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The preparation of gold nanoparticles through reduction of chloroauric acid by trisodium citrate, also known as the Turkevich synthesis, was analyzed both ex situ and in situ. In situ experiments c...

Cell–material interactions are crucial for many biomedical applications, including medical implants, tissue engineering, and biosensors. For implants, while the adhesion of eukaryotic host cells is desirable, bacterial adhesion often leads to infections. Surface free energy (SFE) is an important parameter that controls short- and long-term eukaryotic and prokaryotic cell adhesion. Understanding its effect at a fundamental level is essential for designing materials that minimize bacterial adhesion. Most cell adhesion studies for implants have focused on correlating surface wettability with mammalian cell adhesion and are restricted to short-term time scales. In this work, we used quartz crystal microbalance with dissipation monitoring (QCM-D) and electrical impedance analysis to characterize the adhesion and detachment of S. cerevisiae and E. coli, serving as model eukaryotic and prokaryotic cells within extended time scales. Measurements were performed on surfaces displaying different surface energies (Au, SiO2, and silanized SiO2). Our results demonstrate that tuning the surface free energy of materials is a useful strategy for selectively promoting eukaryotic cell adhesion and preventing bacterial adhesion. Specifically, we show that under flow and steady-state conditions and within time scales up to ∼10 h, a high SFE, especially its polar component, enhances S. cerevisiae adhesion and hinders E. coli adhesion. In the long term, however, both cells tend to detach, but less detachment occurs on surfaces with a high dispersive SFE contribution. The conclusions on S. cerevisiae are also valid for a second eukaryotic cell type, being the human embryonic kidney (HEK) cells on which we performed the same analysis for comparison. Furthermore, each cell adhesion phase is associated with unique cytoskeletal viscoelastic states, which are cell-type-specific and surface free energy-dependent and provide insights into the underlying adhesion mechanisms., info:eu-repo/semantics/published

ispartof: ADVANCED FUNCTIONAL MATERIALS vol:32 issue:5 status: published

The optoelectronic properties of semiconductor nanoparticles make them valuable candidates for the long-term monitoring of transmembrane electric fields in excitable cells. In this work, we show that the electric field sensitivity of the fluorescence intensity of type-I and quasi-type-II quantum dots and quantum rods is enhanced under two-photon excitation compared to single-photon excitation. Based on the superior electric field sensitivity of the two-photon excited fluorescence, we demonstrate the ability of quantum dots and rods to track fast switching E-fields. These findings indicate the potential of semiconductor nanoparticles as cellular voltage probes in multiphoton imaging.

To overcome the problems of refractive index matching and increased disorder when working with traditional heterostructure colloidal photonic crystals (CPCs) with dual or multiple photonic bandgaps (PBGs) for fluorescence enhancement in water, we propose the use of a chemical heterostructure in hollow sphere CPCs (HSCPCs). A partial chemical modification of the HSCPC creates a large contrast in wettability to induce the heterostructure, while the hollow spheres increase the refractive index difference when used in aqueous environment. With the platform, fluorescence enhancement reaches around 160 times in solution, and 72 times (signal-to-background ratio ~7 times) in cells during proof-of-concept live cardiomyocyte contractility experiments. Such photonic platform can be further exploited for chemical sensing, bioassays, and environmental monitoring. Moreover, the introduction of chemical heterostructures provides new design principles for functionalized photonic devices.

Understanding microbial adhesion and retention is crucial for controlling many processes, including biofilm formation, antimicrobial therapy as well as cell sorting and cell detection platforms. Cell detachment is inextricably linked to cell adhesion and retention and plays an important part in the mechanisms involved in these processes. Physico-chemical and biological forces play a crucial role in microbial adhesion interactions and altering the medium ionic strength offers a potential means for modulating these interactions. Real-time studies on the effect of ionic strength on microbial adhesion are often limited to short-term bacterial adhesion. Therefore, there is a need, not only for long-term bacterial adhesion studies, but also for similar studies focusing on eukaryotic microbes, such as yeast. Hereby, we monitored, in real-time, S. cerevisiae adhesion on gold and silica as examples of surfaces with different surface charge properties to disclose long-term adhesion, retention and detachment as a function of ionic strength using quartz crystal microbalance with dissipation monitoring. Our results show that short- and long-term cell adhesion levels in terms of mass-loading increase with increasing ionic strength, while cells dispersed in a medium of higher ionic strength experience longer retention and detachment times. The positive correlation between the cell zeta potential and ionic strength suggests that zeta potential plays a role on cell retention and detachment. These trends are similar for measurements on silica and gold, with shorter retention and detachment times for silica due to strong short-range repulsions originating from a high electron-donicity. Furthermore, the results are comparable with measurements in standard yeast culture medium, implying that the overall effect of ionic strength applies for cells in nutrient-rich and nutrient-deficient media., SCOPUS: ar.j, info:eu-repo/semantics/published

Plasmonic heating finds multiple applications in cell manipulation and stimulation, where heat generated by metal nanoparticles can be used to modify cell adhesion, control membrane currents, and suppress neuronal action potentials among others. Metal nanoparticles can also be easily integrated in artificial extracellular matrices to provide tunable, thermal cueing functionalities with nanometer spatial resolution. In this contribution, we present a platform enabling the combination of plasmonic heating with localized temperature sensing using quantum dots (QDs). Specifically, a functional nanocomposite material was designed with gold nanorods (AuNRs) and QDs incorporated in a cell-permissive hydrogel (e.g., collagen) as well as an optical set-up combining optical heating and temperature imaging, respectively. Specific area stimulation/readout can be realized through structured illumination using digital micromirror device (DMD) projection.

Hematopoietic stem/progenitor cells (HSPCs) are responsible for the generation of blood cells throughout life. It is believed that, in addition to soluble cytokines and niche cells, biophysical cues like elasticity and oxygen tension are responsible for the orchestration of stem cell fate. Although several studies have examined the effects of bone marrow (BM) niche elasticity on HSPC behavior, no study has yet investigated the effects of the elasticity of other niche sites like the fetal liver (FL), where HSPCs expand more extensively. In this study, we evaluated the effect of matrix stiffness values similar to those of the FL on BM-derived HSPC expansion. We first characterized the elastic modulus of murine FL tissue at embryonic day E14.5. Fibrin hydrogels with similar stiffness values as the FL (soft hydrogels) were compared with stiffer fibrin hydrogels (hard hydrogels) and with suspension culture. We evaluated the expansion of total nucleated cells (TNCs), Lin&minus, /cKit+ cells, HSPCs (Lin&minus, /Sca+/cKit+ (LSK) cells), and hematopoietic stem cells (HSCs: LSK- Signaling Lymphocyte Activated Molecule (LSK-SLAM) cells) when cultured in 5% O2 (hypoxia) or in normoxia. After 10 days, there was a significant expansion of TNCs and LSK cells in all culture conditions at both levels of oxygen tension. LSK cells expanded more in suspension culture than in both fibrin hydrogels, whereas TNCs expanded more in suspension culture and in soft hydrogels than in hard hydrogels, particularly in normoxia. The number of LSK-SLAM cells was maintained in suspension culture and in the soft hydrogels but not in the hard hydrogels. Our results indicate that both suspension culture and fibrin hydrogels allow for the expansion of HSPCs and more differentiated progeny whereas stiff environments may compromise LSK-SLAM cell expansion. This suggests that further research using softer hydrogels with stiffness values closer to the FL niche is warranted.

Quantum dots (QDs) have been hypothesized as potential probes for the optical monitoring of electrogenic cell activity given their high optical absorption cross sections compared to organic fluorophores, high brightness, and low photobleaching. Despite the theoretical predictions, less than a handful of papers reported QD-based imaging of cell activity responses because of the critical membrane localization requirements for membrane voltage sensing. Moreover, to the best of our knowledge, two-photon imaging of cellular activity-dependent QD photoluminescence was never demonstrated. The high spatial and temporal resolutions and higher penetration depths, inherent to nonlinear light–tissue interactions, are particularly interesting for biological imaging. In this work, we functionalized fibrous collagen matrices with semiconductor QDs and thereby created artificial extracellular matrices that can optically report cardiomyocyte contractile activity based on QD two-photon fluorescence. We have applied these optically addressable nanofiber matrices to monitor the contractile activity of primary cardiomyocytes, and, for validation, we compared the optical responses with the simultaneously recorded patch-clamp data. Given the long-term stability of QD fluorescence, near-infrared excitation, and high spatiotemporal resolution achievable through multiphoton imaging, this approach can be used for continuous monitoring of cellular functions in cardiac tissue constructs.

Structured illumination microscopy (SIM) is a widely used super resolution imaging technique that can down-modulate a sample's high-frequency information into objective recordable frequencies to enhance the resolution below the diffraction limit. However, classical SIM image reconstruction methods often generate poor results under low illumination conditions, which are required for reducing photobleaching and phototoxicity in cell imaging experiments. Although denoising methods or auxiliary items improved SIM image reconstruction in low signal level situations, they still suffer from decreased reconstruction quality and significant background artifacts, inevitably limiting their practical applications. In order to improve the reconstruction quality, second-order optimized regularized SIM (sorSIM) is designed specifically for image reconstruction in low signal level situations. In sorSIM, a second-order regularization term is introduced to suppress noise effect, and the penalty factor in this term is selected to optimize the resolution enhancement and noise resistance. Compared to classical SIM image reconstruction algorithms as well as to those previously used in low illumination cases, the proposed sorSIM provides images with enhanced resolution and fewer background artifacts. Therefore, sorSIM can be a potential tool for high-quality and rapid super resolution imaging, especially for low signal images. ispartof: Optics Express vol:28 issue:11 pages:16708-16724 ispartof: location:United States status: Published online

The incorporation of functional nanoparticles in scaffolds for tissue constructs has led to the creation of artificial extracellular matrices that more accurately mimic the cues present in the native microenvironment of developing tissue. Additionally, light-sensitive inorganic nanoparticles can act as cell biosensors and report on the physiological parameters during tissue growth and organization. In this work, we functionalized collagen nanofibers with semiconductor quantum dots (QDs) and thereby created artificial extracellular matrices that can optically report on cardiomyocyte activity based on QD two-photon fluorescence. We have applied these optically-addressable nanofiber matrices to monitor activities of primary cardiomyocytes and compared the optical responses with patch-clamp data. Combining the long-term stability of QD fluorescence with the deeper light penetration depths achievable through multiphoton imaging, this approach can be used for continuous monitoring of cellular functions in cardiac tissue engineering.Abstract FigureConcept illustration: optical readout of cardiomyocyte activity with QD-functionalized collagen networks. Whole-cell current-clamp mode is used here to simultaneously monitor changes in the transmembrane voltage while the QD two-photon fluorescence is recorded.

Optical interrogation of cellular electrical activity has proven itself essential for understanding cellular function and communication in complex networks. Voltage-sensitive dyes are important tools for assessing excitability but these highly lipophilic sensors may affect cellular function. Label-free techniques offer a major advantage as they eliminate the need for these external probes. In this work, we show that endogenous second harmonic generation (SHG) from live cells is highly sensitive to changes in membrane potential. Simultaneous electrophysiological control of a living (HEK293T) cell, through whole-cell voltage clamp reveals a linear relation between the SHG intensity and membrane voltage. Our results suggest that due to the high ionic strengths and fast optical response of biofluids, membrane hydration is not the main contributor to the observed field sensitivity. We further provide a conceptual framework that indicates that the SHG voltage sensitivity reflects the electric field within the biological asymmetric lipid bilayer owing to a nonzero tensor. Changing the membrane potential without surface modifications such as electrolyte screening offers high optical sensitivity to membrane voltage (~40% per 100 mV), indicating the power of SHG for label-free read-out. These results hold promise for the design of a non-invasive label-free read-out tool for electrogenic cells.

In this article, we report on the development of a flow cell optimized for the heat-transfer method, a versatile biosensing technique. The design of the flow cell ensures that the heat flow is focused with minimal heat loss through the surroundings of the cell. This results in a more stable measuring signal and an improved sensitivity of the measuring technique. The sensor was characterized by performing background measurements in air, water, and phosphate buffered saline (PBS) solution. Heat flow through the setup was simulated using COMSOL in order to provide insight in the contribution of convection to the heat flow and recommendations for possible future improvements to the cell. Additionally, a two-step algorithm for calculating thermal resistance was defined, allowing the user to accurately derive thermal conductivity from experimental data. Finally, the potential of the flow cell for bacteria (Escherichia coli) detection was assessed and compared with the results obtained in the original HTM setup in a similar experiment. This experiment demonstrates that we were able to improve the limit-of-detection (LoD) to 2.10 × 104 colony forming units (CFU) mL−1 by changing the geometry of the measuring cell. Sensor setup for thermal biodetection experiments a directed heat flow.