Emission Onset Time-Adjustable Chemiluminescent Gold Nanoparticles with Ultrastrong Emission for Smartphone-Based Immunoassay of Severe Acute Respiratory Syndrome Coronavirus 2 Antigen.

Recently, our group reported a chemical timer approach to manipulate the onset time of chemiluminescence (CL) emission. However, it is still in the proof-of-concept stage, and its analytical applications have not been explored yet. Nanomaterials have merits of good catalytic effect, large specific surface area, good biocompatibility, and ease of self-assembly, which are ideal for constructing analytical-interfaces for bioassays. Herein, an emission onset time-adjustable chemiluminescent L012-Au/Mn2+ was synthesized for the first time by modifying Mn2+ on the surface of L012-protected gold nanoparticle. By using H2O2 and NaHCO3 as coreactants, L012-Au/Mn2+ could not only generate an ultrastrong and long-time CL emission but also its CL emission onset time could be adjusted by the addition of thiourea, which could effectively eliminate interference from the addition of coreactants, shorten the exposure time, reduce the detection background, and finally achieve high sensitivity CL imaging analysis. On this basis, a label-free CL immunoassay was constructed with a smartphone-based imaging system for high-throughput and sensitive determination of severe acute respiratory syndrome coronavirus 2 nucleocapsid (N) protein. The CL image of the immunoassay with different concentrations of N proteins was captured in one photograph 100 s after the injection of H2O2 with a short exposure time of 0.5 s. The immunoassay showed good linearity over the concentration range of 1 pg/mL to 10 ng/mL with a detection limit of 0.13 pg/mL, which was much lower than the reported CCD imaging detection method. In addition, it showed good selectivity and stability and was successfully applied in serum samples from healthy individuals and COVID-19 rehabilitation patients.

A chemical timer approach to delayed chemiluminescence.

Although the onset time of chemical reactions can be manipulated by mechanical, electrical, and optical methods, its chemical control remains highly challenging. Herein, we report a chemical timer approach for manipulating the emission onset time of chemiluminescence (CL) reactions. A mixture of Mn2+, NaHCO3, and a luminol analog with H2O2 produced reactive oxygen species (ROS) radicals and other superoxo species (superoxide containing complex) with high efficiency, accompanied by strong and immediate CL emission. Surprisingly, the addition of thiourea postponed CL emission in a concentration-dependent manner. The delay was attributed to a slow-generation-scavenging mechanism, which was found to be generally applicable not only to various types of CL reagents and ROS radical scavengers but also to popular chromogenic reactions. The precise regulation of CL kinetics was further utilized in dynamic chemical coding with improved coding density and security. This approach provides a powerful platform for engineering chemical reaction kinetics using chemical timers, which is of application potential in bioassays, biosensors, CL microscopic imaging, microchips, array chips, and informatics.

Imaging the oxygen wave with a single bioluminescent bacterium.

We developed a capability of a monolayer of bioluminescent (BL) bacteria for spatiotemporally visualizing the heterogeneous distribution and dynamic evolution of interfacial oxygen concentration, resulting in the discovery of spontaneous and stochastic oxygen waves at the interface between the substrate and an undisturbed, apparently still solution. Wild type bacteria, P. phosphoreum, spontaneously emit light during the native metabolism processes, i.e., bioluminescence. The emission intensity is sensitively regulated by oxygen concentration. By taking the electrolysis of water as a model, it was demonstrated that time-lapsed BL imaging of a bacterial monolayer allowed for visualizing the dynamic distribution of oxygen. The results were quantitatively understood with a physical model involving the diffusion equation and Michaelis-Menten equation. Unexpectedly, further study uncovered a spontaneous and stochastic oxygen wave in a standard well of a microtiter plate, which was subsequently attributed to the inevitable micro-convections induced by inhomogeneous evaporation and thermal fluctuation. Because of the wide application of microtiter plates, this study sheds new light to better understand the apparent heterogeneity in cell-culture and bio-assays.

Rational design of functional materials guided by single particle chemiluminescence imaging.

Chemiluminescence (CL) functionalized materials have found tremendous value in developing CL assays for clinical assays and point-of-care tests. To date, the design and optimization of these materials have mainly relied on conventional trial-and-error procedures in which the ensemble performance is evaluated using conditional experiments. Here we have built an optical microscope to acquire the CL emission from single magnetic-polymer hybrid microbeads functionalized with luminol analogues, and to access the CL kinetics of each individual particle. It was incidentally found that a minor subpopulation of microbeads exhibited intense and delayed CL emission while the majority showed transient and weak emission. Structural characterization of the very same individual particles uncovered that the amorphous multi-core microstructures were responsible for the enhanced encapsulation efficiency and optimized CL reaction kinetics. Guided by this knowledge stemming from single particle CL imaging, the synthesis procedure was rationally optimized to enrich the portion of microbeads with better CL performance, which was validated by both single particle imaging and the significantly improved analytical performance at the ensemble level. The present work not only demonstrates the CL imaging and CL kinetics curve of single microbeads for the first time, but also sets a clear example showing the capability of single particle studies to investigate the structure-activity relationship in a bottom-up manner and to help the rational design of ensemble materials with improved performance.

Potential-Resolved Multicolor Electrochemiluminescence of N-(4-Aminobutyl)-N-ethylisoluminol/tetra(4-carboxyphenyl)porphyrin/TiO2 Nanoluminophores.

Most electrochemiluminescence (ECL) studies involve single luminophore with a unique emission process, which severely limits its applications. Recently, multicolor ECL has attracted considerable interests. Herein, we report a novel nanoluminophore prepared by coating 5,10,15,20-tetrakis(4-carboxyphenyl)-porphyrin (TCPP) and N-(4-aminobutyl)-N-ethylisoluminol (ABEI) on the surface of TiO2 nanoparticles (TiO2-TCPP-ABEI), which exhibited unique potential-resolved multicolor ECL emissions using H2O2 and K2S2O8 as coreactants in an aqueous solution. Three ECL peaks, ECL-1 at 458 nm, ECL-2 at 686 nm, and ECL-3 at 529 nm, were obtained with peak potentials of 1.05, -1.65, and -1.85 V, which were attributed to the ECL emission of ABEI, TCPP, and TiO2 moiety of the nanoluminophores, respectively. Potential-resolved multicolor ECL from a nanoluminophore was observed for the first time in an aqueous solution. It opens a new research area of multicolor ECL of nanoluminophores, which is of great importance in ECL field from fundamental studies to practical applications.