RESUMO
Droplet microfluidics has the ability to greatly increase the throughput of screening and sorting of enzymes by carrying reagents in picoliter droplets flowing in inert oils. It was found with the use of a specific surfactant, the interfacial tension of droplets can be very sensitive to droplet pH. This enables the sorting of droplets of different pH when confined droplets encounter a microfabricated trench. The device can be extended to sort enzymes, as a large number of enzymatic reactions lead to the production of an acidic or basic product and a concurrent change in solution pH. The progress of an enzymatic reaction is tracked from the position of a flowing train of droplets. We demonstrate the sorting of esterase isoenzymes based on their enzymatic activity. This label-free technology, that we dub droplet sorting by interfacial tension (SIFT), requires no active components and would have applications for enzyme sorting in high-throughput applications that include enzyme screening and directed evolution of enzymes.
Assuntos
Hidrolases de Éster Carboxílico/isolamento & purificação , Ensaios Enzimáticos/métodos , Acetatos/química , Animais , Hidrolases de Éster Carboxílico/química , Ensaios Enzimáticos/instrumentação , Fluorocarbonos/química , Isoenzimas/química , Isoenzimas/isolamento & purificação , Dispositivos Lab-On-A-Chip , Fígado/enzimologia , Microfluídica/instrumentação , Microfluídica/métodos , Óleos/química , Fenóis/química , Reprodutibilidade dos Testes , Tensão Superficial , Suínos , Água/químicaRESUMO
Selection of live cells from a population is critical in many biological studies and biotechnologies. We present here a novel droplet microfluidic approach that allows for label-free and passive selection of live cells using the glycolytic activity of individual cells. It was observed that with the use of a specific surfactant utilized to stabilize droplet formation, the interfacial tension of droplets was very sensitive to pH. After incubation, cellular lactate release results in droplets containing a live cell to attain a lower pH than other droplets. This enables the sorting of droplets containing live cells when confined droplets flow over a microfabricated trench oriented diagonally with respect to the direction of flow. The technique is demonstrated with human U87 glioblastoma cells for the selection of only droplets containing a live cell while excluding either empty droplets or droplets containing a dead cell. This label-free sorting method, dubbed sorting by interfacial tension (SIFT) presents a new strategy to sort diverse cell types based on metabolic activity.