Characterization of Thermoplastic Fusion Bonding of Microchannels using Pressure Assisted Boiling Point Control System

An innovative method of thermoplastic fusion bonding using a pressure assisted boiling point control (PABP) system was characterized to determine the optimum parameters for bonding polymethyl methacrylate (PMMA) components containing microchannels and thin, 250 µm cover sheets. The PABP system enables precise control of the temperature boundary condition and the applied pressure by immersing the components being bonded in boiling water and varying the vapor pressure. Test structure geometries containing microchannels of two depths and four different aspect ratios were designed: 1:10 (Depth: 10µm, Width: 100 µm and Depth: 5µm, Width: 50 µm), 1:50 (Depth: 10µm, Width: 500µm and Depth: 5µm, Width: 250 µm), 1:100 (Depth: 10µm, Width: 1000µm and Depth: 5µm, Width: 500µm) and 1:200 (Depth: 10µm, Width: 2000µm and Depth: 5µm, Width: 1000µm). Microchannels were hot embossed using micro-milled brass mold inserts. Bonding conditions were optimized by observing microchannel deformation under a microscope. The quality of the bonded samples were rupture and leak tested to determine the integrity and strength of the bonds. Mean rupture pressures for channels of AR of 1:10, 1:50 and 1:100 were 851.02 kPa, 780.14 kPa and 706.09 kPa repectively for shallower channels and 831.93 kPa, 739.3 kPa and 524.38 kPa respectively for deeper channels bonded using open loop system. Rupture pressure decreased with decreasing AR and was higher for shallower channels. A closed loop control system was developed for the automatic temperature control. Results of bonding with both open loop and closed loop systems were compared. Mean rupture pressure for channels of AR 1:10, 1:50 and 1:100 for 5 µm depth were 977.54 kPa, 930.93 kPa and 751.39 kPa respectively and 912.11 kPa, 800.07 kPa and 550.96 kPa respectively for 10 µm depth. It was found that the rupture test results were more consistent and repeatable with closed loop system because of better control of the bonding temperature

Fluidic operation of a polymer-based nanosensor chip for analysing single molecules

Most medical diagnostic tests are expensive, involve slow turnaround times from centralized laboratories and require highly specialized equipment with seasoned technicians to carry out the assay. To facilitate realization of precision medicine at the point of care, we have developed a mixed-scale nanosensor chip featuring high surface area pillar arrays where solid-phase reactions can be performed to detect and identify nucleic acid targets found in diseased patients. Products formed can be identified and detected using a polymer nanofluidic channel. To guide delivery of this platform, we discuss the operation of various components of the device and simulations (COMSOL) used to guide the design by investigating parameters such as pillar array loading, and hydrodynamic and electrokinetic flows. The fabrication of the nanosensor is discussed, which was performed using a silicon (Si) master patterned with a combination of focused ion beam milling and photolithography with deep reactive ion etching. The mixed-scale patterns were transferred into a thermoplastic via thermal nanoimprint lithography, which facilitated fabrication of the nanosensor chip making it appropriate for in vitro diagnostics. The results from COMSOL were experimentally verified for hydrodynamic flow using Rhodamine B as a fluorescent tracer and electrokinetic flow using single fluorescently labelled oligonucleotides (single-stranded DNAs, ssDNAs)

Microfluidic Device for On-Chip Immunophenotyping and Cytogenetic Analysis of Rare Biological Cells

This work is licensed under a Creative Commons Attribution 4.0 International License.The role of circulating plasma cells (CPCs) and circulating leukemic cells (CLCs) as biomarkers for several blood cancers, such as multiple myeloma and leukemia, respectively, have recently been reported. These markers can be attractive due to the minimally invasive nature of their acquisition through a blood draw (i.e., liquid biopsy), negating the need for painful bone marrow biopsies. CPCs or CLCs can be used for cellular/molecular analyses as well, such as immunophenotyping or fluorescence in situ hybridization (FISH). FISH, which is typically carried out on slides involving complex workflows, becomes problematic when operating on CLCs or CPCs due to their relatively modest numbers. Here, we present a microfluidic device for characterizing CPCs and CLCs using immunofluorescence or FISH that have been enriched from peripheral blood using a different microfluidic device. The microfluidic possessed an array of cross-channels (2–4 µm in depth and width) that interconnected a series of input and output fluidic channels. Placing a cover plate over the device formed microtraps, the size of which was defined by the width and depth of the cross-channels. This microfluidic chip allowed for automation of immunofluorescence and FISH, requiring the use of small volumes of reagents, such as antibodies and probes, as compared to slide-based immunophenotyping and FISH. In addition, the device could secure FISH results in <4 h compared to 2–3 days for conventional FISH

Intracellular Delivery of Exogenous Macromolecules into Human Mesenchymal Stem Cells by Double Deformation of the Plasma Membrane

Physical techniques for intracellular delivery of exogeneous materials offer an attractive strategy to enhance the therapeutic efficiency of stem cells. However, these methods are currently limited by poor delivery efficiency as well as cytotoxic effects. Here, a high throughput microfluidic device is designed for efficient (approximate to 85%) cytosolic delivery of exogenous macromolecules with minimal cell death (less than 10%). The designed microfluidic device enables the generation of transient pores as the cells pass through the micron-sized constrictions (6-10 mu m) leading to the passive diffusion of extracellular cargos into the cell cytosol. Specifically, the microfluidic system is designed to induce a double deformation on the cell membrane at the squeezing zones to maximize intracellular delivery. Additionally, the flow rate, ionic concentration, and the molecular weight of the cargo are optimized for maximum efficiency. The optimized device enables cytosolic diffusion of small (3 kDa) and large molecules (70 kDa) without inducing any apoptotic effect. Overall, this double cell deformation platform offers new opportunities to rapidly and efficiently deliver extracellular cargo into stem cells without affecting their viability and functionality