Two-photon fluorescence microscopy is one of the most important recent inventions in biological imaging. This technology enables noninvasive study of biological specimens in three dimensions with submicrometer resolution. Two-photon excitation of fluorophores results from the simultaneous absorption of two photons. This excitation process has a number of unique advantages, such as reduced specimen photodamage and enhanced penetration depth. It also produces higher-contrast images and is a novel method to trigger localized photochemical reactions. Two-photon microscopy continues to find an increasing number of applications in biology and medicine.

The conjugates of gold nanorods and the model drug, fluorescein isothiocyanate (FITC), embedded inside polyelectrolytes (GNRs/FITC@PLE) were synthesized to study the release kinetics of FITC under femtosecond near-infrared (NIR) laser irradiation. The optical and structural properties of GNRs/FITC@PLE conjugates before and after laser treatments were examined using UV-vis spectroscopy, confocal microscopy, and transmission electron microscopy (TEM). The release of FITC from the conjugates was induced by the heat generated from gold nanorods under laser irradiation. The concentration of released FITC was measured as the time of continuous and periodic laser irradiation was varied. Within 5 min of the laser exposure, the release rates of FITC exhibited zero-order and first-order kinetics under continuous and periodic irradiation, respectively. Furthermore, a drug release system was designed based on the conjugates of gold nanorods and the anticancer drug, paclitaxel (PTX), embedded inside polyelectrolytes (GNRs/PTX@PLE). The conjugates were applied for in vitro studies with breast cancer cells. The release of PTX from the conjugates was triggered by NIR laser irradiation, and the inhibition rates of the cells showed strong dependencies on the irradiation modes and time. The results suggested that the multiple releases of PTX from the conjugates can be controlled by laser irradiation within a long period of time. Our system holds great potential for future therapeutic applications on breast cancers.