A fast, low-cost, robust and high-throughput method for viral nucleic acid isolation based on NAxtra magnetic nanoparticles.

The year of 2020 was profoundly marked by a global pandemic caused by a strain of coronavirus named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent of coronavirus disease 2019 (COVID-19). To control disease spread, a key strategy adopted by many countries was the regular testing of individuals for infection. This led to the rapid development of diagnostic testing technologies. In Norway, within a week, our group developed a test kit to quickly isolate viral RNA and safely detect SARS-CoV-2 infection with sensitivity comparable to available kits. Herein, the procedure employed for the detection of SARS-CoV-2 in swab samples from patients using the NTNU-COVID-19 test kit is described in detail. This procedure, based on NAxtra magnetic nanoparticles and an optimized nucleic acid extraction procedure, is robust, reliable, and straightforward, providing high-quality nucleic acids within 14 min. The NAxtra protocol is adaptable and was further validated for extraction of DNA and RNA from other types of viruses. A comparison of the protocol on different liquid handling systems is also presented. Due to the simplicity and low cost of this method, implementation of this technology to diagnose virus infections on a clinical setting would benefit health care systems, promoting sustainability.

Modulating acrylic acid content of nanogels for drug delivery & biocompatibility studies.

Dual stimuli-responsive nanogels (NGs) have gained popularity in the field of bio medicine due to their versatile nature of applicability. Poly(N-isopropylacrylamide)-co-poly(acrylic acid) (pNIPAm-pAAc)-based NGs provide such dual stimuli-response with pNIPAm and pAAc providing thermal and pH-based responses, respectively. Studying the growth of these NGs, as well as, understanding the effect of the incorporation of pAAc in the NG matrix, is important in determining the physico-chemical properties of the NG. Studies have been conducted investigating the effect of increasing pAAc content in the NGs, however, these are not detailed in understanding its effects on the physico-chemical properties of the pNIPAm-pAAc-based NGs. Also, the biocompatibility of the NGs have not been previously reported using human whole blood model. Herein, we report the effect of different reaction parameters, such as surfactant amount and reaction atmosphere, on the growth of pNIPAm-pAAc-based NGs. It is shown that the size of the NGs can be precisely controlled from ~130 nm to ~400 nm, by varying the amount of surfactant and the reaction atmosphere. The effect of increasing incorporation of pAAc in the NG matrix on its physico-chemical properties has been investigated. The potential of these NGs as drug delivery vehicles is investigated by conducting loading and release studies of a model protein drug, cytochrome C (Cyt C) from the NGs at temperature above the volume phase transition temperature (VPTT) and acidic pH. An ex vivo human whole blood model was used to investigate biocompatibility of the NGs by quantifying inflammatory responses during NG exposure. The NGs did not induce any significant production of chemokine IL-8 or pro-inflammatory cytokines (IL-1ß, IL-6, TNF-α), and the cell viability in human whole blood was maintained during 4 h exposure. The NGs did neither activate the complement system, as determined by low Terminal Complement Complex (TCC) activation and Complement Receptor 3 (CR3) activation assays, thereby overall suggesting that the NGs could be potential candidates for biomedical applications.

Magnetic Nanoparticles to Unique DNA Tracers: Effect of Functionalization on Physico-chemical Properties.

To monitor and manage hydrological systems such as brooks, streams, rivers, the use of tracers is a well-established process. Limited number of potential tracers such as salts, isotopes and dyes, make study of hydrological processes a challenge. Traditional tracers find limited use due to lack of multiplexed, multipoint tracing and background noise, among others. In this regard, DNA based tracers possess remarkable advantages including, environmentally friendly, stability, and high sensitivity in addition to showing great potential in the synthesis of ideally unlimited number of unique tracers capable of multipoint tracing. To prevent unintentional losses in the environment during application and easy recovery for analysis, we hereby report DNA encapsulation in silica containing magnetic cores (iron oxide) of two different shapes-spheres and cubes. The iron oxide nanoparticles having size range 10-20 nm, have been synthesized using co-precipitation of iron salts or thermal decomposition of iron oleate precursor in the presence of oleic acid or sodium oleate. Physico-chemical properties such as size, zeta potential, magnetism etc. of the iron oxide nanoparticles have been optimized using different ligands for effective binding of dsDNA, followed by silanization. We report for the first time the effect of surface coating on the magnetic properties of the iron oxide nanoparticles at each stage of functionalization, culminating in silica shells. Efficiency of encapsulation of three different dsDNA molecules has been studied using quantitative polymerase chain reaction (qPCR). Our results show that our DNA based magnetic tracers are excellent candidates for hydrological monitoring with easy recoverability and high signal amplification.

Synthesis, Characterization and Drug Loading of Multiresponsive p[NIPAm-co-PEGMA] (core)/p[NIPAm-co-AAc] (Shell) Nanogels with Monodisperse Size Distributions.

We report the synthesis and properties of temperature- and pH-responsive p([NIPAm-co-PEGMA] (core)/[NIPAm-co-AAc] (shell)) nanogels with narrow size distributions, tunable sizes and increased drug loading efficiencies. The core-shell nanogels were synthesized using an optimized two-stage seeded polymerization methodology. The core-shell nanogels show a narrow size distribution and controllable physico-chemical properties. The hydrodynamic sizes, charge distributions, temperature-induced volume phase transition behaviors, pH-responsive behaviors and drug loading capabilities of the core-shell nanogels were investigated using transmission electron microscopy, zeta potential measurements, dynamic light scattering and UV-Vis spectroscopy. The size of the core-shell nanogels was controlled by polymerizing NIPAm with crosslinker poly(ethylene glycol) dimethacrylate (PEGDMA) of different molecular weights (Mn-200, 400, 550 and 750 g/mol) during the core synthesis. It was found that the swelling/deswelling kinetics of the nanogels was sharp and reversible; with its volume phase transition temperature in the range of 40⁻42 °C. Furthermore, the nanogels loaded with l-3,4-dihydroxyphenylalanine (L-DOPA), using a modified breathing-in mechanism, showed high loading and encapsulation efficiencies, providing potential possibilities of such nanogels for biomedical applications.

The Influence of Differently Shaped Gold Nanoparticles Functionalized with NIPAM-Based Hydrogels on the Release of Cytochrome C.

Here, we report the synthesis and functionalization of five different shapes of Au nanoparticles (NPs), namely nanorods, tetrahexahedral, bipyramids, nanomakura, and spheres with PEG and poly (N-isopropylacrylamide)-acrylic acid (pNIPAm-AAc) hydrogels. The anisotropic NPs are synthesized using seed-mediated growth in the presence of silver. The NPs have been characterized using Dynamic Light Scattering (DLS), zeta potential measurements, UV-Visible spectrophotometry (UV-Vis), and Scanning Transmission Electron Microscopy (S(T)EM). Cyt C was loaded into the PEG-hydrogel-coated AuNPs using a modified breathing-in method. Loading efficiencies (up to 80%), dependent on particle geometry, concentration, and hydrogel content, were obtained. Release experiments conducted at high temperature (40 °C) and acidic pH (3) showed higher release for larger sizes of PEG-hydrogel-coated AuNPs, with temporal transition from spherical to thin film release geometry. AuNP shape, size, number density, and hydrogel content are found to influence the loading as well as release kinetics of Cyt C from these systems.