Protein−ligand interactions are frequently screened using nuclear magnetic resonance (NMR) spectroscopy. The dissociation constant (KD) of a ligand of interest can be determined via a spin-spin relaxation measurement of a reporter ligand, in a single scan when using hyperpolarization by means of dissolution dynamic nuclear polarization (D-DNP). Despite nearly instantaneous signal acquisition, a limitation of D-DNP for the screening of protein−ligand interactions is the required polarization time on the order of tens of minutes. Here, we introduce a multiplexed NMR experiment, where a single hyperpolarized ligand sample is rapidly mixed with protein injected into two flow cells. NMR detection is achieved simultaneously on both channels, resulting in a chemical shift resolved spin relaxation measurement. Spectral resolution allows the use of reference compounds for accurate quantification of concentrations. Simultaneous use of two concentration ratios between protein and ligand broadens the range of KD that is accurately measurable in a single experiment to at least an order of magnitude. In a comparison of inhibitors for the protein trypsin, the average KD values of benzamidine and benzylamine were found to be 12.6±1.4 μM and 207±22.3 μM from three measurements, based on KD = 142 μM assumed known for the reporter ligand 4-(trifluoromethyl)benzene-1-carboximidamide. Typical confidence ranges at 95% evaluated for single experiments were (8.3 μM, 20 μM) and (151 μM, 328 μM). The multiplexed detection of two or more hyperpolarized samples increases throughput of D-DNP by the same factor, improving the applicability to most multi-point measurements that would traditionally be achieved using titrations.

Despite much progress in functionalized gold nanomaterial (GNMs), chemiluminescent (CL) functionalized GNMs with high CL efficiency are far from fully developed. In this work, we report a general strategy for the synthesis of gold nanoparticles (GNPs) bifunctionalized by CL reagent and catalyst metal complexes (BF-GNPs) by taking N-(aminobutyl)-N-(ethylisoluminol) (ABEI) as a model of CL reagents. The complexes of 2-[bis[2-[carboxymethyl-[2-oxo-2-(2-sulfanylethylamino)ethyl]amino]ethyl]amino]acetic acid (DTDTPA) with various metal ions, including Co(2+), Cu(2+), Pb(2+), Ni(2+), Hg(2+), Cr(3+), Eu(3+), La(3+), Gd(3+), Sm(3+), Er(3+), Dy(3+), Ce(4+), and Ce(3+), were grafted on the surface of ABEI functionalized GNPs (ABEI-GNPs) to form a series of BF-GNPs. These BF-GNPs exhibited excellent CL activity. In particular, the CL intensity of DTDTPA/Co(2+)-ABEI-GNPs was over 3 orders of magnitude higher than ABEI-GNPs. This work demonstrates for the first time that metal complexes grafted on the surface of GNPs have unique catalytic activity on the CL reaction, superior to that in the liquid phase. Such BF-GNPs may find future applications in bioassays, microchips, and molecular/cellular imaging.

Non-pharmaceutical interventions (NPIs) and vaccination are two fundamental approaches for mitigating the coronavirus disease 2019 (COVID-19) pandemic. However, the real-world impact of NPIs versus vaccination, or a combination of both, on COVID-19 remains uncertain. To address this, we built a Bayesian inference model to assess the changing effect of NPIs and vaccination on reducing COVID-19 transmission, based on a large-scale dataset including epidemiological parameters, virus variants, vaccines, and climate factors in Europe from August 2020 to October 2021. We found that (1) the combined effect of NPIs and vaccination resulted in a 53% (95% confidence interval: 42–62%) reduction in reproduction number by October 2021, whereas NPIs and vaccination reduced the transmission by 35% and 38%, respectively; (2) compared with vaccination, the change of NPI effect was less sensitive to emerging variants; (3) the relative effect of NPIs declined 12% from May 2021 due to a lower stringency and the introduction of vaccination strategies. Our results demonstrate that NPIs were complementary to vaccination in an effort to reduce COVID-19 transmission, and the relaxation of NPIs might depend on vaccination rates, control targets, and vaccine effectiveness concerning extant and emerging variants.