Biophysical and Biochemical Regulation of Cell Dynamics in Magnetically Assembled Cellular Structures.

Soluble signaling molecules and extracellular matrix (ECM) regulate cell dynamics in various biological processes. Wound healing assays are widely used to study cell dynamics in response to physiological stimuli. However, traditional scratch-based assays can damage the underlying ECM-coated substrates. Here, we use a rapid, non-destructive, label-free magnetic exclusion technique to form annular aggregates of bronchial epithelial cells on tissue-culture treated (TCT) and ECM-coated surfaces within 3 h. The cell-free areas enclosed by the annular aggregates are measured at different times to assess cell dynamics. The effects of various signaling molecules, including epidermal growth factor (EGF), oncostatin M, and interleukin 6, on cell-free area closures are investigated for each surface condition. Surface characterization techniques are used to measure the topography and wettability of the surfaces. Further, we demonstrate the formation of annular aggregates on human lung fibroblast-laden collagen hydrogel surfaces, which mimic the native tissue architecture. The cell-free area closures on hydrogels indicate that the substrate properties modulate EGF-mediated cell dynamics. The magnetic exclusion-based assay is a rapid and versatile alternative to traditional wound healing assays.

SARS-CoV-2 detection with aptamer-functionalized gold nanoparticles.

A rapid detection test for SARS-CoV-2 is urgently required to monitor virus spread and containment. Here, we describe a test that uses nanoprobes, which are gold nanoparticles functionalized with an aptamer specific to the spike membrane protein of SARS-CoV-2. An enzyme-linked immunosorbent assay confirms aptamer binding with the spike protein on gold surfaces. Protein recognition occurs by adding a coagulant, where nanoprobes with no bound protein agglomerate while those with sufficient bound protein do not. Using plasmon absorbance spectra, the nanoprobes detect 16 nM and higher concentrations of spike protein in phosphate-buffered saline. The time-varying light absorbance is examined at 540 nm to determine the critical coagulant concentration required to agglomerates the nanoprobes, which depends on the protein concentration. This approach detects 3540 genome copies/µl of inactivated SARS-CoV-2.

Fibroblasts Accelerate Formation and Improve Reproducibility of 3D Cellular Structures Printed with Magnetic Assistance.

Fibroblasts (mouse, NIH/3T3) are combined with MDA-MB-231 cells to accelerate the formation and improve the reproducibility of 3D cellular structures printed with magnetic assistance. Fibroblasts and MDA-MB-231 cells are cocultured to produce 12.5 : 87.5, 25 : 75, and 50 : 50 total population mixtures. These mixtures are suspended in a cell medium containing a paramagnetic salt, Gd-DTPA, which increases the magnetic susceptibility of the medium with respect to the cells. A 3D monotypic MDA-MB-231 cellular structure is printed within 24 hours with magnetic assistance, whereas it takes 48 hours to form a similar structure through gravitational settling alone. The maximum projected areas and circularities, and cellular ATP levels of the printed structures are measured for 336 hours. Increasing the relative amounts of the fibroblasts mixed with the MDA-MB-231 cells decreases the time taken to form the structures and improves their reproducibility. Structures produced through gravitational settling have larger maximum projected areas and cellular ATP, but are deemed less reproducible. The distribution of individual cell lines in the cocultured 3D cellular structures shows that printing with magnetic assistance yields 3D cellular structures that resemble in vivo tumors more closely than those formed through gravitational settling. The results validate our hypothesis that (1) fibroblasts act as a "glue" that supports the formation of 3D cellular structures, and (2) the structures are produced more rapidly and with greater reproducibility with magnetically assisted printing than through gravitational settling alone. Printing of 3D cellular structures with magnetic assistance has applications relevant to drug discovery, lab-on-chip devices, and tissue engineering.

Label-Free Magnetic-Field-Assisted Assembly of Layer-on-Layer Cellular Structures.

Controlled cell assembly is essential for fabricating in vitro 3D models that mimic the physiology of in vivo cellular architectures. Whereas tissue engineering techniques often rely on intrusive magnetic nanoparticles placed in cells and hydrogel encapsulation of cells to produce multilayered cellular constructs, we describe a high-throughput, label-free, and scaffold-free magnetic field-guided technique that assembles cells into a layered aggregate. An inhomogeneous magnetic field influences the diamagnetic cells suspended in a paramagnetic culture medium. Driven by the magnetic susceptibility difference and the field gradient, the cells are displaced toward the region of lowest field strength. Two cell lines are used to demonstrate the sequential assembly of layer-on-layer aggregates in microwells within 6 h. The effect of magnet size on the assembly dynamics is characterized and a microwell size criterion for the highest cell aggregation provided. Label-free magnetic-field-assisted assembly is relevant for on-demand scalable biofabrication of complex layered structures. Potential applications include drug discovery, developmental biology, lab-on-chip devices, and cancer research.

Gadopentatic acid affects in vitro proliferation and doxorubicin response in human breast adenocarcinoma cells.

Contrasting agents (CAs) that are administered to patients during magnetic resonance imaging to facilitate tumor identification are generally considered harmless. However, gadolinium (Gd) based contrast agents can be retained in the body, inflicting specific cell line cytotoxicity. We investigate the effect of Gadopentatic acid (Gd-DTPA) on human breast adenocarcinoma MCF-7 cells. These cells exhibit a toggle switch response: exposure to 0.1 and 1 mM concentrations of Gd-DTPA enhances proliferation, which is hindered at a higher 10 mM concentration. Proliferation is enhanced when cells transition to 3D morphologies in post confluent conditions. The proliferation dependence on the concentration of CA is absent for Hs 578T and MDA-MB-231 triple negative cell lines. MCF-7 cells reveal a double toggle switch related to the expression of VEGF, which goes through high-low-high downregulation when cells are exposed to 0.1, 1, and 10 mM Gd-DTPA, respectively. Finally, doxorubicin drug response is assessed, which also reveals a double toggle switch behavior, where drug cytotoxicity exhibits a nonlinear dependence on the CA concentration. A toggle switch in cell characteristics that are exposed to 1 mM of Gd-DTPA amplifies the importance of this threshold, affecting several cell behaviors if surpassed. This work emphasizes the important effects that CAs can have on cells, specifically Gd-DTPA on MCF-7 cells, and the implications for cell growth and drug response during clinical and synthetic biology procedures.

3D cellular structures and co-cultures formed through the contactless magnetic manipulation of cells on adherent surfaces.

A magnet array is employed to manipulate diamagnetic cells that are contained in paramagnetic medium to demonstrate for the first time the contactless bioprinting of three-dimensional (3D) cellular structures and co-cultures of breast cancer MCF-7 and endothelial HUVEC at prescribed locations on tissue culture treated well plates. Sequential seeding of different cell lines and the spatial displacement of the magnet array creates co-cultured cellular structures within a well without using physically intrusive well inserts. Both monotypic and co-culture experiments produce morphologically rich 3D cell structures that are otherwise absent in regular monolayer cell cultures. The magnetic contactless bioprinting of cells provides further insight into cell behaviour, invasion strategies and transformations that are useful for potential applications in drug screening, 3D cell culture formation and tissue engineering.

Magnetic Printing of a Biosensor: Inexpensive Rapid Sensing To Detect Picomolar Amounts of Antigen with Antibody-Functionalized Carbon Nanotubes.

When an antibody (Ab) is immobilized on its surface, a carbon nanotube (CNT) becomes a biosensor that detects the corresponding antigen (Ag) because Ag-Ab complexes formed on the CNT surface moderate the current flow through it. We synthesized a biological ink containing CNTs that are twice functionalized, first with magnetic nanoparticles and thereafter with the anti-c-Myc monoclonal Ab. The ink is pipetted and dynamically self-organized by an external magnetic field into a dense electrically conducting sensor strip that measures the decrease in current when a sample containing c-Myc Ag is deposited on it. Prototypes are rapidly fabricated materials that cost less than 20 cents (Canadian) for each sensor. With larger current decreases due to real-time specific Ag-Ab binding for higher c-Myc concentrations, the biosensor distinguishes between picomolar c-Myc concentrations within a minute, offering proof of concept of a simple, rapid, economical, and sensitive method to detect specific molecules recognizable by Abs.

Nickel oxide nanotube synthesis using multiwalled carbon nanotubes as sacrificial templates for supercapacitor application.

A novel approach for the fabrication of nickel oxide nanotubes based on multiwalled carbon nanotubes as a sacrificial template is described. Electroless deposition is employed to deposit nickel onto carbon nanotubes. The subsequent annealing of the product in the presence of air oxidizes nickel to nickel oxide, and carbon is released as gaseous carbon dioxide, leaving behind nickel oxide nanotubes. Electron microscopy and elemental mapping confirm the formation of nickel oxide nanotubes. New chelating polyelectrolytes are used as dispersing agents to achieve high colloidal stability for both the nickel-coated carbon nanotubes and the nickel oxide nanotubes. A gravimetric specific capacitance of 245.3 F g-1 and  an areal capacitance of 3.28 F cm-2 at a scan rate of 2 mV s-1 is achieved, with an electrode fabricated using nickel oxide nanotubes as the active element with a mass loading of 24.1 mg cm-2.

Tailoring Material Stiffness by Filler Particle Organization.

In the context of emerging methods to control particle organization in particle-matrix composite materials, we explore, using finite element analysis, how to modulate the material bulk mechanical stiffness. Compared to a composite containing randomly distributed particles, material stiffness is enhanced 100-fold when filler particles are aligned into linear chains lying parallel to the loading direction. In contrast, chains aligned perpendicular to that direction produce negligible stiffness change. These outcomes reveal how zigzag chains, which provide intermediate results, can modulate stiffness. The stiffness decreases gradually with increasing zigzag angle θ over a range spanning 2 orders of magnitude.

High gradient magnetic field microstructures for magnetophoretic cell separation.

Microfluidics has advanced magnetic blood fractionation by making integrated miniature devices possible. A ferromagnetic microstructure array that is integrated with a microfluidic channel rearranges an applied magnetic field to create a high gradient magnetic field (HGMF). By leveraging the differential magnetic susceptibilities of cell types contained in a host medium, such as paramagnetic red blood cells (RBCs) and diamagnetic white blood cells (WBCs), the resulting HGMF can be used to continuously separate them without attaching additional labels, such as magnetic beads, to them. We describe the effect of these ferromagnetic microstructure geometries have on the blood separation efficacy by numerically simulating the influence of microstructure height and pitch on the HGMF characteristics and resulting RBC separation. Visualizations of RBC trajectories provide insight into how arrays can be optimized to best separate these cells from a host fluid. Periodic microstructures are shown to moderate the applied field due to magnetic interference between the adjacent teeth of an array. Since continuous microstructures do not similarly weaken the resultant HGMF, they facilitate significantly higher RBC separation. Nevertheless, periodic arrays are more appropriate for relatively deep microchannels since, unlike continuous microstructures, their separation effectiveness is independent of depth. The results are relevant to the design of microfluidic devices that leverage HGMFs to fractionate blood by separating RBCs and WBCs.

Printing Three-Dimensional Heterogeneities in the Elastic Modulus of an Elastomeric Matrix.

We present a rapid and controllable method to create microscale heterogeneities in the 3D stiffness of a soft material by printing patterns with a ferrofluid ink. An ink droplet moved through a liquid polydimethylsiloxane (PDMS) volume using an externally applied magnetic field sheds clusters of magnetic nanoparticles (MNPs) in its wake. By varying the field spatiotemporally, a well-defined three-dimensional curvilinear feature is printed that contains MNP clusters. Subsequent cross-linking of the PDMS preserves the feature in place after the magnetic field is removed. Since the ferrofluid ink interferes with the cross-linking of PDMS, a 3D print containing ink density variations leads to corresponding spatial deviations in the elastic modulus of the matrix. The modulus is mapped in the experiments with atomic force microscopy. This rapid method to print 3D heterogeneities in soft matter promises the ability to mimic mechanical variations that occur in natural biomaterials.

Nickel Nanoparticles Entangled in Carbon Nanotubes: Novel Ink for Nanotube Printing.

We report the serendipitous discovery of a rapid and inexpensive method to attach nanoscale magnetic chaperones to carbon nanotubes (CNTs). Nickel nanoparticles (NiNPs) become entangled in CNTs after both are dispersed in kerosene by sonication and form conjugates. An externally applied magnetic field manipulates the resulting CNTs-NiNP ink without NiNP separation, allowing us to print an embedded circuit in an elastomeric matrix and fabricate a strain gage and an oil sensor. The new method to print a circuit in a soft material using an NiNP-CNT ink is more rapid and inexpensive than the complex physical and chemical means typically used to magnetize CNTs.

In Situ 3D Label-Free Contactless Bioprinting of Cells through Diamagnetophoresis.

Using whole blood, we demonstrate the first realization of a novel macroscale, contactless, label-free method to print in situ three-dimensional (3D) cell assemblies of different morphologies and sizes. This novel bioprinting method does not use nozzles that can contaminate the cell suspension, or to which cells can adhere. Instead, we utilize the intrinsic diamagnetic properties of whole blood cells to magnetically manipulate them in situ in a nontoxic paramagnetic medium, creating (a) rectangular bar, (b) three-pointed star, and (c) spheroids of varying sizes. We envision the technique to be transferable to other cell lines, with potential applications in tissue engineering and drug screening.

Changing the magnetic properties of microstructure by directing the self-assembly of superparamagnetic nanoparticles.

Magnetic nanoparticles (MNPs) in a liquid dispersion can be organized through controlled self-assembly by applying an external magnetic field that regulates inter-particle interactions. Thus, micro- and nanostructures of desired morphology and superlattice geometry that show emergent magnetic properties can be fabricated. We describe how superferromagnetism, which is a specific type of emergence, can be produced. Here, superparamagnetic nanoparticles that show no individual residual magnetization are organized into structures with substantial residual magnetization that behave as miniature permanent magnets. We investigate the emergence of superferromagnetism in an idealized system consisting of two MNPs, by considering the influence that interparticle magnetostatic interactions have on the dynamics of the magnetic moments. We use this model to illustrate the design principles for self-assembly in terms of the choice of material and MNP particle size. We simulate the dynamics of the interacting magnetic moments by applying the stochastic Landau-Lifshitz-Gilbert equation to verify our principles. The findings enable a method to pattern material magnetization with submicron resolution, a useful feature that has potential applications for magnetic recording and microfluidic particle traps. The analysis also yields useful empirical generalizations that could facilitate other theoretical developments.

Thermal AND Gate Using a Monolayer Graphene Nanoribbon.

The first ever implementation of a thermal AND gate, which performs logic calculations with phonons, is presented using two identical thermal diodes composed of asymmetric graphene nanoribbons (GNRs). Employing molecular dynamics simulations, the characteristics of this AND gate are investigated and compared with those for an electrical AND gate. The thermal gate mechanism originates through thermal rectification due to asymmetric phonon boundary scattering in the two diodes, which is only effective at the nanoscale and at the temperatures much below the room temperature. Due to the high phonon velocity in graphene, the gate has a fast switching time of ≈100 ps.

Patterning the Stiffness of Elastomeric Nanocomposites by Magnetophoretic Control of Cross-linking Impeder Distribution.

We report a novel method to pattern the stiffness of an elastomeric nanocomposite by selectively impeding the cross-linking reactions at desired locations while curing. This is accomplished by using a magnetic field to enforce a desired concentration distribution of colloidal magnetite nanoparticles (MNPs) in the liquid precursor of polydimethysiloxane (PDMS) elastomer. MNPs impede the cross-linking of PDMS; when they are dispersed in liquid PDMS, the cured elastomer exhibits lower stiffness in portions containing a higher nanoparticle concentration. Consequently, a desired stiffness pattern is produced by selecting the required magnetic field distribution a priori. Up to 200% variation in the reduced modulus is observed over a 2 mm length, and gradients of up to 12.6 MPa·mm-1 are obtained. This is a significant improvement over conventional nanocomposite systems where only small unidirectional variations can be achieved by varying nanoparticle concentration. The method has promising prospects in additive manufacturing; it can be integrated with existing systems thereby adding the capability to produce microscale heterogeneities in mechanical properties.

Thermal rectification in a polymer-functionalized single-wall carbon nanotube.

Thermal rectification occurs when heat current through a material is favored in one direction but not in the opposite direction. These materials, often called thermal diodes, have the potential to perform logic calculations with phonons. Rectification obtained with existing material systems is either too minor or too difficult to implement practically. Hence, we present a scheme to enable higher rectification using a single-wall carbon nanotube (SWCNT) that is covalently functionalized near one end with polyacetylene (PA) chains. This composite structure allows rectification R up to 204%, which is higher than the values reported for SWCNTs. Here, [Formula: see text], where [Formula: see text] and [Formula: see text] are the heat currents for forward and reverse bias, respectively. The interatomic interactions in the SWCNT-PA nanocomposite are nonlinear, i.e., they are anharmonic, which is a requirement for thermal rectification. Through atomistic simulations, we identify two additional conditions to accomplish thermal rectification at the nanoscale, namely, (1) structural asymmetry, and (2) that the influence of this asymmetry on thermal transport is temperature dependent. The optimum temperature difference to achieve the highest thermal rectification with the structure is 40-80 K.

Communication: A tractable design for a thermal transistor.

We propose a conceptual design for a logic device that is the thermal analog of a transistor. It has fixed hot (emitter) and cold (collector) temperatures, and a gate controls the heat current. Thermal logic could be applied for thermal digital computing, enhance energy conservation, facilitate thermal rheostats, and enable the transport of phononic data. We demonstrate such a device using molecular dynamics simulations that consider thermal transport across hot and cold solid Si regions that seal water within them. Changes in the hot side, or emitter, heat current are linear with respect to varying gate temperature but the corresponding variation in the collector current is nonlinear. This nonlinear variation in collector current defines the ON and OFF states of the device. In its OFF state, the thermal conductivity of the device is positive. In the ON state, however, more heat is extracted through the cold terminal than is provided at the hot terminal due to the intervention of the base terminal. This makes it possible to alter the transport factor by varying the gate conditions. When the device is ON, the transport factor is greater than unity, i.e., more heat is rejected at the collector than is supplied to the emitter.

Soft Polymer Magnetic Nanocomposites: Microstructure Patterning by Magnetophoretic Transport and Self-Assembly.

A method to produce and pattern magnetic microstructure in a soft-polymer matrix is demonstrated. An externally applied magnetic field is used to influence the dynamics of magnetophoretic transport and dipolar self-assembly of magnetic nanoparticle clusters in the liquid precursor of poly-dimethylsiloxane (PDMS). Magnetic nanoparticles agglomerate by an interplay of van der Waals forces and dipolar interactions to form anisotropic clusters. These clusters are concentrated on a substrate by magnetophoresis, wherein they self-organize by dipolar interactions to form microscopic filaments. The polymer is cured in the presence of the magnetic field to preserve the microstructure shape. The externally applied magnetic field and its gradient are the two main control variables of interest when considering magnetic control during nanoparticle self-assembly. Their influence on microstructure geometry is investigated through correlations with the height of a characteristic self-assembled filament, fraction of the substrate area covered by the microstructure and its shape anisotropy. These relations enable a priori design.