# A Low-Cost Laser-Prototyped Microfluidic Device for Separating Cells and Bacteria
## 1 Introduction
Two popular types of inertial cell separation methods are serpentine channels and spiral microfluidics [36; 37; 38; 39]. Both serpentine channels and spiral microfluidics offer several advantages over traditional cell separation methods, such as higher throughput, lower sample volume, and increased purity. These methods also require minimal external equipment and can be easily integrated with other microfluidic devices, making them ideal for a wide range of applications, such as point-of-care diagnostics, drug discovery, and cancer research. Even though the passive microfluidic methods are simpler compared to the active ones, the majority of the passive devices rely on expensive and costly cleanroom microfabrication facilities which are not readily available in developing countries [40; 41]. While there are some preliminary reports on 3D-printed microfluidic devices for cell separation, more examples and demonstrations are critical to extend and advancing this field with simpler and better tools [42; 43]. Therefore, more efforts are still needed to investigate low-cost fabrication alternatives to conventional microfluidic devices to achieve low-cost and accessible cell separation applications. Microfluidic-based separation techniques have emerged as a powerful tool for the efficient isolation and sorting of cells and bacteria [44; 45]. These techniques utilize the unique properties of microscale fluid flow to manipulate and separate biological entities with high precision and throughput. Numerous studies have demonstrated the effectiveness of microfluidic platforms in various applications, including cell sorting, cell enrichment, and bacterial analysis. For example, in one study by [PERSON] et al., a microfluidic device was developed for the separation of heterogeneous particles or cell mixtures in a continuous flow using acoustophoresis [46]. In this device, a standing surface acoustic wave (SSAW) field was created across a microchannel by propagating two identical surface acoustic waves (SAWs) generated by interdigital transducers (IDTs). The developed device successfully separated two types of fluorescent microspheres with different sizes. Furthermore, Escherichia coli bacteria mixed with peripheral blood mononuclear cells (PBMCs) were efficiently isolated using the SSAW-based separation technique. Flow cytometric analysis of the collected samples indicated a purity of 95.65% for the separated _E. coli_ bacteria. In another study, a label-free, size-based bacteria separation method from whole blood was achieved using elasto-inertial microfluidics [47]. In elasto-inertial microfluidics, the migration of blood cells into a non-Newtonian solution is facilitated by viscoelastic flow, while smaller bacteria remain in the streamline at the blood sample entrance and can be effectively separated. The flow conditions were optimized using particles, resulting in a continuous separation of 5 \(\upmu\)m particles from 2 \(\upmu\)m particles with yields of 95% and 93% at respective outlets. Subsequently, bacteria were continuously separated from undiluted whole blood samples with an efficiency of 76%. Even though these examples demonstrate good separation efficiencies, the fabrication of the device requires lengthy and expensive procedures that limit the application of such devices in rapid disease screenings. Such wide-scale applications require low-cost single-use, disposable devices. Thus, to enable disposable and affordable point-of-care microfluidic devices, alternative fabrication approaches should be explored to further prove the feasibility of low-cost manufacturing for lab-on-chip applications. In this work, we demonstrate a simple and low-cost microfluidic device fabricated by implementing rapid laser prototyping using polymethyl methacrylate (PMMA). The device is designed to have a single inlet, a serpentine section, and a spiral microfluidic channel to achieve sheath-less cell separation.