Magnetically localized and wash-free fluorescent immuno-assay: From a research platform (MLFIA) to a multiplexed POC system (MagIA).

Sexually transmitted infections (STI) remain one of the world's public health priorities: Nearly 400 million people are infected not only in emerging, but also in western countries. HIV, HBV and HCV share common infection pathways; thus these 3 diseases are recommended to be tested at the same time. However, this combined approach is currently mainly available in laboratories, and seldomly at the Point-of-care (POC). Consequently, there is a need for a STI screening POC platform with laboratory-like performance. Such a platform should be autonomous and portable and enable multiplexed screening from capillary blood. The previously developed and introduced MLFIA (Magnetically Localized and wash-free Fluorescent Immuno-Assay) technology has the potential to address these needs, as the MLFIA 18-chamber microfluidic cartridge and the MLFIA Analyzer were previously characterized and evaluated with plasma and serum from patients infected with HIV, Hepatitis B (Hep B) or C (Hep C). Here, we present the efforts to transfer this research platform (MLFIA) to a fully integrated multi-analysis solution (MagIA). First, we present the design changes of the consumable enabling to perform multiple assays in parallel, a fast filling of the cartridge with patient samples, and a homogeneous reagent/sample incubation. Second, we describe the development a piezoelectric actuator integrated into the Analyzer: this mixing module allows for an automated, fully integrated and portable workflow, with homogeneous in-situ mixing capabilities. The obtained MagIA platform was further characterized and validated for immunoassays (LOD, cartridge stability over time), using various biological models including OVA and IgG. We discuss the performances of the MLFIA and MagIA platforms for the detection of HIV / Hep B / Hep C using results from 102 patient plasma samples. Lastly, we assessed the compatibility of the MagIA platform with veinous and capillary blood samples as a final step towards its POC validation.

Magnetically localized and wash-free fluorescence immunoassay (MLFIA): proof of concept and clinical applications.

Immunoassays are used for many applications in various markets, from clinical diagnostics to the food industry, generally relying on gold-standard ELISAs that are sensitive, robust, and cheap but also time-consuming and labour intensive. As an alternative, we propose here the magnetically localized and wash-free fluorescence immunoassay (MLFIA): a no-wash assay to directly measure a biomolecule concentration, without mixing nor washing steps. To do so, a fluorescence no-wash measurement is performed to generate a detectable signal. It consists of a differential measurement between the fluorescence of fluorophores bound to magnetic nanoparticles specifically captured by micro-magnets against the residual background fluorescence of unbound fluorophores. Targeted biomolecules (antibodies or antigens) are locally concentrated on micro-magnet lines, with the number of captured biomolecules quantitatively measured without any washing step. The performance of the MLFIA platform is assessed and its use is demonstrated with several biological models as well as clinical blood samples for HIV, HCV and HBV detection, with benchmarking to standard analyzers of healthcare laboratories. Thus, we demonstrated for the first time the versatility of the innovative MLFIA platform. We highlighted promising performances with the successful quantitative detection of various targets (antigens and antibodies), in different biological samples (serum and plasma), for different clinical tests (HCV, HBV, HIV).

Magnetophoretic induced convective capture of highly diffusive superparamagnetic nanoparticles.

Micro-magnets producing magnetic field gradients as high as 106 T m-1 have been used to efficiently trap nanoparticles with a magnetic core of just 12 nm in diameter. Particle capture efficiency increases with increasing particle concentration. Comparison of measured capture kinetics with numerical modelling reveals that a threshold concentration exists below which capture is diffusion-driven and above which it is convectively-driven. This comparison also shows that two-way fluid-particle coupling is responsible for the formation of convective cells, the size of which is governed by the height of the droplet. Our results indicate that for a suspension with a nanoparticle concentration suitable for bioassays (around 0.25 mg ml-1), all particles can be captured in less than 10 minutes. Since nanoparticles have a significantly higher surface-to-volume ratio than the more widely used microparticles, their efficient capture should contribute to the development of next generation digital microfluidic lab-on-chip immunoassays.