Lifetime-Engineered Ruby Nanoparticles (Tau-Rubies) for Multiplexed Imaging of μ-Opioid Receptors

Abstract

To address the growing demand for simultaneous imaging of multiple biomarkers in highly scattering media such as organotypic cell cultures, we introduce a new type of photoluminescent nanomaterial termed “tau-ruby” composed of ruby nanocrystals (Al2O3:Cr3+) with tunable emission lifetime. The lifetime tuning range from 2.4 to 3.2 ms was achieved by varying the Cr3+ dopant concentration from 0.8% to 0.2%, affording facile implementation of background-free detection. We developed inexpensive scalable production of tau-ruby characterized by bright emission, narrow spectrum (693 ± 2 nm), and virtually unlimited photostability upon excitation with affordable excitation/detection sources, noncytotoxic and insensitive to microenvironmental fluctuations. By functionalizing the surface of tau-rubies with targeting antibodies, we obtained different biomarkers suitable for multiplexed lifetime imaging. As a proof of principle, three tau-ruby bioprobes, characterized by three mean lifetimes, were deployed to label three μ-opioid receptor species expressed on transfected cancer cells, each fused to a unique epitope, so that three types of cells were lifetime-encoded. Robust decoding of photoluminescent signals that report on each cell type was achieved by using a home-built lifetime imaging system and resulted in high-contrast multiplexed lifetime imaging of the cells.

Synthesis of Ultrafine Aluminum-Rich Corundum Nanocrystals

“…Excitation of the solid solution nanocrystals as a powder using a 532 nm laser produced a sharp emission line at 695 nm characteristic of substitutional Cr 3+ ions in an Al-rich corundum lattice (Figure 7). 7,37,38 Two shoulders were observed on either side of the sharp emission (Figure 7a). These shoulders, between 670 and 680 nm and between 705 and 715 nm, have been attributed to coupled pairs of Cr 3+ ions.…”

Ruby Nanoflakes (Rubyene) for Efficient 2D Förster Resonance Energy Transfer: Implications for Engineered Emitters in Multiplexed Imaging

“…much greater than the Rn thickness (Figure S6). Rn coating with SiO 2 changed the zeta potential to negative ∼ −30 mV 30 and hindered ICG attaching to nanoparticles surface (right panel in Figure 6e). The FRET efficiency, η, is the fraction of the donor excitation energy transferred to the acceptors.…”

Nanometer-Thick Carbon Coatings with Covalent Chemical Functionalization of Metal Oxide Nanoparticles for Environmental and Biological Applications

“…The small size and wide range of optical and magnetic properties make metal oxide nanomaterials particularly attractive candidates for use as analytical probes in chemistry and biology. , For example, a nanoruby (Al 2 O 3 :Cr 3+ ) has a broad absorption spectrum coupled with a single very sharp emission line and quantum efficiency exceeding 80%; these properties make it well suited to applications based on imaging and tracking in complex biological systems. − Iron oxide nanoparticles exhibit useful magnetic properties, facilitating fundamental studies of protein adsorption − and potential use in biomedical diagnostics and therapy and diagnostics. − Recent studies have shown that metal oxides can also be highly effective micronutrient delivery vehicles for nano-enhanced agriculture. − Control over the particle surface could increase their efficiency and effectiveness, targeting specific parts of plants or bodies for delivery of nutrients or drugs.…”