Structural interactions in polymer-stabilized magnetic nanocomposites.

Superparamagnetic iron oxide nanoparticles (SPIONs) have attracted significant attention because of their nanoscale magnetic properties. SPION aggregates may afford emergent properties, resulting from dipole-dipole interactions between neighbors. Such aggregates can display internal order, with high packing fractions (>20%), and can be stabilized with block co-polymers (BCPs), permitting design of tunable composites for potential nanomedicine, data storage, and electronic sensing applications. Despite the routine use of magnetic fields for aggregate actuation, the impact of those fields on polymer structure, SPION ordering, and magnetic properties is not fully understood. Here, we report that external magnetic fields can induce ordering in SPION aggregates that affect their structure, inter-SPION distance, magnetic properties, and composite Tg. SPION aggregates were synthesized in the presence or absence of magnetic fields or exposed to magnetic fields post-synthesis. They were characterized using transmission electron microscopy (TEM), small angle X-ray scattering (SAXS), superconducting quantum interference device (SQUID) analysis, and differential scanning calorimetry (DSC). SPION aggregate properties depended on the timing of field application. Magnetic field application during synthesis encouraged preservation of SPION chain aggregates stabilized by polymer coatings even after removal of the field, whereas post synthesis application triggered subtle internal reordering, as indicated by increased blocking temperature (TB), that was not observed via SAXS or TEM. These results suggest that magnetic fields are a simple, yet powerful tool to tailor the structure, ordering, and magnetic properties of polymer-stabilized SPION nanocomposites.

Localized Plasmonic Heating for Single-Molecule DNA Rupture Measurements in Optical Tweezers.

To date, studies on the thermodynamic and kinetic processes that underlie biological function and nanomachine actuation in biological- and biology-inspired molecular constructs have primarily focused on photothermal heating of ensemble systems, highlighting the need for probes that are localized within the molecular construct and capable of resolving single-molecule response. Here we present an experimental demonstration of wavelength-selective, localized heating at the single-molecule level using the surface plasmon resonance of a 15 nm gold nanoparticle (AuNP). Our approach is compatible with force-spectroscopy measurements and can be applied to studies of the single-molecule thermodynamic properties of DNA origami nanomachines as well as biomolecular complexes. We further demonstrate wavelength selectivity and establish the temperature dependence of the reaction coordinate for base-pair disruption in the shear-rupture geometry, demonstrating the utility and flexibility of this approach for both fundamental studies of local (nanometer-scale) temperature gradients and rapid and multiplexed nanomachine actuation.

Cooperative control of a DNA origami force sensor.

Biomolecular systems are dependent on a complex interplay of forces. Modern force spectroscopy techniques provide means of interrogating these forces, but they are not optimized for studies in constrained environments as they require attachment to micron-scale probes such as beads or cantilevers. Nanomechanical devices are a promising alternative, but this requires versatile designs that can be tuned to respond to a wide range of forces. We investigate the properties of a nanoscale force sensitive DNA origami device which is highly customizable in geometry, functionalization, and mechanical properties. The device, referred to as the NanoDyn, has a binary (open or closed) response to an applied force by undergoing a reversible structural transition. The transition force is tuned with minor alterations of 1 to 3 DNA oligonucleotides and spans tens of picoNewtons (pN). The DNA oligonucleotide design parameters also strongly influence the efficiency of resetting the initial state, with higher stability devices (≳10 pN) resetting more reliably during repeated force-loading cycles. Finally, we show the opening force is tunable in real time by adding a single DNA oligonucleotide. These results establish the potential of the NanoDyn as a versatile force sensor and provide fundamental insights into how design parameters modulate mechanical and dynamic properties.

Cooperative control of a DNA origami force sensor.

Most biomolecular systems are dependent on a complex interplay of forces. Modern force spectroscopy techniques provide means of interrogating these forces. These techniques, however, are not optimized for studies in constrained or crowded environments as they typically require micron-scale beads in the case of magnetic or optical tweezers, or direct attachment to a cantilever in the case of atomic force microscopy. We implement a nanoscale force-sensing device using a DNA origami which is highly customizable in geometry, functionalization, and mechanical properties. The device, referred to as the NanoDyn, functions as a binary (open or closed) force sensor that undergoes a structural transition under an external force. The transition force is tuned with minor alterations of 1 to 3 DNA oligonucleotides and spans tens of picoNewtons (pN). This actuation of the NanoDyn is reversible and the design parameters strongly influence the efficiency of resetting the initial state, with higher stability devices (≳10 pN) resetting more reliably during repeated force-loading cycles. Finally, we show that the opening force can be adjusted in real time by the addition of a single DNA oligonucleotide. These results establish the NanoDyn as a versatile force sensor and provide fundamental insights into how design parameters modulate mechanical and dynamic properties.

DNA-caged nanoparticles via electrostatic self-assembly.

DNA-modified nanoparticles enable DNA sensing and therapeutics in nanomedicine and are also crucial for nanoparticle self-assembly with DNA-based materials. However, methods to conjugate DNA to nanoparticle surfaces are limited, inefficient, and lack control. Inspired by DNA tile nanotechnology, we demonstrate a new approach to nanoparticle modification based on electrostatic attraction between negatively charged DNA tiles and positively charged nanoparticles. This approach does not disrupt nanoparticle surfaces and leverages the programmability of DNA nanotechnology to control DNA presentation. We demonstrated this approach using a vareity of nanoparticles, including polymeric micelles, polystyrene beads, gold nanoparticles, and superparamagnetic iron oxide nanoparticles with sizes ranging from 5-20 nm in diameter. DNA cage formation was confirmed through transmission electron microscopy (TEM), neutralization of zeta potential, and a series of fluorescence experiments. DNA cages present "handle" sequences that can be used for reversible target attachment or self-assembly. Handle functionality was verified in solution, at the solid-liquid interface, and inside fixed cells, corresponding to applications in biosensing, DNA microarrays, and erasable immunocytochemistry. These experiments demonstrate the versatility of the electrostatic DNA caging approach and provide a new pathway to nanoparticle modification with DNA that will empower further applications of these materials in medicine and materials science.

Evaluating glioblastoma tumour sphere growth and migration in interaction with astrocytes using 3D collagen-hyaluronic acid hydrogels.

Glioblastoma (GB) is an astrocytic brain tumour with a low survival rate, partly because of its highly invasive nature. The GB tumour microenvironment (TME) includes its extracellular matrix (ECM), a variety of brain cell types, unique anatomical structures, and local mechanical cues. As such, researchers have attempted to create biomaterials and culture models that mimic features of TME complexity. Hydrogel materials have been particularly popular because they enable 3D cell culture and mimic TME mechanical properites and chemical composition. Here, we used a 3D collagen I-hyaluronic acid hydrogel material to explore interactions between GB cells and astrocytes, the normal cell type from which GB likely derives. We demonstrate three different spheroid culture configurations, including GB multi-spheres (i.e., GB and astrocyte cells in spheroid co-culture), GB-only mono-spheres cultured with astrocyte-conditioned media, and GB-only mono-spheres cultured with dispersed live or fixed astrocytes. Using U87 and LN229 GB cell lines and primary human astrocytes, we investigated material and experiment variability. We then used time-lapse fluorescence microscopy to measure invasive potential by characterizing the sphere size, migration capacity, and weight-averaged migration distance in these hydrogels. Finally, we developed methods to extract RNA for gene expression analysis from cells cultured in hydrogels. U87 and LN229 cells displayed different migration behaviors. U87 migration occurred primarily as single cells and was reduced with higher numbers of astrocytes in both multi-sphere and mono-sphere plus dispersed astrocyte cultures. In contrast, LN229 migration exhibited features of collective migration and was increased in monosphere plus dispersed astrocyte cultures. Gene expression studies indicated that the most differentially expressed genes in these co-cultures were CA9, HLA-DQA1, TMPRSS2, FPR1, OAS2, and KLRD1. Most differentially expressed genes were related to immune response, inflammation, and cytokine signalling, with greater influence on U87 than LN229. These data show that 3D in vitro hydrogel co-culture models can be used to reveal cell line specific differences in migration and to study differential GB-astrocyte crosstalk.

Oxidation of quantum dots encapsulated in block copolymer micelles as a function of polymer terminal charge.

Most high-quality quantum dots (QDs) are synthesized in the organic phase, and are often coated with polymers for use in aqueous biological environments. QDs can exhibit fluorescence losses during phase transfer, but evaluating underlying mechanisms (e.g., oxidation, surface etching, loss of colloidal stability) can be challenging because of variation in synthesis methods. Here, fluorescence stability of QDs encapsulated in block co-polymer (BCP) micelles was investigated as a function of BCP terminal functionalization (i.e., -OH, -COOH, and -NH2 groups) and synthesis method (i.e., electrohydrodynamic emulsification-mediated selfassembly (EE-SA), sonication, and manual shaking). Fluorescence losses, fluorescence intensity, energy spectra, and surface composition were assessed using spectrofluorometry and cathodoluminescence spectroscopy (CL) with integrated X-ray photoemission spectroscopy (XPS). QDs passivated using charged BCPs exhibited 50-80% lower fluorescence intensity than those displaying neutral groups (e.g., -OH), which CL/XPS revealed to result from oxidation of surface Cd to CdO. Fluorescence losses were higher for processes with slow formation speed, but minimized in the presence of poly(vinyl alcohol) (PVA) surfactant. These data suggest slower BCP aggregation kinetics rather than electrostatic chain repulsion facilitated QD oxidation. Thus, polymer coating method and BCP structure influence QD oxidation during phase transfer and should be selected to maximize fast aggregation kinetics.

One step closer to cancer nanomedicine.

High-throughput tool uncovers links between cell signaling and nanomaterial uptake.

Nanoparticles caged with DNA nanostructures.

Nanoparticles (NPs) offer many benefits in biotechnology because of their small size and unique properties. However, many applications require precise positioning of the NPs or biological targeting molecules on their surfaces. DNA cages constructed from DNA tile, origami, or wireframe nanostructures offer a promising path forward because of their simplicity and programmability that can be used to generate complex, dynamic 2D and 3D geometries. Such materials can be used to pattern DNA on NP surfaces and organize NPs into specific supramolecular structures. DNA-caged NPs can be implemented in biosensing and drug delivery applications with cavities precisely designed to encapsulate-specific biomolecules. Ultimately, such approaches provide a springboard for future DNA robot designs that will enable controlled interactions with biological systems.

Synthesis of polymer nanoparticles via electrohydrodynamic emulsification-mediated self-assembly.

HYPOTHESIS: Electrospray can rapidly produce fine, organic solvent-in-water emulsions in the absence of surfactant via electrohydrodynamic emulsification (EE), a reverse configuration of traditional electrospray. This paper investigates whether EE can produce high-quality nanocomposites comprised of block co-polymers and organic nanoparticles (NPs) via the interfacial instability (IS) self-assembly method. Surfactant-free approaches may increase encapsulation efficiency and product uniformity, process speed, and ease of downstream product purification. EXPERIMENTS: All particles were produced using EE-mediated self-assembly (SA) (EE-SA). Particles were produced using poly(lactic-co-glycolic acid) (PLGA) polymers as proof of concept. Then, block copolymer (BCP) micelles were synthesized from polystyrene-block-poly(ethylene oxide) (PS-b-PEO) (PS 9.5 kDa:PEO 18.0 kDa) in the presence and absence of superparamagnetic iron oxide nanoparticles (SPIONs) or quantum dots (QDs). Encapsulant concentration was varied, and the effect of encapsulant NP ligands on final particle size was investigated. FINDINGS: EE-SA generated both pure polymer NPs and nanocomposites containing SPIONs and QDs. PLGA particles spanned from sub- to super-micron sizes. PS-b-PEO NPs and nanocomposites were highly monodisperse, and more highly loaded than those made via a conventional, surfactant-rich IS process. Free ligands decreased the size of pure BCP particles. Increasing encapsulant levels led to a morphological transition from spherical to worm-like to densely loaded structures.

Comparative Encapsulation Efficiency of Lutein in Micelles Synthesized via Batch and High Throughput Methods.

PURPOSE: Black raspberries (BRBs) and their anthocyanin-rich hydrophilic fractions (BRB-H) have exhibited significant chemopreventative activity across aerodigestive cancers. Lutein, the primary component of the BRB lipophilic fraction (BRB-L), also demonstrates bioactivity potential, but is less well characterized, in part because of its poor, innate bioavailability. For these lipophilic compounds to be accurately evaluated for anticancer efficacy, it is necessary to increase their functional bioavailability using delivery vehicles. Lutein has been delivered in commercial settings in emulsion form. However, emulsions are unstable, particularly in the gastrointestinal tract, which limit their use as an oral nutraceutical. Here, we evaluated lutein encapsulation and cellular uptake for nanoparticle (NP) delivery vehicles composed of three different materials synthesized via two different approaches. METHODS: Specifically, NPs were synthesized via smaller scale batch interfacial instability (II) sonication and semi-continuous high throughput electrohydrodynamic-mediated mixing nanoprecipitation (EM-NP) methods using polystyrene-polyethylene oxide (PSPEO) or polycaprolactone-polyethylene glycol (PCLPEG) block copolymers and PHOSPHOLIPON 90G® (P90G, Lipoid GmbH) lipids. Size distribution, lutein encapsulation efficiency (EE), and cellular uptake and delivery were evaluated for each NP formulation. RESULTS: NPs produced via high throughput EM-NP had higher EEs than NPs produced via batch II sonication, and P90G had the greatest EE (55%) and elicited faster cellular uptake in premalignant oral epithelial cells (SCC83) compared to other delivery systems. CONCLUSION: These qualities suggest P90G could be a beneficial candidate for future lutein in vitro delivery research and clinical translation for oral cancer prevention.

Hyaluronic acid induces ROCK-dependent amoeboid migration in glioblastoma cells.

Glioblastoma (GBM) is the most aggressive and deadly adult brain tumor, primarily because of its high infiltrative capacity and development of resistance to therapy. Although GBM cells are typically believed to migrate via mesenchymal (e.g., fibroblast-like) migration modes, amoeboid (e.g., leucocyte-like) migration modes have been identified and may constitute a salvage pathway. However, the mesenchymal to amoeboid transition (MAT) process in GB is not well characterized, most likely because most culture models induce MAT via pharmacological or genetic inhibition conditions that are far from physiological. In this study, we examined the ability of hyaluronic acid (HA) content in three-dimensional collagen (Col) hydrogels to induce MAT in U87 GBM cells. HA and Col are naturally-occurring components of the brain extracellular matrix (ECM). In pure Col gels, U87 cells displayed primarily mesenchymal behaviors, including elongated cell morphology, clustered actin and integrin expression, and crawling migration behaviors. Whereas an increasing population of cells displaying amoeboid behaviors, including rounded morphology, cortical actin expression, low/no integrin expression, and squeezing or gliding motility, were observed with increasing HA content (0.1-0.2 wt% in Col). Consistent with amoeboid migration, these behaviors were abrogated by ROCK inhibition with the non-specific small molecule inhibitor Y27632. Toward identification of histological MAT classification criteria, we also examined the correlation between cell and nuclear aspect ratio (AR) in Col and Col-HA gels, finding that nuclear AR has a small variance and is not correlated to cell AR in HA-rich gels. These results suggest that HA may regulate GBM cell motility in a ROCK-dependent manner.

Self-Assembly and sedimentation of 5 nm SPIONs using horizontal, high magnetic fields and gradients.

Superparamagnetic iron oxide nanoparticles (SPIONs) are employed in multiple applications, especially within medical and chemical engineering fields. However, their magnetic separation is very challenging as the magnetophoretic motion is hindered by thermal energy and viscous drag. Recent studies have addressed the recovery of SPIONs by a combination of cooperative magnetophoresis and sedimentation. Nevertheless, the effect of horizontal, high fields and gradients on the vertical sedimentation of SPIONs has not been described. In this work, we report, for the first time, the magnetically facilitated sedimentation of 5 nm particles by applying fields and gradients perpendicular to gravity. The magnetic field was generated by quadrupole magnetic sorters and the process was measured with time by tracking the concentration along the length of a channel contacting the 5 nm SPIONs within the quadrupole field. Our experimental data suggest that aggregates of 60-90 particles are formed in the system; thus, particle agglomeration by dipole-dipole interactions was promoted, and these clusters settled down as a result of gravitational forces. Multiple variables and parameters were evaluated, including the initial SPION concentration, the temperature, the magnetic field and gradient and operation time. It was found that the process was improved by decreasing the initial concentration and the temperature, but the magnitude of the magnetic field and gradient did not significantly affect the sedimentation. Finally, the separation process was rapid, with the systems reaching the equilibrium in approximately 20 minutes, which is a significant advantage in comparison to other systems that require longer times and larger particle sizes.

MicroRNA-mRNA Interactions at Low Levels of Compressive Solid Stress Implicate mir-548 in Increased Glioblastoma Cell Motility.

Glioblastoma (GBM) is an astrocytic brain tumor with median survival times of <15 months, primarily as a result of high infiltrative potential and development of resistance to therapy (i.e., surgical resection, chemoradiotherapy). A prominent feature of the GBM microenvironment is compressive solid stress (CSS) caused by uninhibited tumor growth within the confined skull. Here, we utilized a mechanical compression model to apply CSS (<115 Pa) to well-characterized LN229 and U251 GBM cell lines and measured their motility, morphology, and transcriptomic response. Whereas both cell lines displayed a peak in migration at 23 Pa, cells displayed differential response to CSS with either minimal (i.e., U251) or large changes in motility (i.e., LN229). Increased migration of LN229 cells was also correlated to increased cell elongation. These changes were tied to epigenetic signaling associated with increased migration and decreases in proliferation predicted via Ingenuity® Pathway Analysis (IPA), characteristics associated with tumor aggressiveness. miRNA-mRNA interaction analysis revealed strong influence of the miR548 family (i.e., mir-548aj, mir-548az, mir-548t) on differential signaling induced by CSS, suggesting potential targets for pharmaceutical intervention that may improve patient outcomes.

Biomolecular detection, tracking, and manipulation using a magnetic nanoparticle-quantum dot platform.

Fluorescent and magnetic materials play a significant role in biosensor technology, enabling sensitive quantification and separations with applications in diagnostics, purification, quality control, and therapeutics. Here, we present a magneto-fluorescent biosensor/separations platform consisting of quantum dots (QDs) and superparamagnetic iron oxide nanoparticles (SPIONs) that are separately encapsulated in amphiphilic block co-polymer micelles conjugated to DNA or protein (i.e., single-stranded (ss) DNA derived from the mRNA of the tumor suppressor protein p53 or avidin protein). Analytes were detected via an aggregation sandwich assay upon binding of at least 1 QD and 1 SPION-containing micelle to result in a fluorescent/magnetic composite. Multiplexed isolation of protein and DNA biomolecules was demonstrated by using QDs of varying emission wavelength; QD fluorescence intensity could be correlated with analyte concentration. Sequential or parallel biomolecule separation was achieved by adding appropriately functionalized SPION-containing micelles and applying user-controlled magnetic fields via patterned magnetic disks and wires. QD fluorescence was used to continuously visualize analyte separation during this process. This QD/SPION platform is simple to use, demonstrates ∼10-16 M sensitivity in analyte detection (comparable to competing QD biosensors based on energy transfer) with specificity against 1 and 2 basepair mismatches in DNA detection, molecular separations capability in solutions of ∼10-10 M, and permits simultaneous or parallel, multiplexed separation of protein and DNA. Thus, this versatile platform enables self-assembly-based rapid, sensitive, and specific detection and separation of biomolecules, simultaneously and with real-time visualization. This technology demonstrates potential for nanoscale assembly, biosensing, and bioseparations.

Reciprocal Control of Hierarchical DNA Origami-Nanoparticle Assemblies.

DNA origami mechanisms offer promising tools for precision nanomanipulation of molecules or nanomaterials. Recent advances have extended the function of individual DNA origami devices to material scales via hierarchical assemblies. However, achieving rapid and precise control of large conformational changes in hierarchical assemblies remains a critical challenge. Here, we demonstrate a method for controlling DNA origami-nanoparticle assemblies through a multiscale approach, in which nanoparticles impart control on the conformation of individual DNA origami mechanisms, whereas DNA origami assemblies control the conformation of nanoparticle arrays. Specifically, we show that the angular distributions of DNA origami hinge mechanisms are tunable as a function of nanoparticle size and distance from the hinge vertex. We selectively adjust the affinity of nanoparticle binding sites, resulting in hinge actuation via DNA melting without releasing the nanoparticle, thereby enabling rapid and reversible temperature-based actuation. Finally, we demonstrate this rapid actuation in DNA origami-nanoparticle arrays of length scales extending over a micron. These results provide guiding principles toward the design of dynamic, DNA-origami hierarchical materials capable of storing and releasing mechanical energy.

Compact quantum dot surface modification to enable emergent behaviors in quantum dot-DNA composites.

Quantum dot (QD) biological imaging and sensing applications often require surface modification with single-stranded deoxyribonucleic acid (ssDNA) oligonucleotides. Furthermore, ssDNA conjugation can be leveraged for precision QD templating via higher-order DNA nanostructures to exploit emergent behaviors in photonic applications. Use of ssDNA-QDs across these platforms requires compact, controlled conjugation that engenders QD stability over a wide pH range and in solutions of high ionic strength. However, current ssDNA-QD conjugation approaches suffer from limitations, such as the requirement for thick coatings, low control over ssDNA labeling density, requirement of large amounts of ssDNA, or low colloidal or photostability, restraining implementation in many applications. Here, we combine thin, multidentate, phytochelatin-3 (PC3) QD passivation techniques with strain-promoted copper-free alkyne-azide click chemistry to yield functional ssDNA-QDs with high stability. This process was broadly applicable across QD sizes (i.e., λem = 540, 560, 600 nm), ssDNA lengths (i.e., 10-16 base pairs, bps), and sequences (poly thymine, mixed bps). The resulting compact ssDNA-QDs displayed a fluorescence quenching efficiency of up to 89% by hybridization with complementary ssDNA-AuNPs. Furthermore, ssDNA-QDs were successfully incorporated with higher-order DNA origami nanostructure templates. Thus, this approach, combining PC3 passivation with click chemistry, generates ssDNA-PC3-QDs that enable emergent QD properties in DNA-based devices and applications.

Effect of Electrospun Fiber Mat Thickness and Support Method on Cell Morphology.

Electrospun fiber mats (EFMs) are highly versatile biomaterials used in a myriad of biomedical applications. Whereas some facets of EFMs are well studied and can be highly tuned (e.g., pore size, fiber diameter, etc.), other features are under characterized. For example, although substrate mechanics have been explored by several groups, most studies rely on Young's modulus alone as a characterization variable. The influence of fiber mat thickness and the effect of supports are variables that are often not considered when evaluating cell-mechanical response. To assay the role of these features in EFM scaffold design and to improve understanding of scaffold mechanical properties, we designed EFM scaffolds with varying thickness (50-200 µm) and supporting methodologies. EFM scaffolds were comprised of polycaprolactone and were either electrospun directly onto a support, suspended across an annulus (3 or 10 mm inner diameter), or "tension-released" and then suspended across an annulus. Then, single cell spreading (i.e., Feret diameter) was measured in the presence of these different features. Cells were sensitive to EFM thickness and suspended gap diameter. Overall, cell spreading was greatest for 50 µm thick EFMs suspended over a 3 mm gap, which was the smallest thickness and gap investigated. These results are counterintuitive to conventional understanding in mechanobiology, which suggests that stiffer materials, such as thicker, supported EFMs, should elicit greater cell polarization. Additional experiments with 50 µm thick EFMs on polystyrene and polydimethylsiloxane (PDMS) supports demonstrated that cells can "feel" the support underlying the EFM if it is rigid, similar to previous results in hydrogels. These results also suggest that EFM curvature may play a role in cell response, separate from Young's modulus, possibly because of internal tension generated. These parameters are not often considered in EFM design and could improve scaffold performance and ultimately patient outcomes.

Beyond Linear Elastic Modulus: Viscoelastic Models for Brain and Brain Mimetic Hydrogels.

With their high degree of specificity and investigator control, in vitro disease models provide a natural complement to in vivo models. Especially in organs such as the brain, where anatomical limitations make in vivo experiments challenging, in vitro models have been increasingly used to mimic disease pathology. However, brain mimetic models may not fully replicate the mechanical environment in vivo, which has been shown to influence a variety of cell behaviors. Specifically, many disease models consider only the linear elastic modulus of brain, which describes the stiffness of a material with the assumption that mechanical behavior is independent of loading rate. Here, we characterized porcine brain tissue using a modified stress relaxation test, and across a panel of viscoelastic models, showed that stiffness depends on loading rate. As such, the linear elastic modulus does not accurately reflect the viscoelastic properties of native brain. Among viscoelastic models, the Maxwell model was selected for further analysis because of its simplicity and excellent curve fit (R2 = 0.99 ± 0.0006). Thus, mechanical response of native brain and hydrogel mimetic models was analyzed using the Maxwell model and the linear elastic model to evaluate the effects of strain rate, time post mortem, region, tissue type (i.e., bulk brain vs white matter), and in brain mimetic models, hydrogel composition, on observed mechanical properties. In comparing the Maxwell and linear elastic models, linear elastic modulus is consistently lower than the Maxwell elastic modulus across all brain regions. Additionally, the Maxwell model is sensitive to changes in viscosity and small changes in elasticity, demonstrating improved fidelity. These findings demonstrate the insufficiency of linear elastic modulus as a primary mechanical characterization for brain mimetic materials and provide quantitative information toward the future design of materials that more closely mimic mechanical features of brain.