Micronit Microfluidics (Netherlands)

Microfluidics is a very attractive discipline for implementing more versatile high-throughput screening methods to improve enzymes. However, state-of-the-art microfluidics set-ups are mainly devoted to screening enzymes in solution, while screening of enzyme immobilization protocols using microfluidics is scarce. In this work, a microreactor device is designed, fabricated and applied to test different heterogeneous biocatalysts with transaminase activity. This microsystem set-up can be operated in two different modes: packed-bed (PBR) and fluidized (FBR) microreactors without exchanging the sample. Using the same sample to run the two modes, the fidelity of the comparison is increased between the two fluid dynamic regimes. Moreover, by testing different carriers under different modes, a Histagged transaminase is found from Pseudomonas fluorescens immobilized on agarose porous microbeads functionalized with cobalt-chelates and operated as FBR maximizes the STY, and minimizes the equilibration times. This device exemplifies the potential of microreactors for prototyping more efficient heterogeneous biocatalysts.

Understanding the complex landscape of protein interactions, especially those involving intrinsically disordered proteins (IDPs), is fundamental yet challenging due to their structural heterogeneity and flexibility. Traditional sequence-based homology methods frequently fall short in characterizing IDP functions and interactions. Here, we present a novel approach leveraging supervised and unsupervised machine learning techniques, focusing exclusively on the compositional features of proteins. An Edmond-Ogston-inspired mixing model can reliably predict the degree of survivin (BIRC5) binding as a function of peptide composition alone, revealing a first interesting connection with the composition diagrams of chemical thermodynamics. By representing protein sequences through their functional group compositions, we demonstrate that specific compositions robustly predict binding interactions with survivin, an important human protein in cellular regulation pathways. Experimental validation via peptide microarray confirms the predictive power of our simplified compositional model, independent of exact amino acid sequences. Extending this method across the human proteome, we identified distinct compositional signatures correlating with survivin interactions and revealed fine grained biologically meaningful functional clusters based on compositional similarity. Our findings suggest a compositional blueprint underpinning protein interactions, offering a powerful, simplified framework to decode complex biological networks.

This study demonstrates the use of degenerate primers in combination with CRISPR-based detection for high selectivity measurements of high-risk HPV types. We found that the E6 genes are more suitable for this purpose than the L1 gene, and the lbCas12a performs better than AsCas12a in differentiating between HPV types. Initial steps toward integrating this method into an easy-to-use device have been taken, with future research focusing on incorporating the entire process, including DNA extraction, target amplification, and target detection, into a single device. This advancement aims to make primary HPV screening feasible in low-resource settings, improving accessibility and early detection capabilities.

This hypothesis demonstrates that the efficiency of Loop-Mediated Isothermal Amplification (LAMP) for nucleic acid detection can be positively influenced by preconcentration of microbial cells onto hydrophobic paper surfaces. The mechanism of this model is based on the high attraction of microbes towards hydrophobic surfaces. Extensive studies have demonstrated that hydrophobic surfaces exhibit enhanced bacterial and fungal adhesion. The preconcentration approach by exploiting this inherent affinity of hydrophobic paper substrates enables the adherence of a greater number of target cells, resulting in a higher concentration of target templates for amplification directly from urine samples. In contrast to conventional methods, which often involve complex procedures, this approach offers a simpler, cost-effective, and user-friendly alternative. Moreover, the integration of cell adhesion, LAMP amplification, and signal readout within paper origami-based devices can provide a portable, robust, and highly efficient platform for rapid nucleic acid detection. This innovative hypothesis holds significant potential for point-of-care (POC) diagnostics and field surveillance applications. Further research and development in this field will advance the implementation of this technology, contributing to improved healthcare systems and public health outcomes.

Abstract Displacing viscous oil by water leads to poor displacement efficiency owing to the high mobility ratio and viscous fingering. Polymer injection increases oil recovery by reducing viscous fingering and improving sweep efficiency. We are showing how Alkali-Polymer (AP) flooding is substantially improving production of reactive viscous oil from a Romanian oil field. IFT measurements, coreflood and micro-model experiments were used to understand and optimize the physico-chemical processes leading to incremental oil recovery. Extensive IFT measurements were performed at different alkali and AP concentrations. In addition, phase behavior tests were done. Furthermore, micro-model experiments were used to elucidate effects at the pore-scale and as screening tool for which chemicals to use. Single and two-phase coreflood experiments helped defining the displacement efficiency on a core scale. Various sequences and concentrations of alkali and polymers were injected to reduce costs and maximize incremental recovery of the reactive viscous oil. IFT measurements showed that saponification (110 μmol/g saponifiable acids) at the oil-alkali solution interface is very effectively reducing the IFT. With time, the IFT is increasing owing to diffusion of the generated soaps away from the interface. Phase experiments confirmed that emulsions are formed initially. Micro-models revealed that injection of polymers or alkali only leads to limited incremental oil recovery over waterflooding. For alkali injection, oil is emulsified due to in-situ saponification at the edges of viscous fingers. AP injection after waterflooding is very effective. The emulsified oil at the edges of the viscous fingers is effectively dragged by the viscous fluid substantially increasing recovery. Corefloods confirmed the findings of the micromodels. In addition, the effect of di-valent cations for the selection of the polymer concentration was investigated. Water softening leads to significantly higher viscosity of the AP slug than non-softened brine. Reducing the polymer concentration to obtain the same viscosity as the polymer solution containing divalent cations resulted in similar displacement efficiency. Hence, significant cost savings can be realized for the field conditions, for which AP injection is planned after polymer injection. The results show that alkali solutions lead to initial low IFT of reactive viscous oil owing to soap generation at the oil-alkali solution interface increasing with time due to diffusion. Injecting alkali solutions into reactive viscous oil is not effective to reduce remaining oil saturation, a limited amount of oil is mobilized at the edges of viscous fingers. AP flooding of reactive viscous oil is substantially increasing incremental oil recovery. The reason is the effective dragging of the mobilized oil with the viscous fluid and associated exposure of additional oil to the alkali solutions. Furthermore, the economics of AP flooding projects can be substantially improved by adjusting the polymer concentration to the AP slug containing softened water.

Blood plasma separation is a prerequisite in numerous biomedical assays involving low abundance plasma-borne biomarkers and thus is the fundamental step before many bioanalytical steps. High-capacity refrigerated centrifuges, which have the advantage of handling large volumes of blood samples, are widely utilized, but they are bulky, non-transportable, and prohibitively expensive for low-resource settings, with prices starting at $1,500. On the other hand, there are low-cost commercial and open-source micro-centrifuges available, but they are incapable of handling typical clinical amounts of blood samples (2-10mL). There is currently no low-cost CE marked centrifuge that can process large volumes of clinical blood samples on the market. As a solution, we customised the rotor of a commercially available low-cost micro-centrifuge (~$125) using 3D printing to enable centrifugation of large clinical blood samples in resource poor-settings. Our custom adaptor ($15) can hold two 9 mL S-Monovette tubes and achieve the same separation performance (yield, cell count, hemolysis, albumin levels) as the control benchtop refrigerated centrifuge, and even outperformed the control in platelet separation by at least four times. This low-cost open-source centrifugation system capable of processing clinical blood tubes could be valuable to low-resource settings where centrifugation is required immediately after blood withdrawal for further testing.

Abstract Blood plasma separation is a prerequisite in numerous biomedical assays involving low abundance plasma-borne biomarkers and thus is the fundamental step before many bioanalytical steps. Conventionally, plasma separation is performed using high-capacity refrigerated centrifuges which have the advantage of handling large volume blood samples. These centrifuges are bulky, and prohibitively expensive for low-resource settings, with prices starting from $1,500. Although commercial and existing open-source micro-centrifuges are relatively low-cost, they cannot handle large volume blood samples. Microfluidic blood plasma separation also has been adopted by many researchers to enable low-cost plasma separation, however, these systems still present yield and purity issues for extremely low abundance biomarker detection such as the detection of various fractions of circulating cell-free DNA. To overcome this, we customised the rotor of a commercially available micro-centrifuge ∼$125) using fused filament fabrication to enable centrifugation of large clinical blood samples in resource poor-settings. Our designed adaptor ($15) can hold two 9 mL S-Monovette tubes and maintain the same separation performance (yield, cell count, hemolysis, albumin levels) as the control benchtop refrigerated centrifuge. This low-cost open-source centrifugation system capable of processing clinical blood tubes could be valuable to low-funded laboratories or low-resource settings where centrifugation is required immediately after blood withdrawal for further testing.

This study focuses on the development of a fluorocarbon-free directional silicon etching process, called CORE (Clear, Oxidize, Remove, and Etch) in which a switching sequence of SF6 and O2 is operated at room temperature. This distinguishes it from the old-fashioned room temperature and cryogenic mixed RIE processes as CORE enables a higher selectivity, creates pattern independency of etching profiles and works excellent at room temperature. The CORE process resembles the well-known SF6-based Bosch process, but the usual C4F8 inhibitor is replaced by O2 oxidation with self-limiting characteristics. Therefore the CORE result is similar to Bosch, however has the advantage of preventing the pile-up of fluorocarbon deposits at the topside of deep-etched or nano-sized features. At the same time, process drift is minimized as the reactor wall is staying perfectly clean. The CORE process has shown an excellent performance in high aspect ratio (3D) nanoscale structures with an accurate and controllable etch rate between 1 and 50 nm min−1 (and SiO2-selectivity of ca. 35) using the etch-tool in the RIE-mode. By adding the ICP source (DRIE-mode), a directional etch rate up to 1 μm min−1 (at 50 sccm SF6 flow) and selectivity >200 for SiO2 is possible.