Tunable Magneto-Plasmonic Nanosensor for Sensitive Detection of Foodborne Pathogens.

Frequent outbreaks of food-borne pathogens, particularly E. coli O157:H7, continue to impact human health and the agricultural economy tremendously. The required cell count for this pathogenic strain of E. coli O157:H7 is relatively low and hence it is vital to detect at low colony forming unit (CFU) counts. Available detection methods, though sensitive, fall short in terms of timeliness and often require extensive sample processing. To overcome these limitations, we propose a novel magneto-plasmonic nanosensor (MPnS) by integrating surface plasmon resonance (SPR) properties with spin-spin magnetic relaxation (T2 MR) technology. We engineered MPnS by encapsulating several gold nanoparticles (GNPs) within the polymer-coating of iron oxide nanoparticles (IONPs). First, the polyacrylic acid (PAA)-coated IONPs were synthesized using a solvent precipitation method, then gold chloride solution was used to synthesize GNPs and encapsulate them within the PAA-coatings of IONPs in one step. A magnetic separation technique was used to purify the MPnS and the presence of GNPs within IONPs was characterized using transmission electron microscopy (TEM), energy dispersive x-ray spectroscopy (EDS), and other spectroscopic methods. The synthesized MPnS exhibits MR relaxation properties while possessing amplified optical properties than conventional GNPs. This allows for rapid and ultrasensitive detection of E. coli O157:H7 by SPR, T2 MR, and colorimetric readout. Experiments conducted in simple buffer and in milk as a complex media demonstrated that our MPnS-based assay could detect as low as 10 CFUs of this pathogenic strain of E. coli O157:H7 in minutes with no cross-reactivity. Overall, the formulated MPnS is robust and holds great potential for the ultrasensitive detection of E. coli O157:H7 in a simple and timely fashion. Moreover, this platform is highly customizable and can be used for the detection of other foodborne pathogens.

Rapid Detection and One-Step Differentiation of Cross-Reactivity Between Zika and Dengue Virus Using Functional Magnetic Nanosensors.

Infection with the Zika virus (ZIKV) is an ongoing problem especially as accurate, cost-effective testing remains unresolved. In addition, coinfection occurs with both the Dengue virus (DENV) and ZIKV which leads to cross-reactivity between the flaviviruses and can result in false positives and inaccurate testing. This supports the current need for a simple assay that can detect Zika antibodies sensitively that at the same time can differentiate between cross-reactive antibodies. In this study, we developed customizable magnetic relaxation nanosensors (MRnS) conjugated to various ligands, which included ZIKV (ZENV, zika domain III and NS1) and DENV proteins for specific detection of cross-reactive Zika and Dengue antibodies. Binding interactions between functional MRnS and corresponding targets resulted in the change in spin-spin magnetic relaxation time (T2MR) of water protons, allowing for a rapid and simple means by which these interactions were detected and quantified. Our results show the detection of Zika antibodies within minutes at concentrations as low as 20 nM and display high specificity, reproducibility, and analytical sensitivity. Furthermore, a mixture of functional MRnS was used for the one-step simultaneous detection and differentiation of Zika and Dengue infections. These results demonstrate high specificity and sensitivity for the detection of ZIKV and DENV despite coinfections in both simple and complex media. Overall, our magnetic nanoplatform could be used as a rapid and sensitive assay for the detection of not only Zika- and Dengue-related testing but can be further applied to serological samples of any other pathogens.

A Bimodal Nanosensor for Probing Influenza Fusion Protein Activity Using Magnetic Relaxation.

Viral fusion is a critical step in the entry pathway of enveloped viruses and remains a viable target for antiviral exploration. The current approaches for studying fusion mechanisms include ensemble fusion assays, high-resolution cryo-TEM, and single-molecule fluorescence-based methods. While these methods have provided invaluable insights into the dynamic events underlying fusion processes, they come with their own limitations. These often include extensive data and image analysis in addition to experimental time and technical requirements. This work proposes the use of the spin-spin T2 relaxation technique as a sensitive bioanalytical method for the rapid quantification of interactions between viral fusion proteins and lipids in real time. In this study, new liposome-coated iron oxide nanosensors (LIONs), which mimic as magnetic-labeled host membranes, are reported to detect minute interactions occurring between the membrane and influenza's fusion glycoprotein, hemagglutinin (HA). The influenza fusion protein's interaction with the LION membrane is detected by measuring changes in the sensitive spin-spin T2 magnetic relaxation time using a bench-top NMR instrument. More data is gleaned from including the fluorescent dye DiI into the LION membrane. In addition, the effects of environmental factors on protein-lipid interaction that affect fusion such as pH, time of incubation, trypsin, and cholesterol were also examined. Furthermore, the efficacy and sensitivity of the spin-spin T2 relaxation assay in quantifying similar protein/lipid interactions with more native configurations of HA were demonstrated using virus-like particles (VLPs). Shorter domains derived from HA were used to start a reductionist path to identify the parts of HA responsible for the NMR changes observed. Finally, the known fusion inhibitor Arbidol was employed in our spin-spin T2 relaxation-based fusion assay to demonstrate the application of LIONs in real-time monitoring of this aspect of fusion for evaluation of potential fusion inhibitors.