Printing of In Situ Functionalized Mesoporous Silica with Digital Light Processing for Combinatorial Sensing.

Combinatorial sensing is especially important in the context of modern drug development to enable fast screening of large data sets. Mesoporous silica materials offer high surface area and a wide range of functionalization possibilities. By adding structural control, the combination of structural and functional control along all length scales opens a new pathway that permits larger amounts of analytes being tested simultaneously for complex sensing tasks. This study presents a fast and simple way to produce mesoporous silica in various shapes and sizes between 0.27-6 mm by using light-induced sol-gel chemistry and digital light processing (DLP). Shape-selective functionalization of mesoporous silica is successfully carried out either after printing using organosilanes or in situ while printing through the use of functional mesopore template for the in situ functionalization approach. Shape-selective adsorption of dyes is shown as a demonstrator toward shape selective screening of potential analytes.

Sensing Levofloxacin with an RNA Aptamer as a Bioreceptor.

To combat the growing threat of antibiotic resistance, environmental testing for antibiotic contamination is gaining an increasing role. This study aims to develop an easy-to-use assay for the detection of the fluoroquinolone antibiotic levofloxacin. Levofloxacin is used in human and veterinary medicine and has been detected in wastewater and river water. An RNA aptamer against levofloxacin was selected using RNA Capture-SELEX. The 73 nt long aptamer folds into three stems with a central three-way junction. It binds levofloxacin with a Kd of 6 µM and discriminates the closely related compound ciprofloxacin. Furthermore, the selection process was analyzed using a next-generation sequencing approach to better understand the sequence evolution throughout the selection. The aptamer was used as a bioreceptor for the development of a lateral flow assay. The biosensor exploited the innate characteristic of RNA Capture-SELEX to select aptamers that displace a complementary DNA oligonucleotide upon ligand binding. The lateral flow assay achieved a limit of visual detection of 100 µM. While the sensitivity of this assay constrains its immediate use in environmental testing, the present study can serve as a template for the selection of RNA aptamer-based biosensors.

Device for measuring part adhesion in FFF process.

The adhesion of parts to the build surface plays a central role in the Fused Filament Fabrication (FFF) process. Without sufficient adhesion, the part will deform (so called warping) due to thermal shrinkage, so that no defined geometries can be created. Nevertheless, there is no established method to measure the adhesion of printed parts and therefore it is not possible to targeted improve it. This article presents a measurement method based on the DIN EN 28510-1 standard and a corresponding test device which makes it possible to identify the optimum build surface for a filament and also to improve the process parameters in a targeted manner. The test device combines a FFF printer with a measuring unit so that all common filaments can be tested close to the process up to a processing temperature of 400 °C in the nozzle and around 150 °C on the build platform. The test device uses only open-source parts and software and costs about 1700€.

Carbohydrate binding module-fused antibodies improve the performance of cellulose-based lateral flow immunoassays.

Since the pandemic outbreak of Covid-19 in December 2019, several lateral flow assay (LFA) devices were developed to enable the constant monitoring of regional and global infection processes. Additionally, innumerable lateral flow test devices are frequently used for determination of different clinical parameters, food safety, and environmental factors. Since common LFAs rely on non-biodegradable nitrocellulose membranes, we focused on their replacement by cellulose-composed, biodegradable papers. We report the development of cellulose paper-based lateral flow immunoassays using a carbohydrate-binding module-fused to detection antibodies. Studies regarding the protein binding capacity and potential protein wash-off effects on cellulose paper demonstrated a 2.7-fold protein binding capacity of CBM-fused antibody fragments compared to the sole antibody fragment. Furthermore, this strategy improved the spatial retention of CBM-fused detection antibodies to the test area, which resulted in an enhanced sensitivity and improved overall LFA-performance compared to the naked detection antibody. CBM-assisted antibodies were validated by implementation into two model lateral flow test devices (pregnancy detection and the detection of SARS-CoV-2 specific antibodies). The CBM-assisted pregnancy LFA demonstrated sensitive detection of human gonadotropin (hCG) in synthetic urine and the CBM-assisted Covid-19 antibody LFA was able to detect SARS-CoV-2 specific antibodies present in serum. Our findings pave the way to the more frequent use of cellulose-based papers instead of nitrocellulose in LFA devices and thus potentially improve the sustainability in the field of POC diagnostics.

Analysis of free chlorine in aqueous solution at very low concentration with lateral flow tests.

Test strips are convenient tools for rapid, semi-quantitative analysis of a variety of parameters by dipping them for a few seconds in a sample solution followed by a simple colorimetric read-out. Their sensitivity is mainly determined by the reactivity of the test dyes on the reaction zone and is not sufficient for some applications. The detection limit of commercially available free chlorine test strips, for example, is at present not low enough to confirm the absence of this analyte as disinfectant in rinsing solutions after disinfection or to control required residual amounts of chlorine in drinking water. Therefore, we developed a user-friendly lateral flow test which is capable to detect very low amounts of free chlorine. The latter relies on a larger sample volume passing the reaction zone as compared to simple dip test strips. An amount of as low as 0.05 ppm chlorine can, however, only be detected if oxidation stable flow test substrates are used. The eventually developed flow test reaches a 10x higher sensitivity than a commercial dip test. The result is obtained within 4-5 min flow time, whereby no action is required by the user during this analysis time.

Characterization and Inkjet Printing of an RNA Aptamer for Paper-Based Biosensing of Ciprofloxacin.

The excessive use of antibiotics in food-producing animals causes a steady rise of multiple antibiotic resistance in foodborne bacteria. Next to sulfonamides, the most common antibiotics groups are fluoroquinolones, aminoglycosides, and ß-lactams. Therefore, there is a need for a quick, efficient, and low-cost detection procedure for antibiotics. In this study, we propose an inkjet-printed aptamer-based biosensor developed for the detection of the fluoroquinolone ciprofloxacin. Due to their extraordinary high affinity and specificity, aptamers are already widely used in various applications. Here we present a ciprofloxacin-binding RNA aptamer developed by systematic evolution of ligands by exponential enrichment (SELEX). We characterized the secondary structure of the aptamer and determined the KD to 36 nM that allow detection of antibiotic contamination in a relevant range. We demonstrate that RNA aptamers can be inkjet-printed, dried, and resolved while keeping their functionality consistently intact. With this proof of concept, we are paving the way for a potential range of additional aptamer-based, printable biosensors.

Gravure printing for mesoporous film preparation.

This study presents gravure printing as a new strategy for rapid printing of ceramic mesoporous films and highlights its advantages over conventional mesoporous film preparation using evaporation induced self-assembly together with dip-coating. By varying the printing process parameters, the mesoporous coating thicknesses can be adjusted between 20 and 200 nm while maintaining a very high film homogeneity allowing the printing of ultrathin mesoporous films. Step gradients in film composition are accessible by consecutively printing two different "inks". Thereby, gravure printing is a much faster process than mesoporous single- and multilayer preparation using conventional dip-coating because lower amounts of solution are transferred and dissolution of previously deposited layers is avoided. The effect of printing process parameters on resulting film characteristics as well as the resulting mesoporous film's ionic accessibility is systematically investigated.

Combining Wax Printing with Hot Embossing for the Design of Geometrically Well-Defined Microfluidic Papers.

A simple, efficient, and repeatable combination of wax printing and hot embossing is reported. This combination yields microfluidic channels in paper, where fluid transport driven by paper-intrinsic capillary forces takes place inside the noncompressed areas, whereas embossed and wax-bonded areas serve as hydrophobic barriers laterally confining the fluid flow. Lab-made paper sheets first coated with a hydrophobic wax were hot-embossed with a tailor-made metal stamp. Both paper-intrinsic (e.g., grammage, fiber type) and paper-extrinsic parameters (e.g., embossing force) were studied for their influence on the geometry of the embossed structures and the resulting redistribution of the wax within the paper matrix. Embossing of wax-printed paper at temperatures above the wax melting point was completed within 15 s. Cotton linters papers required higher embossing forces than eucalyptus papers, which can be explained by their different intrinsic mechanical properties. In summary, both paper-intrinsic and paper-extrinsic parameters were found to have strong impact on resolution and reproducibility of the channels. All in all, the approach yields microfluidic channels in a fast and robust and reproducible manner with comparably low constrains on the precision of manufacturing parameters, such as embossing time, force, or temperature. Most importantly, embossing greatly reduces the lateral spreading of the wax as seen with melting approaches and therefore allows for a much higher feature density than the latter.