A low cost and high throughput magnetic bead-based immuno-agglutination assay in confined droplets

Abstract

Although passive immuno-agglutination assays consist of one step and simple procedures, they are usually not adapted for high throughput analyses and they require expensive and bulky equipment for quantitation steps. Here we demonstrate a low cost, multimodal and high throughput immuno-agglutination assay that relies on a combination of magnetic beads (MBs), droplets microfluidics and magnetic tweezers. Antibody coated MBs were used as a capture support in the homogeneous phase. Following the immune interaction, water in oil droplets containing MBs and analytes were generated and transported in Teflon tubing. When passing in between magnetic tweezers, the MBs contained in the droplets were magnetically confined in order to enhance the agglutination rate and kinetics. When releasing the magnetic field, the internal recirculation flows in the droplet induce shear forces that favor MBs redispersion. In the presence of the analyte, the system preserves specific interactions and MBs stay in the aggregated state while in the case of a non-specific analyte, redispersion of particles occurs. The analyte quantitation procedure relies on the MBs redispersion rate within the droplet. The influence of different parameters such as magnetic field intensity, flow rate and MBs concentration on the agglutination performances have been investigated and optimized. Although the immuno-agglutination assay described in this work may not compete with enzyme linked immunosorbent assay (ELISA) in terms of sensitivity, it offers major advantages regarding the reagents consumption (analysis is performed in sub microliter droplet) and the platform cost that yields to very cheap analyses. Moreover the fully automated analysis procedure provides reproducible analyses with throughput well above those of existing technologies. We demonstrated the detection of biotinylated phosphatase alkaline in 100 nL sample volumes with an analysis rate of 300 assays per hour and a limit of detection of 100 pM.

Chromatin immunoprecipitation in microfluidic droplets: towards fast and cheap analyses

“…In this context, we developed an integrated ChIP procedure, from chromatin immunoprecipitation to DNA purification, using a lab on tube platform that has been previously described (see Fig.1, Supplementary Fig.2) [20][21][22][23] . This platform comprises a single 300 µm diameter Teflon tubing and a syringe pump that enables the back and forth movement of droplets.…”

Rapid and continuous magnetic separation in droplet microfluidic devices

“…We could also expect that given the spacing between consecutive droplets there is no recirculating zones at the front and back of droplets 51 . In addition, the presence of bead aggregates is likely to influence the flow fields such that the latter will depend on the presence and the strength of the magnetic field 60 . We therefore focus on our engineering and experimental efforts to improve separation efficiency of magnetic beads inside droplets, and refer a more quantitative explanation for our observed distribution patterns to future efforts in flow simulations and hydrodynamic theory.…”

Microfluidics for DNA and Protein Analysis with Multiplex Microbead-Based Assays

“…(b) Bright-field micrographs of magnetic bead agglutination with analyte in droplet with the concentration of 0 and 2 nM before, during, and after magnetic confinement. Reproduced from [103] with permission of The Royal Society of Chemistry aggregated state while in the case of a nonspecific analyte, redispersion of particles occurred (Fig. 8.5b).…”

“…8.5a) [103]. Water-in-oil droplets containing antibody-coated magnetic beads (MBs) and analytes were generated and transported in Teflon tubing.…”