Hydrogel Microvalves as Control Elements for Parallelized Enzymatic Cascade Reactions in Microfluidics

Abstract

Compartmentalized microfluidic devices with immobilized catalysts are a valuable tool for overcoming the incompatibility challenge in (bio) catalytic cascade reactions and high-throughput screening of multiple reaction parameters. To achieve flow control in microfluidics, stimuli-responsive hydrogel microvalves were previously introduced. However, an application of this valve concept for the control of multistep reactions was not yet shown. To fill this gap, we show the integration of thermoresponsive poly(N-isopropylacrylamide) (PNiPAAm) microvalves (diameter: 500 and 600 µm) into PDMS-on-glass microfluidic devices for the control of parallelized enzyme-catalyzed cascade reactions. As a proof-of-principle, the biocatalysts glucose oxidase (GOx), horseradish peroxidase (HRP) and myoglobin (Myo) were immobilized in photopatterned hydrogel dot arrays (diameter of the dots: 350 µm, amount of enzymes: 0.13–2.3 µg) within three compartments of the device. Switching of the microvalves was achieved within 4 to 6 s and thereby the fluid pathway of the enzyme substrate solution (5 mmol/L) in the device was determined. Consequently, either the enzyme cascade reaction GOx-HRP or GOx-Myo was performed and continuously quantified by ultraviolet-visible (UV-Vis) spectroscopy. The functionality of the microvalves was shown in four hourly switching cycles and visualized by the path-dependent substrate conversion.

Enzymatic Synthesis of Sialic Acids in Microfluidics to Overcome Cross-Inhibitions and Substrate Supply Limitations

“…A design of a microfluidic reaction chamber filled with a hexagonal array of 98 hydrogel−enzyme-dots was previously established (see S1.10). 29 In this design, the t r of the fluid at the hydrogel array is 10 min at a FR of 1 μL/min, corresponding to a fluid volume of 10 μL in the microfluidic chamber.…”

“…Enzymatic (cascade) reactions were previously integrated in microfluidic devices and various immobilization techniques were established, such as the binding to silica nanosprings, the attachment to surfaces and microbeads, − or the entrapment in microstructured hydrogels. − Even compartmentalized enzyme immobilization for performing stepwise cascade reactions was demonstrated, but incompatibility issues were not addressed under continuous flow. − …”

Digital manufacturing for accelerating organ-on-a-chip dissemination and electrochemical biosensing integration

“…The electrode role is not restricted only to real-time sensing of chemical and physical information in OoC. Electrodes have been also employed in MPS platforms and lab-on-chip devices as heating elements to study sperm thermotaxis, 128 smart hydrogel valve control systems, 129 or micropattern hydrogels inside microchannels. 130,131 Hydrogels may not only be used to embed biorecognition elements or to improve electrode longterm performance, 132 but also, they can be used to embed the MPS under study.…”

Fabrication of Thermo-Responsive Controllable Shape-Changing Hydrogel

“…Thermal modulation reversibly switches the polymer through the volume phase transition temperature (VPTT) around 33–35 °C from a water-swollen, hydrophilic state into a de-swollen, hydrophobic state. [ 18 , 19 , 20 ] PNIPAAm hydrogels were applied in various fields with soft actuator actions, such as analytical separation and detection [ 21 ], antifouling coatings [ 22 ], soft robotics [ 23 , 24 ], microfluidic flow controlling [ 25 , 26 ], and also in additive manufacturing (AM) [ 27 , 28 , 29 ]. PNIPAAm hydrogels, however, have a limited efficiency as an actuator because of their limited mechanical property profile.…”