Microscale Interfacial Polymerization on a Chip.

Forming hydrogels with precise geometries is challenging and mostly done using photopolymerization, which involves toxic chemicals, rinsing steps, solvents, and bulky optical equipment. Here, we introduce a new method for in situ formation of hydrogels with a well-defined geometry in a sealed microfluidic chip by interfacial polymerization. The geometry of the hydrogel is programmed by microfluidic design using capillary pinning structures and bringing into contact solutions containing hydrogel precursors from vicinal channels. The characteristics of the hydrogel (mesh size, molecular weight cut-off) can be readily adjusted. This method is compatible with capillary-driven microfluidics, fast, uses small volumes of reagents and samples, and does not require specific laboratory equipment. Our approach creates opportunities for filtration, hydrogel functionalization, and hydrogel-based assays, as exemplified by a rapid, compact competitive immunoassay that does not require a rinsing step.

Complex Nucleic Acid Hybridization Reactions inside Capillary-Driven Microfluidic Chips.

Nucleic acid hybridization reactions play an important role in many (bio)chemical fields, for example, for the development of portable point-of-care diagnostics, and often such applications require nucleic acid-based reaction systems that ideally run without enzymes under isothermal conditions. The use of novel capillary-driven microfluidic chips to perform two isothermal nucleic acid hybridization reactions, the simple opening of molecular beacon structures and the complex reaction cascade of a clamped-hybridization chain reaction (C-HCR), is reported here. For this purpose, reagents are arranged in a self-coalescence module (SCM) of a passive silicon microfluidic chip using inkjet spotting. The SCM occupies a footprint of ≈7 mm2 of a ≈0.4 × 2 cm2 microfluidic chip. By means of fluorophore-labeled DNA probes, the hybridization reactions can be analyzed in just ≈2 min and using only ≈3 µL of the sample. Furthermore, the SCM chip offers a variety of reagent delivery options, allowing, for example, the influence of the initiator concentration on the kinetics of C-HCR to be investigated systematically with minimal sample and time requirements. These results suggest that self-powered microfluidic chips equipped with a SCM provide a powerful platform for performing and investigating complex reaction systems.

Bottom-up Construction of a Primordial Carboxysome Mimic.

We have constructed a synthetic mimic of the carboxysome, a cyanobacterial carbon-fixing organelle. Using an electrostatic tagging system, we coencapsulated the two key carboxysomal enzymes, ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and carbonic anhydrase (CA), in an engineered protein cage based on lumazine synthase. A statistically significant kinetic effect of coencapsulated CA on RuBisCO activity was not observed under ambient or oxygen saturated conditions, suggesting that enzyme proximity alone may not be the key determinant in carboxysome function. The capsid shell protected the enzyme from proteolytic damage, a factor that could have provided early cyanobacteria with an evolutionary benefit. Our strategy to coencapsulate different proteins can easily be extended to other sequentially acting enzymes and lays down principles for developing artificial organelles to control biosynthetic pathways in vivo.

Rapid quantitative assays for glucose-6-phosphate dehydrogenase (G6PD) and hemoglobin combined on a capillary-driven microfluidic chip.

Rapid tests for glucose-6-phosphate dehydrogenase (G6PD) are extremely important for determining G6PD deficiency, a widespread metabolic disorder which triggers hemolytic anemia in response to primaquine and tafenoquine medication, the most effective drugs for the radical cure of malaria caused by Plasmodium parasites. Current point-of-care diagnostic devices for G6PD are either qualitative, do not normalize G6PD activity to the hemoglobin concentration, or are very expensive. In this work we developed a capillary-driven microfluidic chip to perform a quantitative G6PD test and a hemoglobin measurement within 2 minutes and using less than 2 µL of sample. We used a powerful microfluidic module to integrate and resuspend locally the reagents needed for the G6PD assay and controls. We also developed a theoretical model that successfully predicts the enzymatic reactions on-chip, guides on-chip reagent spotting and allows efficient integration of multiple assays in miniaturized formats with only a few nanograms of reagents.

Programmable hydraulic resistor for microfluidic chips using electrogate arrays.

Flow rates play an important role in microfluidic devices because they affect the transport of chemicals and determine where and when (bio)chemical reactions occur in these devices. Flow rates can conveniently be determined using external peripherals in active microfluidics. However, setting specific flow rates in passive microfluidics is a significant challenge because they are encoded on a design and fabrication level, leaving little freedom to users for adjusting flow rates for specific applications. Here, we present a programmable hydraulic resistor where an array of "electrogates" routes an incoming liquid through a set of resistors to modulate flow rates in microfluidic chips post-fabrication. This approach combines a battery-powered peripheral device with passive capillary-driven microfluidic chips for advanced flow rate control and measurement. We specifically show a programmable hydraulic resistor composed of 7 parallel resistors and 14 electrogates. A peripheral and smartphone application allow a user to activate selected electrogates and resistors, providing 127 (27-1) flow resistance combinations with values spanning on a 500 fold range. The electrogates feature a capillary pinning site (i.e. trench across the flow path) to stop a solution and an electrode, which can be activated in a few ms using a 3 V bias to resume flow based on electrowetting. The hydraulic resistor and microfluidic chip shown here enable flow rates from ~0.09 nL.s-1 up to ~5.66 nL.s-1 with the resistor occupying a footprint of only 15.8 mm2 on a 1 × 2 cm2 microfluidic chip fabricated in silicon. We illustrate how a programmable hydraulic resistor can be used to set flow rate conditions for laminar co-flow of 2 liquids and the enzymatic conversion of a substrate by stationary enzymes (alkaline phosphatase) downstream of the programmable hydraulic resistor.

Embedded Multimaterial Extrusion Bioprinting.

Embedded extrusion bioprinting allows for the generation of complex structures that otherwise cannot be achieved with conventional layer-by-layer deposition from the bottom, by overcoming the limits imposed by gravitational force. By taking advantage of a hydrogel bath, serving as a sacrificial printing environment, it is feasible to extrude a bioink in freeform until the entire structure is deposited and crosslinked. The bioprinted structure can be subsequently released from the supporting hydrogel and used for further applications. Combining this advanced three-dimensional (3D) bioprinting technique with a multimaterial extrusion printhead setup enables the fabrication of complex volumetric structures built from multiple bioinks. The work described in this paper focuses on the optimization of the experimental setup and proposes a workflow to automate the bioprinting process, resulting in a fast and efficient conversion of a virtual 3D model into a physical, extruded structure in freeform using the multimaterial embedded bioprinting system. It is anticipated that further development of this technology will likely lead to widespread applications in areas such as tissue engineering, pharmaceutical testing, and organs-on-chips.

Ondansetron inhibits the analgesic effects of tramadol: a possible 5-HT(3) spinal receptor involvement in acute pain in humans.

UNLABELLED: To investigate a possible antinociceptive role of serotonin receptor subtype 3 (5-HT(3)), we evaluated the effects of a coadministration of ondansetron, a 5-HT(3) selective antagonist, and tramadol, a central analgesic dependent on enhanced serotonergic transmission. Fifty-nine patients undergoing ear, throat, and nose surgery, using tramadol for 24-h postoperative patient-controlled analgesia (bolus = 30 mg; lockout interval = 10 min) were randomly allocated either to a group receiving ondansetron continuous infusion (1 mg. mL(-1). h(-1)) for postoperative nausea and vomiting (Group O) or to a control group receiving saline (Group T). Pain and vomiting scores and tramadol consumption were evaluated at 4, 8, 12, and 24 h. Pain scores were never >4, according to a 0-10 numerical rating scale, in both groups. Group O required significantly larger doses of tramadol at 4 h (213 versus 71 mg, P < 0.001), 8 h (285 versus 128 mg, P < 0.002), and 12 h (406 versus 190 mg, P < 0.002). Vomiting scores were higher in Group O at 4 h (P < 0.05) and 8 h (P = 0.05). We conclude that ondansetron reduced the overall analgesic effect of tramadol, probably blocking spinal 5-HT(3) receptors. IMPLICATIONS: Serotonin is an important neurotransmitter of the descending pathways that down-modulate spinal nociception. In postoperative pain, ondansetron, a selective 5-HT(3) receptor antagonist, increased the analgesic dose of tramadol. We suggest that, when antagonized for antiemetic purpose, 5-HT(3) receptors foster nociception, because of their site-dependent action.