Automated Miniaturized Digital Microfluidic Antimicrobial Susceptibility Test Using a Chip-Integrated Optical Oxygen Sensor

We present the first digital microfluidic (DMF) antimicrobial susceptibility test (AST) using an optical oxygen sensor film for in-situ and real-time continuous measurement of extracellular dissolved oxygen (DO). The device allows one to monitor bacterial growth across the entire cell culture area, and the fabricated device was utilized for a miniaturized and automated AST. The oxygen-sensitive probe platinum(II)-5,10,15,20-tetrakis-(2,3,4,5,6-pentafluorophenyl)-porphyrin was embedded in a Hyflon AD 60 polymer and spin-coated as a 100 nm thick layer onto an ITO glass serving as the DMF ground electrode. This DMF-integrated oxygen sensing film was found to cause no negative effects to the droplet manipulation or cell growth on the chip. The developed DMF platform was used to monitor the DO consumption during growth caused by cellular respiration. A rapid and reliable twofold dilution procedure was developed and performed, and the AST with ATCC 25922 in the presence of ampicillin, chloramphenicol, and tetracycline at different concentrations from 0.5 to 8 μg mL was investigated. All sample dispensation, dilution, and mixing were performed automatically on the chip within 10 min. The minimum inhibitory concentration values measured from the DMF chip were consistent with those from the standard broth microdilution method but requiring only minimal sample handling and working with much smaller sample volumes.