A Label-Free Droplet Sorting Platform Integrating Dielectrophoretic Separation for Estimating Bacterial Antimicrobial...

Antimicrobial resistance (AMR) has become a crucial global health issue. Antibiotic-resistant bacteria can survive after antibiotic treatments, lowering drug efficacy and increasing lethal risks. A microfluidic water-in-oil emulsion droplet system can entrap microorganisms and antibiotics within the tiny bioreactor, separate from the surroundings, enabling independent assays that can be performed in a high-throughput manner. This study presents the development of a label-free dielectrophoresis (DEP)-based microfluidic platform to sort droplets that co-encapsulate Escherichia coli (E. coli) and ampicillin (Amp) and droplets that co-encapsulate Amp-resistant (AmpR) E. coli with Amp only based on the conductivity-dependent DEP force (F) without the assistance of optical analyses. The 9.4% low conductivity (LC) Luria-Bertani (LB) broth diluted with 170 mM mannitol can maintain E. coli and AmpR E. coli growth for 3 h and allow Amp to kill almost all E. coli, which can significantly increase the LCLB conductivity by about 100 μS/cm. Therefore, the AmpR E. coli/9.4%LCLB/Amp where no cells are killed and the E. coli/9.4%LCLB/Amp-containing droplets where most of the cells are killed can be sorted based on this conductivity difference at an applied electric field of 2 MHz and 100 V_pp that generates positive F. Moreover, the sorting ratio significantly decreased to about 50% when the population of AmpR E. coli was equal to or higher than 50% in droplets. The conductivity-dependent DEP-based sorting platform exhibits promising potential to probe the ratio of AmpR E. coli in an unknown bacterial sample by using the sorting ratio as an index.