Abstract
Digital Light Processing (DLP) stereolithography (SLA) as a high-resolution 3D printing process offers a low-cost alternative for prototyping of microfluidic geometries, compared to traditional clean-room and workshop-based methods. Here, we investigate DLP-SLA printing performance for the production of micro-chamber chip geometries suitable for Polymerase Chain Reaction (PCR), a key process in molecular diagnostics to amplify nucleic acid sequences. A DLP-SLA fabrication protocol for printed micro-chamber devices with monolithic micro-channels is developed and evaluated. Printed devices were post-processed with ultraviolet (UV) light and solvent baths to reduce PCR inhibiting residuals and further treated with silane coupling agents to passivate the surface, thereby limiting biomolecular adsorption occurences during the reaction. The printed devices were evaluated on a purpose-built infrared (IR) mediated PCR thermocycler. Amplification of 75 base pair long target sequences from genomic DNA templates on fluorosilane and glass modified chips produced amplicons consistent with the control reactions, unlike the non-silanized chips that produced faint or no amplicon. The results indicated good functionality of the IR thermocycler and good PCR compatibility of the printed and silanized SLA polymer. Based on the proposed methods, various microfluidic designs and ideas can be validated in-house at negligible costs without the requirement of tool manufacturing and workshop or clean-room access. Additionally, the versatile chemistry of 3D printing resins enables customised surface properties adding significant value to the printed prototypes. Considering the low setup and unit cost, design flexibility and flexible resin chemistries, DLP-SLA is anticipated to play a key role in future prototyping of microfluidics, particularly in the fields of research biology and molecular diagnostics. From a system point-of-view, the proposed method of thermocycling shows promise for portability and modular integration of funcitonalitites for diagnostic or research applications that utilize nucleic acid amplification technology.
Highlights
Several research groups demonstrated complete systems, integrating all steps for PCRbased diagnosis of various pathogens in crude samples for field-use [17,18,19]; to an extent, the potential of POC molecular diagnostic technology is met in some commercial cartridgebased Polymerase Chain Reaction (PCR) assays with pre-printed reagents for diagnosis of common infectious diseases and for drug sensitivity tests performed by trained personnel [20, 21]
Low-cost research prototyping of customised PCR microfluidics is highly desirable within the molecular diagnostics community, driven by the increased demand for low-cost POC molecular diagnostics
In this paper we have presented specific processing routes to repeatedly deliver low-cost microfluidic chamber PCR devices printed with Digital Light Processing (DLP) SLA in a two-step printing protocol with sacrificial wax
Summary
Several research groups demonstrated complete systems, integrating all steps for PCRbased diagnosis of various pathogens in crude samples for field-use [17,18,19]; to an extent, the potential of POC molecular diagnostic technology is met in some commercial cartridgebased PCR assays with pre-printed reagents for diagnosis of common infectious diseases and for drug sensitivity tests performed by trained personnel [20, 21]. Commercial quality microfluidics are manufactured at large volumes, mainly with microforming processes and medical grade thermoplastics, due to low unit costs Such processes are highly expensive for prototyping, due to the requirement of forming tool manufacturing and only validated designs justify investment in a tool [30]. Soft lithography of polydimethylsiloxane (PDMS) [17, 31], clean-room based photolithographic patterning or etching of SU-8, glass or silicon [18, 32] and material removal processes such as mechanical and laser micromachining [33] of a wide range of microfluidic materials are typically preferred for research prototyping and low volume production
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