Assembly-line manipulation of droplets in microfluidic platform for fluorescence encoding and simultaneous multiplexed DNA detection

In this article, a new mode of droplets manipulation is presented and applied for simultaneous multiplexed DNA detection. We call this droplets manipulation, “assembly-line manipulation of droplets (ALMD)”. Firstly, multiple droplets containing the same target mixtures are generated in the microchannel, and then fused with later generated different droplets containing corresponding probes, respectively. Finally, all the fused droplets were fluorescence imaged on-line and real-time. The successful implementation of droplets fluorescence encoding based on ALMD shows the reproducibility and accuracy of this manipulation mode. As a proof-of-concept application, the simultaneous multiplexed DNA detection was carried out through the model of human immunodeficiency virus (HIV) gene sequence and variola virus (small pox, VV) gene sequence based on ALMD in the microfluidic system. It is proved that this method achieves simultaneous multiplexed DNA measurements with a significantly time-saving way and without different dye-labelled probes or complex operation procedures. In addition, it reveals the possibility of high-throughput biosensing with simple chip design and detection equipment. [Display omitted] •A novel strategy, assembly-line manipulation of droplets (ALMD) is presented.•The microfluidic platform based on ALMD can be used to fluorescence encoding.•Based on ALMD, simultaneous multiplexed DNA on-line detection has been realized.•The method based on ALMD has potential for high-throughput DNA assay.