An inexpensive microfluidic device for three-dimensional hydrodynamic focusing in imaging flow cytometry

Abstract

We present design, characterization, and testing of an inexpensive, sheath-flow based microfluidic device for three-dimensional (3D) hydrodynamic focusing of cells in imaging flow cytometry. In contrast to other 3D sheathing devices, our device hydrodynamically focuses the cells in a single-file near the bottom wall of the microchannel that allows imaging cells with high magnification and low working distance objectives, without the need for small device dimensions. The relatively large dimensions of the microchannels enable easy fabrication using less-precise fabrication techniques, and the simplicity of the device design avoids the need for tedious alignment of various layers. We have characterized the performance of the device with 3D numerical simulations and validated these simulations with experiments of hydrodynamic focusing of a fluorescently dyed sample fluid. The simulations show that the width and the height of the 3D focused sample stream can be controlled independently by varying the heights of main and side channels of the device, and the flow rates of sample and sheath fluids. Based on simulations, we also provide useful guidelines for choosing the device dimensions and flow rates for focusing cells of a particular size. Thereafter, we demonstrate the applicability of our device for imaging a large number of RBCs using brightfield microscopy. We also discuss the choice of the region of interest and camera frame rate so as to image each cell individually in our device. The design of our microfluidic device makes it equally applicable for imaging cells of different sizes using various other imaging techniques such as phase-contrast and fluorescence microscopy.

A Novel Detachable, Reusable, and Versatile Acoustic Tweezer Manipulation Platform for Biochemical Analysis and Detection Systems

“…In flow cytometry, cells need to be lined up before they pass through the monitoring site [ 32 , 33 ]. Many scholars use hydrodynamic methods to gather particles together [ 34 , 35 , 36 ]. By adding glass with a high acoustic speed, the acoustic attenuation angle area was smaller.…”

An optimized PDMS microfluidic device for ultra-fast and high-throughput imaging flow cytometry

“…Another kind of vertical hydrodynamic focusing is achieved in multi-layer T 37 or Y 38 shaped channels, where the sample channel has a lower height than the sheath channels, 39 and vertical focusing could be realized by increasing the velocity or Reynolds number. 40 Additionally, the inertial effects on fluid is pretty strong at high velocities, which could affect the focusing of cells in the microchannels, 41 leading to a result like cell separation, 42,43 but unstable. Even worse, inertial focusing always demands a long channel, 44 which keeps a high flow resistance limited to a higher velocity.…”

Simplified 3D hydrodynamic flow focusing for lab-on-chip single particle study

“…Nonetheless, also these devices show a limited throughput ranging from µL/min to 30 µL/min, mainly due to the de-formability of polydimethylsiloxane (PDMS), of which they are made. Instead, Patel et al 33 have used a similar strategy, but their device flow rate is not limited thanks to the polymethyl methacrylate (PMMA) micromilling. However, since their focused particles flow on the bottom of the main channel, the device is exposed to clogging risks.…”