*3.1.8 Droplet microfluidics*

Droplet microfluidic techniques involve mixture of the aqueous phase and oil phase, and generation of water-in-oil microdroplets in the microfluidic chips [98]. Droplet shape offers encapsulation of a single cell in a picoliter droplet, which confines individual cell and entraps its secretion, restrains the interference from surrounding environment. To achieve real-time monitoring of cell secretion, droplets microfluidic techniques are conventionally integrated with cell-surface affinity sensors or bead-based biosensors. Qiu et al. [94] developed a membrane-anchored aptamer sensor and combined it with droplet microfluidics for IFN-γ detection. This aptamer sensor has a hairpin structure, resulting in quenched fluorescence. Upon binding with IFN-γ, the aptamer switches on the fluorescence signal by aptamer structure change. The limit of detection was approximately 10.0 nM. Due to the proximity of the aptamer sensor to the cell surface, this method achieved monitoring of cytokine secretion at a single-cell level. Chokkalingam et al. [99] took advantage of free capture beads in droplets to detect the secreted cytokines from cells. The cytokine capture step and fluorescent detection step were separated by the gelation of droplets, and de-emulsification. Wei et al. [100] combined LSPR with droplet technique, developing a sensor with a detection limit of 6–7 ng/ ml for VEGF and IL-8. The LSPR spectrum shift was obtained by dark-field spectroscope. Due to the advantages of droplet technique, the plasmonic droplets in a continuous flow allow high-throughput detection of 600–1800 droplets/min, 100–150 cells/min.
