Multiplexed electrochemical liposomes applied to the detection of nucleic acids for Influenza A, Influenza B and SARS-CoV-2.

Multiplexing is a relevant strategy for biosensors to improve accuracy and decision-making due to the increased amount of simultaneously obtained information. Liposomes offer unique benefits for label-based multiplexing since a variety of different marker molecules can be encapsulated, leading to intrinsic signal amplification and enabling a variety of detection formats. We successfully developed an electrochemical (EC) liposome-based platform technology for the simultaneous detection of at least three analytes by studying parameters to ensure specific and sensitive bioassay performance. Influenza A and B and SARS-CoV-2 sequences served as model system in a standard sandwich hybridization assay. Studies included encapsulants, probe distribution on liposomes and capture beads, assay setup and interferences between liposomes to also ensure a generalization of the platform. Ruthenium hexamine(III), potassium hexacyanoferrate(II) and m-carboxy luminol, when encapsulated separately into a liposome, provided desirable long-term stability of at least 12 months and no cross-signals between liposomes. Through the optimization process, low limits of detections of 1.6 nmol L-1, 125 pmol L-1 and 130 pmol L-1, respectively, were achieved in a multiplexed assay setup, which were similar to singleplex assays. Non-specific interactions were limited to 25.1%, 7.6% and 7.5%, respectively, through sequential liposome incubations and singleplex capture bead designs. Here, ruthenium hexamine liposomes had only mediocre performance so that low overall signal strength translated into higher LODs and worse specificity. A different marker such as ferroin may be an option in the future. The identification of further electrochemical markers will provide new opportunities for liposomes to function as multiplex, orthogonal or internal standard labels in electrochemical bioassays.

Signaling strategies of silver nanoparticles in optical and electrochemical biosensors: considering their potential for the point-of-care.

Silver nanoparticles (AgNPs) have long been overshadowed by gold NPs' success in sensor and point-of-care (POC) applications. However, their unique physical, (electro)chemical, and optical properties make them excellently suited for such use, as long as their inherent higher instability toward oxidation is controlled. Recent advances in this field provide novel strategies that demonstrate that the AgNPs' inherent capabilities improve sensor performance and enable the specific detection of analytes at low concentrations. We provide an overview of these advances by focusing on the nanosized Ag (in the range of 1-100 nm) properties with emphasis on optical and electrochemical biosensors. Furthermore, we critically assess their potential for point-of-care sensors discussing advantages as well as limitations for each detection technique. We can conclude that, indeed, strategies using AgNP are ready for sensitive POC applications; however, research focusing on the simplification of assay procedures is direly needed for AgNPs to make the successful jump into actual applications.