Highly fluorescent conjugated polymer dots were developed for demanding applications such as fluorescence imaging in live cells. These nanoparticles exhibit small particle diameters, extraordinary fluorescence brightness, and excellent photostability. Single particle fluorescence imaging and kinetic studies indicate much higher emission rates (∼108 s-1) and little or no blinking of the nanoparticles as compared to typical results for single dye molecules and quantum dots. Analysis of single particle photobleaching trajectories reveals excellent photostability — as many as 109 or more photons emitted per nanoparticle prior to irreversible photobleaching. The superior figures of merit of these new fluorescent probes, together with the demonstration of cellular imaging, indicate their enormous potential for demanding fluorescence-based imaging and sensing applications such as high speed super-resolution single molecule/particle tracking and highly sensitive assays.

Semiconducting polymer dots (Pdots) represent a new class of ultrabright fluorescent probes for biological imaging. They exhibit several important characteristics for experimentally demanding in vitro and in vivo fluorescence studies, such as their high brightness, fast emission rate, excellent photostability, non-blinking, and non-toxic feature. However, controlling the surface chemistry and bioconjugation of Pdots has been a challenging problem that prevented their widespread applications in biological studies. Here, we report a facile yet powerful conjugation method that overcomes this challenge. Our strategy for Pdot functionalization is based on entrapping heterogeneous polymer chains into a single dot, driven by hydrophobic interactions during nanoparticle formation. A small amount of amphiphilic polymer bearing functional groups is cocondensed with the majority of semiconducting polymers to modify and functionalize the nanoparticle surface for subsequent covalent conjugation to biomolecules, such as streptavidin and immunoglobulin G (IgG). The Pdot bioconjugates can effectively and specifically label cellular targets, such as cell surface marker in human breast cancer cells, without any detectable nonspecific binding. Single-particle imaging, cellular imaging, and flow cytometry experiments indicate a much higher fluorescence brightness of Pdots compared to those of Alexa dye and quantum dot probes. The successful bioconjugation of these ultrabright nanoparticles presents a novel opportunity to apply versatile semiconducting polymers to various fluorescence measurements in modern biology and biomedicine.

A facile method has been developed to prepare aqueous dispersions of encapsulated conjugated polymer nanoparticles exhibiting high fluorescence brightness. Salient features of the nanoparticles include their small diameter and spherical morphology. Encapsulation of the nanoparticles with a silica shell reduces the rate of photooxidation and allows facile attachment of functional groups for subsequent bioconjugation and nanoparticle assembly. Functionalization of the nanoparticle with amine groups followed by the addition of Au nanoparticles resulted in the formation of nanoparticle assemblies, as evidenced by the efficient quenching of the conjugated polymer fluorescence by the Au nanoparticles.

We report on the fluorescence properties and the combined effects of energy diffusion and energy transfer in polyfluorene nanoparticles doped with a variety of fluorescent dyes. As the doping host, polyfluorene possesses extraordinary "light harvesting" ability, resulting in higher per-particle brightness as compared to dye-loaded silica nanoparticles of similar dimensions. Both the steady-state fluorescence spectra and time-resolved fluorescence measurements indicate highly efficient energy transfer from the host polymer to the acceptor dye molecules. A model that takes into account the combined effects of energy diffusion, Förster transfer, and particle size was developed. Comparisons of experimental data to the model results elucidate the importance of particle size and energy diffusion within the polymer in determining the optical properties of the doped conjugated polymer nanoparticles. Fluorescence quantum yields of ~40% and peak extinction coefficients of 1.5 × 10(9) M(-1)cm(-1) were determined for aqueous suspensions of ~30 nm diameter polymer nanoparticles doped with perylene or coumarin 6 (2 wt %). Photobleaching experiments indicate that energy transfer phenomena strongly influence the photostability of these dye-doped nanoparticles. Significant features of these nanoparticles include the high brightness, highly red-shifted emission spectrum, and excellent photostability, which are promising for biological labeling and sensing applications. In addition, the nanoparticles are a useful model system for studying energy transfer in dense, nanostructured, multichromophoric systems.