Pressure-driven formation of sub-femtoliter droplets in constricted T-shaped nanofluidic channel and application to isolation and counting of single nanoparticles

Nanofluidics has developed, enabling the analysis of single nanoparticles such as viruses and macromolecules using nanochannels. However, since nanoparticles migrate randomly by Brownian motion with displacements comparable to the nanochannel size, it is difficult to array and process nanoparticles individually in continuous phase flows with biological analyte concentrations of pM–nM. The present study developed a method to generate aqueous droplets of 10 2 aL–10 0 fL volumes in an organic phase in a nanochannel by pressure-driven flow control, and applied the method to the isolation of single nanoparticles with 10 1 −10 2 pM concentrations. A T-shaped nanochannel with a narrow constriction section was proposed to locally enhance the shear force to form smaller droplets by 10 2 kPa external pressure, without increasing the pressure loss by downscaling the channel. The proposed channel design was validated, droplets with 0.8–1.0 fL volumes were formed with pL/min flow rates at Ca = 0.5–1.4 × 10 −3 , and the dynamics of droplet generation was well characterized by the linear scaling law. Using the developed device, encapsulation and counting of single fluorescent nanoparticles and single fluorescently-labeled DNA molecules in droplets were successfully demonstrated. This work will contribute to the development of droplet nanofluidics and ultrasmall analytical applications. • A method of sub-femtoliter droplet formation in a nanochannel by pressure-driven flow control was developed. • A constricted T-shaped nanochannel was proposed to locally enhance the shear force for generating smaller aqueous droplets. • Droplet formation in the nanochannel was achieved at pL/min flow rates with the dynamics similar to that in microspaces. • Encapsulation and counting of single nanoparticles in sub-femtoliter droplets were achieved at 35−140 pM concentrations. • The developed method was successfully applied to the concentration measurement of fluorescently-labeled DNA molecules.