of the Raman spectrum. The cellular GNS distribution was visualized by combining the epi-detected signal with forwards-detected CARS at the CH2 resonance. The applicability of this technique to real-world nanoparticle dosing problems was demonstrated in a study of the effect of H2S on nanoshell uptake using two donor molecules, NaHS and GYY4137. As GYY4137 concentration was increased from 10 µM to 1 mM, the nanoshell pixel percentage as a function of cell volume (PPCV) increased from 2.15% to 3.77%. As NaHS concentration was increased over the same range, the nanoshell PPCV decreased from 12.67% to 11.47%. The most important factor affecting uptake in this study was found to be the rate of H2S release, with rapid-release from NaHS resulting in significantly greater uptake

Nano-Biophotonics

Photonic techniques are the methods of choice for probing biological systems, as they are non-invasive, non-ionising, inexpensive, and are ubiquitous. When applied to the treatment and prevention of disease and for pathology in general, biophotonics offers a means to bridge the gap between understanding of molecular structures and their role in physiological functions. There is a wide range of such techniques used in imaging, assaying, bio-sensing, optical diagnosis, each of which has limitations as well as benefits. The experiments outlined in this thesis use nanotechnology to overcome the limitations of resolution, contrast and chemical specificity with photonic techniques in biology. The experimental work outlined in this thesis is divided over three chapters, the first of which is concerned with nanostructured metallic surfaces for use in surface enhanced Raman scattering (SERS) for protein assay applications. This chapter gives details of the methods used to produce and characterise SERS substrates using gold and silver thermally evaporated onto butterfly wing sections, together with the protocols developed for manufacturing biomimetic analogues of these naturally occurring nanostructures. The conjugation system designed to modify the metal surfaces for use in an avidin/biotin model protein binding assay is described, together with an account of the efficacy of the final assay. The results obtained show that such naturally occurring nanostructures, and their biomimetic analogues, are suitable for use as SERS substrates for wet protein binding assays. This work represents a major advance in the field of SERS assay. The experimental chapters describe experiments that use coherent Raman scattering (CRS) methods to probe the interactions between nanoparticles and live cell cultures, as well as provide chemically selective images of tissue samples

BALL SIZE AND WEIGHT EFFECTS ON THROWING KINEMATICS AND KINETICS IN YOUTH BASEBALL ATHLETES

In baseball, youth players play on smaller fields with shorter base path distance, pitching distance, and smaller mounds. Despite this, the baseball itself remains unchanged for youth athletes. This prospective cohort analyzed the kinematics and kinetics of 38 youth baseball pitchers while using modified sized and weighted baseballs. An ANOVA was used to determine statistical significance amongst ball modifications. ANOVA results show significance between the 3oz-5oz baseball with the 3oz baseball decreasing elbow varus torque. This is a preliminary study on the effects of modified baseballs on youth athletes