Multifuntional Gold Nanoparticles for the SERS Detection of Pathogens Combined with a LAMP-in-Microdroplets Approach.

Alexandra Teixeira,Laura Rodriguez-Lorenzo,Alejandro Garrido-Maestu,Marta Prado,Foteini Roumani,Begoña Espiña,Lorena Diéguez,Sara Abalde-Cela,Juan L Paris

doi:10.3390/ma13081934

Abstract

We developed a droplet-based optofluidic system for the detection of foodborne pathogens. Specifically, the loop-mediated isothermal amplification (LAMP) technique was combined with surface-enhanced Raman scattering (SERS), which offers an excellent method for DNA ultradetection. However, the direct SERS detection of DNA compromises the simplicity of data interpretation due to the variability of its SERS fingerprints. Therefore, we designed an indirect SERS detection method using multifunctional gold nanoparticles (AuNPs) based on the formation of pyrophosphate generated during the DNA amplification by LAMP. Towards this goal, we prepared multifunctional AuNPs involving three components with key roles: (1) thiolated poly(ethylene glycol) as stabilizing agent, (2) 1-naphthalenethiol as Raman reporter, and (3) glutathione as a bioinspired chelating agent of magnesium (II) ions. Thus, the variation in the SERS signal of 1-naphthalenethiol was controlled by the aggregation of AuNPs triggered by the complexation of pyrophosphate and glutathione with free magnesium ions. Using this strategy, we detected Listeria monocytogenes, not only in buffer, but also in a food matrix (i.e., ultra-high temperaturemilk) enabled by the massive production of hotspots as a result of the self-assemblies that enhanced the SERS signal. This allowed the development of a microdroplet-LAMP-SERS platform with isothermal amplification and real-time identification capabilities.

Highlights

In recent years, microfluidics technology has demonstrated increasing relevance in the development of analytical tools in different fields of research and applications [1,2]
GSH is included in these biosynthesized small molecules and it was selected to functionalize a surface-enhanced Raman scattering (SERS) substrate based on AuNPs [37,38]
The chelating capacity of GHS has been used to design bioinspired sensors for heavy metals detection such as cadmium (II) [37,60] and lead (II) [37,61]. This simple interaction was not enough to develop a detection method for nucleic acids via loop-mediated isothermal amplification (LAMP) due to magnesium concentration being constant in the reaction mixture

Summary

Introduction

Microfluidics technology has demonstrated increasing relevance in the development of analytical tools in different fields of research and applications [1,2]. Microfluidic devices are very attractive in food safety analysis because they offer economic, portable, and disposable systems that can be used for on-site detection without the need of specific expertise when fully integrated. The implementation of point-of-care approaches for the detection of foodborne pathogens in the food chain is pivotal to food safety management [5]. The utilization of microfluidics, and microdroplets technology, a branch of microfluidics [6,7], may increase the options to achieve the implementation of point-of-care systems based on molecular techniques for the fast detection of pathogens in foodstuffs.

Objectives

Methods

Conclusion