This approach is extremely promising for flow cytometry applications, since it presents the great advantage of performing flow focusing within a single-inlet single-outlet microfluidic device without the need for external fields (that complicate the setups) and at high throughput. In recent years, inertial microfluidics has been successfully proposed to effectively manipulate particle positions in microfluidic channels 13– 15, by simply exploiting the channel geometry and the flow rate. So far, different methods have been proposed to obviate this limitation, but they usually envisage the use of external fields (such as acoustic 6, electrical 7 or optical fields 8) or the presence of multi-lateral sheaths flows 9– 12. Unfortunately, due to the intrinsic 2D nature of the standard fabrication techniques, this achievement is difficult to obtain. Moreover, a fundamental requirement is the capability to perform 3D particle focusing that forces the cells to flow one behind the other to enable enhanced optical alignment with better signal 4, 5. ![]() To be truly competitive with standard instrumentation, these microfluidic devices should satisfy several challenging constraints: (i) a high throughput, necessary to process large amounts of samples in a reasonable amount of time (ii) a low pressure drop over the microfluidic device, which basically means low fluidic resistance, thus allowing vacuum driven flows (iii) optical accessibility inside the microchannel to allow high quality optical detection 3. Indeed, the microfluidic approach favors analysis at the single-cell level, a reduced sample contamination, an increased analysis sensitivity and a higher processing automation 1, 2. On-chip flow cytometers promise several advantages in terms of cost reduction, portability, and increased device performance with respect to standard bench-top approaches. ![]() The resulting device is quite interesting for high-throughput on-chip flow cytometry. In addition, the low fluidic resistance of the channel network is compatible with vacuum driven flows. The use of out-of-plane loops favors a compact parallelization of multiple focusing channels, allowing one to process large amounts of samples. The use of tightly curving three-dimensional microfluidic loops produces strong Dean drag forces along the whole loop but also induces an asymmetric Dean flow decay in the subsequent straight channel, thus producing rapid cross-sectional mixing flows that assist with 3D particle focusing. The device is fabricated by femtosecond laser irradiation followed by chemical etching, which is a simple two-step process enabling the creation of 3D microfluidic networks in fused silica glass substrates. This device exploits inertial fluidic effects that occur in a laminar regime at sufficiently high flow rates, manipulating the particle positions by the combination of inertial lift forces and Dean drag forces. In this work, we present a compact microfluidic device capable of 3D particle focusing at high flow rates and with a small footprint, without the requirement of external fields or lateral sheath flows, but using only a single-inlet, single-outlet microfluidic sequence of straight channels and tightly curving vertical loops. Currently, 3D focusing is achieved mainly in devices with complex layouts, additional sheath fluids, and complex pumping systems. ![]() Three-dimensional (3D) particle focusing in microfluidics is a fundamental capability with a wide range of applications, such as on-chip flow cytometry, where high-throughput analysis at the single-cell level is performed.
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