Metal-Loaded Synthetic Melanin via Oxidative Polymerization of Neurotransmitter Norepinephrine Exhibiting High Photothermal Conversion.

Polydopamine (PDA)-derived melanin-like materials exhibit significant photothermal conversion owing to their broad-spectrum light absorption. However, their low near-infrared (NIR) absorption and inadequate hydrophilicity compromise their utilization of solar energy. Herein, we developed metal-loaded poly(norepinephrine) nanoparticles (PNE NPs) by predoping metal ions (Fe3+, Mn3+, Co2+, Ca2+, Ga3+, and Mg2+) with norepinephrine, a neuron-derived biomimetic molecule, to address the limitations of PDA. The chelation between catechol and metal ions induces a ligand-to-metal charge transfer (LMCT) through the formation of donor-acceptor pairs, modulating the light absorption behavior and reducing the band gap. Under 1 sun illumination, the Fe-loaded PNE coated wood evaporator achieved a high seawater evaporation rate and efficiency of 1.75 kg m-2 h-1 and 92.4%, respectively, owing to the superior hydrophilicity and photothermal performance of PNE. Therefore, this study offers a comprehensive exploration of the role of metal ions in enhancing the photothermal properties of synthetic melanins.

A novel Y-shaped photoiniferter used for the construction of polydimethylsiloxane surfaces with antibacterial and antifouling properties.

The simultaneous introduction of two new functionalities into the same polymeric substrate under mild reaction conditions is an interesting and important topic. Herein, dual-functional polydimethylsiloxane (PDMS) surfaces with antibacterial and antifouling properties were conveniently developed via a novel Y-shaped asymmetric dual-functional photoiniferter (Y-iniferter). The Y-iniferter was initially immobilized onto the PDMS surface by radical coupling under visible light irradiation. Afterwards, poly(2-hydroxyethyl methacrylate) (PHEMA) brushes and antibacterial ionic liquid (IL) fragments were simultaneously immobilized on the Y-iniferter-modified PDMS surfaces by combining the sulfur(VI)-fluoride exchange (SuFEx) click reaction and UV-photoinitiated polymerization. Experiments using E. coli as a model bacterium demonstrated that the modified PDMS surfaces had both the expected antibacterial properties of the IL fragments and the excellent antifouling properties of PHEMA brushes. Furthermore, the cytotoxicity of the modified PDMS surfaces to L929 cells was examined in vitro with a CCK-8 assay, which showed that the modified surfaces maintained excellent cytocompatibility. Briefly, this strategy of constructing an antibacterial and antifouling PDMS surface has the advantages of simplicity and convenience and might inspire the construction of diverse dual-functional surfaces by utilizing PDMS more effectively.

Contributions of photochemistry to bio-based antibacterial polymer materials.

Surgical site infections constitute a major health concern that may be addressed by conferring antibacterial properties to surgical tools and medical devices via functional coatings. Bio-sourced polymers are particularly well-suited to prepare such coatings as they are usually safe and can exhibit intrinsic antibacterial properties or serve as hosts for bactericidal agents. The goal of this Review is to highlight the unique contribution of photochemistry as a green and mild methodology for the development of such bio-based antibacterial materials. Photo-generation and photo-activation of bactericidal materials are illustrated. Recent efforts and current challenges to optimize the sustainability of the process, improve the safety of the materials and extend these strategies to 3D biomaterials are also emphasized.

Radiation-induced graft polymerization of elastin onto polyvinylpyrrolidone as a possible wound dressing.

The purpose of our study was to obtain new wound dressings in the form of hydrogels that promote wound healing taking advantage of the broad activities of elastin (ELT) in physiological processes. The hydrogel of ELT and polyvinylpyrrolidone (PVP; ELT-PVP) was obtained by cross-linking induced by gamma irradiation at a dose of 25 kGy. The physicochemical changes attributed to cross-linking were analyzed through scanning electron microscopy (SEM), infrared spectroscopy analysis with Fourier transform (FTIR), differential scanning calorimetry (DSC), and thermogravimetric analysis (TGA). Furthermore, we performed a rheological study to determine the possible changes in the fluidic macroscopic properties produced by the cross-linking method. Finally, we accomplished viability and proliferation analyses of human dermal fibroblasts in the presence of the hydrogel to evaluate its biological characteristics. The hydrogel exhibited a porous morphology, showing interconnected porous with an average pore size of 16 ± 8.42 µm. The analysis of FTIR, DSC, and TGA revealed changes in the chemical structure of the ELT-PVP hydrogel after the irradiation process. Also, the hydrogel exhibited a rheological behavior of a pseudoplastic and thixotropic fluid. The hydrogel was biocompatible, demonstrating high cell viability, whereas ELT presented low biocompatibility at high concentrations. In summary, the hydrogel obtained by gamma irradiation revealed the appropriate morphology to be applied as a wound dressing. Interestingly, the hydrogel exhibited a higher percentage of cell viability compared with ELT, suggesting that the cross-linking of ELT with PVP is a suitable strategy for biological applications of ELT without generating cellular damage.

Stretchable and Bioadhesive Gelatin Methacryloyl-Based Hydrogels Enabled by in Situ Dopamine Polymerization.

Hydrogel patches with high toughness, stretchability, and adhesive properties are critical to healthcare applications including wound dressings and wearable devices. Gelatin methacryloyl (GelMA) provides a highly biocompatible and accessible hydrogel platform. However, low tissue adhesion and poor mechanical properties of cross-linked GelMA patches (i.e., brittleness and low stretchability) have been major obstacles to their application for sealing and repair of wounds. Here, we show that adding dopamine (DA) moieties in larger quantities than those of conjugated counterparts to the GelMA prepolymer solution followed by alkaline DA oxidation could result in robust mechanical and adhesive properties in GelMA-based hydrogels. In this way, cross-linked patches with ∼140% stretchability and ∼19 000 J/m3 toughness, which correspond to ∼5.7 and ∼3.3× improvement, respectively, compared to that of GelMA controls, were obtained. The DA oxidization in the prepolymer solution was found to play an important role in activating adhesive properties of cross-linked GelMA patches (∼4.0 and ∼6.9× increase in adhesion force under tensile and shear modes, respectively) due to the presence of reactive oxidized quinone species. We further conducted a parametric study on the factors such as UV light parameters, the photoinitiator type (i.e., lithium phenyl-2,4,6-trimethylbenzoylphosphinate, LAP, versus 2-hydroxy-4'-(2-hydroxyethoxy)-2-methylpropiophenone, Irgacure 2959), and alkaline DA oxidation to tune the cross-linking density and thereby hydrogel compliance for better adhesive properties. The superior adhesion performance of the resulting hydrogel along with in vitro cytocompatibility demonstrated its potential for use in skin-attachable substrates.

A dissipative pathway for the structural evolution of DNA fibres.

Biochemical networks interconnect, grow and evolve to express new properties as different chemical pathways are selected during a continuous cycle of energy consumption and transformation. In contrast, synthetic systems that push away from equilibrium usually return to the same self-assembled state, often generating waste that limits system recyclability and prevents the formation of adaptable networks. Here we show that annealing by slow proton dissipation selects for otherwise inaccessible morphologies of fibres built from DNA and cyanuric acid. Using single-molecule fluorescence microscopy, we observe that proton dissipation influences the growth mechanism of supramolecular polymerization, healing gaps within fibres and converting highly branched, interwoven networks into nanocable superstructures. Just as the growth kinetics of natural fibres determine their structural attributes to modulate function, our system of photoacid-enabled depolymerization and repolymerization selects for healed materials to yield organized, robust fibres. Our method provides a chemical route for error-checking, distinct from thermal annealing, that improves the morphologies and properties of supramolecular materials using out-of-equilibrium systems.

Photo-Polymerization Damage Protection by Hydrogen Sulfide Donors for 3D-Cell Culture Systems Optimization.

Photo-polymerized hydrogels are ideally suited for stem-cell based tissue regeneration and three dimensional (3D) bioprinting because they can be highly biocompatible, injectable, easy to use, and their mechanical and physical properties can be controlled. However, photo-polymerization involves the use of potentially toxic photo-initiators, exposure to ultraviolet light radiation, formation of free radicals that trigger the cross-linking reaction, and other events whose effects on cells are not yet fully understood. The purpose of this study was to examine the effects of hydrogen sulfide (H2S) in mitigating cellular toxicity of photo-polymerization caused to resident cells during the process of hydrogel formation. H2S, which is the latest discovered member of the gasotransmitter family of gaseous signalling molecules, has a number of established beneficial properties, including cell protection from oxidative damage both directly (by acting as a scavenger molecule) and indirectly (by inducing the expression of anti-oxidant proteins in the cell). Cells were exposed to slow release H2S treatment using pre-conditioning with glutathione-conjugated-garlic extract in order to mitigate toxicity during the photo-polymerization process of hydrogel formation. The protective effects of the H2S treatment were evaluated in both an enzymatic model and a 3D cell culture system using cell viability as a quantitative indicator. The protective effect of H2S treatment of cells is a promising approach to enhance cell survival in tissue engineering applications requiring photo-polymerized hydrogel scaffolds.

Simultaneous Grafting Polymerization of Acrylic Acid and Silver Aggregates Formation by Direct Reduction Using γ Radiation onto Silicone Surface and Their Antimicrobial Activity and Biocompatibility.

The modification of medical devices is an area that has attracted a lot of attention in recent years; particularly, those developments which search to modify existing devices to render them antimicrobial. Most of these modifications involve at least two stages (modification of the base material with a polymer graft and immobilization of an antimicrobial agent) which are both time-consuming and complicate synthetic procedures; therefore, as an improvement, this project sought to produce antimicrobial silicone (PDMS) in a single step. Using gamma radiation as both an energy source for polymerization initiation and as a source of reducing agents in solution, PDMS was simultaneously grafted with acrylic acid and ethylene glycol dimethacrylate (AAc:EGDMA) while producing antimicrobial silver nanoparticles (AgNPs) onto the surface of the material. To obtain reproducible materials, experimental variables such as the effect of the dose, the intensity of radiation, and the concentration of the silver salt were evaluated, finding the optimal reaction conditions to obtain materials with valuable properties. The characterization of the material was performed using electronic microscopy and spectroscopic techniques such as 13C-CPMAS-SS-NMR and FTIR. Finally, these materials demonstrated good antimicrobial activity against S. aureus while retaining good cell viabilities (above 90%) for fibroblasts BALB/3T3.

Ionic Carbazole-Based Water-Soluble Two-Photon Photoinitiator and the Fabrication of Biocompatible 3D Hydrogel Scaffold.

Two-photon polymerization of a three-dimensional (3D) hydrogel structure has been widely applied in biological tissue engineering. For improving the biocompatibility of hydrogel structures, a new kind of ionic carbazole water-soluble photoinitiator was prepared to realize the fabrication of a 3D hydrogel structure in aqueous phase. 3,6-Bis[2-(1-methyl-pyridinium)vinyl]-9-methyl-carbazole diiodide (BMVMC) and cucurbit[7]uril (CB7) have been employed to generate a complex with better water solubility by host-guest interactions. The binding ratio of the complex was demonstrated to be 1:1 through the characterization of isothermal titration calorimetry (ITC). The two-photon absorption (TPA) cross section of the complex increases to 2500 GM compared with the 750 GM of the BMVMC molecule. Then, an aqueous-phase photoresist was obtained using the CB7/BMVMC complex as the photoinitiator and poly(ethylene glycol) diacrylate (PEGda) as the hydrogel monomer. Two-photon fabrication capability in aqueous phase has been studied using the as-prepared photoresist. A low laser threshold of 3.7 mW as well as a high resolution of 180 nm are achieved. Benefiting from the fluorescence properties of the photoinitiator, we can achieve the confocal fluorescence images without any assistance of fluorescent probes. Subsequently, a 3D engineered hydrogel scaffold microstructure was fabricated by the two-photon polymerization technology, whose biocompatibility was demonstrated by culturing the structure with living cells of L929. The BMVMC-CB7 complex and the as-prepared photoresist are demonstrated to have good biocompatibility, which is prospective for further application in tissue engineering.

Spatiotemporal Control of Supramolecular Polymerization and Gelation of Metal-Organic Polyhedra.

In coordination-based supramolecular materials such as metallogels, simultaneous temporal and spatial control of their assembly remains challenging. Here, we demonstrate that the combination of light with acids as stimuli allows for the spatiotemporal control over the architectures, mechanical properties, and shape of porous soft materials based on metal-organic polyhedra (MOPs). First, we show that the formation of a colloidal gel network from a preformed kinetically trapped MOP solution can be triggered upon addition of trifluoroacetic acid (TFA) and that acid concentration determines the reaction kinetics. As determined by time-resolved dynamic light scattering, UV-vis absorption, and 1H NMR spectroscopies and rheology measurements, the consequences of the increase in acid concentration are (i) an increase in the cross-linking between MOPs; (ii) a growth in the size of the colloidal particles forming the gel network; (iii) an increase in the density of the colloidal network; and (iv) a decrease in the ductility and stiffness of the resulting gel. We then demonstrate that irradiation of a dispersed photoacid generator, pyranine, allows the spatiotemporal control of the gel formation by locally triggering the self-assembly process. Using this methodology, we show that the gel can be patterned into a desired shape. Such precise positioning of the assembled structures, combined with the stable and permanent porosity of MOPs, could allow their integration into devices for applications such as sensing, separation, catalysis, or drug release.

Synthesis of an UV-Curable Divinyl-Fumarate Poly-ε-Caprolactone for Stereolithography Applications.

The limited number of commercially available photocrosslinkable resins for stereolithography has often been considered the main limitation of this technique. In this manuscript, a photocrosslinkable poly-ε-caprolactone (PCL) has been synthesized by a two-step method starting from ring opening polymerization (ROP) of ε-caprolactone. Hydroxyethyl vinyl ether (HEVE) has been used both as the initiator of ROP and as photo-curable functional group to obtain a vinyl poly-ε-caprolactone (VPCL). The following reaction of VPCL with fumaryl chloride (FuCl) results in a divinyl-fumarate polycaprolactone (VPCLF). Moreover, a catalyst based on Al, instead of the most popular Tin(II) 2-ethylhexanoate, has been employed to reduce the cytotoxicity of the material. VPCLF has been successfully used, in combination with N-vinyl-pyrrolidone (NVP), to fabricate 3D porous scaffolds by micro-stereolithography (µ-SL) with mathematically defined architectures.

Influence of curing duration and mixing techniques of bulk fill resin composites on bi-axial flexural strength and degree of conversion.

OBJECTIVES: The aim was to assess the influence of polymerization duration, method and resin manipulation techniques on the biaxial flexural strength (BFS) and degree of conversion (DC) of bulk fill resin composites (BFRC). METHODS: One hundred and eighty disc specimens were fabricated using MultiCore (MC) and Core-It (CI) bulk fill resin composite. Each material group, specimens were divided into nine subgroups based on curing methods (Light cure for 10 and 20 s; and auto-cure) and mixing techniques (first auto-mix, second automix, and hand mix). BFS was tested with a ball indenter at a crosshead speed of 0.50 mm/min. DC was assessed for MC and CI materials for 10 s and 20 s light cure; and auto cure specimens using Fourier Transform-Infrared Spectroscopy (FTIR). Statistical data comparisons were performed using ANOVA, Bonferroni and Tukey-Kramer tests. RESULTS: For MC and CI, BFS was highest in 10 s light cure specimens, however comparable to specimens cured for 20 s (p > 0.05). Auto cure specimens showed lower BFS than light cured samples for both materials (p < 0.05). Hand mixed specimens showed significantly compromised BFS compared to automix technique for MC and CI. DC % was comparable for 10 s and 20 s light cure methods for both materials (p > 0.05), which was higher than DC % of auto cure bulk fill resins (p < 0.05). CI showed higher DC % and BFS compared to MC bulk fill resin composite. CONCLUSION: Photo-polymerization duration of 10 and 20 s showed similar outcomes for BFS and DC %; and BFS for auto-mixed resins (MC and CI) was significantly higher than hand mixed resin. BFS and DC was higher in photopolymerized groups as compared to auto-cured resin regardless of the manipulation technique for both materials (MultiCore and Core it).

Evaluation of degree of conversion, rate of cure, microhardness, depth of cure, and contraction stress of new nanohybrid composites containing pre-polymerized spherical filler.

The aim of the present study was to characterize nanohybrid and nanofilled composites in terms of degree of conversion (DC), rate of cure (RC), microhardness (Vickers hardness number; VHN), depth of cure, and contraction stress (CS). Ceram.X® universal- A3, duo enamel E2, and duo dentin D3 composites were compared to Tetric EvoCeram® and FiltekTMSupreme XTE composites of equivalent dentin and enamel shades under a 40 s photopolymerization protocol. DC was measured by infrared spectroscopy, calculating RC from the kinetic curve. Top and bottom VHN were determined using a Vickers indenter, and bottom/top surface ratio (Vickers hardness ratio; VHR) calculated. CS vs. time was assessed by a universal testing machine and normalized for the specimen bonding area. All materials showed DC < 60%, Ceram.X® composites reaching higher values than the other composites of corresponding shades. RC at 5 s of photopolymerization was always higher than that at 10 s. All the Ceram.X® composites and the lighter-shaded Tetric EvoCeram® and FiltekTMSupreme XTE composites reached the RC plateau after 25 s, the remaining materials showed a slower kinetic trend. Tetric EvoCeram® and FiltekTMSupreme XTE composites displayed the softest and the hardest surfaces, respectively. Differently from darker-shaded materials, the universal and the three enamel-shaded composites resulted optimally cured (VHR > 80%). The tested composites differed in CS both during and after light cure, Tetric EvoCeram® and FiltekTMSupreme XTE composites displaying the highest and the lowest CS, respectively. Only the Ceram.X® universal-A3 reached a CS plateau value. The tested composites exhibited material-dependent chemo-mechanical properties. Increasing the curing time and/or reducing the composite layer thickness for dentin-shaded composites appears advisable.

Femtosecond laser programmed artificial musculoskeletal systems.

Natural musculoskeletal systems have been widely recognized as an advanced robotic model for designing robust yet flexible microbots. However, the development of artificial musculoskeletal systems at micro-nanoscale currently remains a big challenge, since it requires precise assembly of two or more materials of distinct properties into complex 3D micro/nanostructures. In this study, we report femtosecond laser programmed artificial musculoskeletal systems for prototyping 3D microbots, using relatively stiff SU-8 as the skeleton and pH-responsive protein (bovine serum albumin, BSA) as the smart muscle. To realize the programmable integration of the two materials into a 3D configuration, a successive on-chip two-photon polymerization (TPP) strategy that enables structuring two photosensitive materials sequentially within a predesigned configuration was proposed. As a proof-of-concept, we demonstrate a pH-responsive spider microbot and a 3D smart micro-gripper that enables controllable grabbing and releasing. Our strategy provides a universal protocol for directly printing 3D microbots composed of multiple materials.

NIR Sensitizer Operating under Long Wavelength (1064 nm) for Free Radical Photopolymerization Processes.

Free radical polymerization upon near-infrared (NIR) light is still the subject of intense research efforts and remains a huge challenge particularly for long wavelengths (>1000 nm). In this study, a NIR sensitizer operating upon long wavelength (1064 nm) is proposed for an efficient polymerization of acrylate monomers. A new three-component photoinitiating system is developed comprising the NIR sensitizer in combination with an Iodonium salt (Iod) and an amine. Remarkably, the NIR sensitizer (IR 1064) absorbing strongly in all the near infrared region (700-1200 nm) offers the possibility to use a broad range of irradiation wavelengths, i.e., examples are provided at 785 and 1064 nm. Such long wavelengths are characterized by many advantages such as a deeper penetration of light and therefore a better curing of the monomer but it is also much safer than UV light. Excellent performance is observed for the three-component IR 1064/Iod/Amine system under air: high conversion of acrylate functions associated with a fast polymerization time. The use of IR 1064 as NIR sensitizer with a broad NIR absorption is-to the best of current knowledge-never proposed in the literature. The photoinitiating performances are studied using real-time Fourier transform infrared spectroscopy.

Propagation of THz irradiation energy through aqueous layers: Demolition of actin filaments in living cells.

The effect of terahertz (THz) radiation on deep tissues of human body has been considered negligible due to strong absorption by water molecules. However, we observed that the energy of THz pulses transmits a millimeter thick in the aqueous solution, possibly as a shockwave, and demolishes actin filaments. Collapse of actin filament induced by THz irradiation was also observed in the living cells under an aqueous medium. We also confirmed that the viability of the cell was not affected under the exposure of THz pulses. The potential of THz waves as an invasive method to alter protein structure in the living cells is demonstrated.

Fabrication of 3D-Printed Fish-Gelatin-Based Polymer Hydrogel Patches for Local Delivery of PEGylated Liposomal Doxorubicin.

3D printing technology has been applied to various fields and its medical applications are expanding. Here, we fabricated implantable 3D bio-printed hydrogel patches containing a nanomedicine as a future tailored cancer treatment. The patches were prepared using a semi-solid extrusion-type 3D bioprinter, a hydrogel-based printer ink, and UV-LED exposure. We focused on the composition of the printer ink and semi-synthesized fish gelatin methacryloyl (F-GelMA), derived from cold fish gelatin, as the main component. The low viscosity of F-GelMA due to its low melting point was remarkably improved by the addition of carboxymethyl cellulose sodium (CMC), a pharmaceutical excipient. PEGylated liposomal doxorubicin (DOX), as a model nanomedicine, was incorporated into the hydrogel and liposome stability after photo-polymerization was evaluated. The addition of CMC inhibited particle size increase. Three types of 3D-designed patches (cylinder, torus, gridlines) were produced using a 3D bioprinter. Drug release was dependent on the shape of the 3D-printed patches and UV-LED exposure time. The current study provides useful information for the preparation of 3D printed nanomedicine-based objects.

Surface grafting of fluorescent polymers on halloysite nanotubes through metal-free light-induced controlled polymerization: Preparation, characterization and biological imaging.

Halloysite nanotubes (HNTs) are a kind of aluminosilicate clay with a unique hollow tubular structure that has been intensively explored for various applications especially in biomedical fields owing to their excellent biocompatibility, biodegrading potential and low cost. Surface modification of HNTs with functional polymers will greatly improve their properties and endow new functions for biomedical applications. In this work, a light-induced reversible addition-fragmentation chain transfer (RAFT) polymerization was introduced to successfully prepare HNTs based fluorescent HNTs/poly(PEGMA-Fl) composites in the presence of oxygen using diacrylate-fluorescein and poly (ethylene glycol) methyl ether methacrylate (PEGMA) as the monomers. Without other catalysts, heating, and deoxygenation procedure, the polymerization process can take place under mild conditions. Besides, owing to the introduction of fluorescein and PEGMA on the surface of HNTs, the resultant HNTs/poly(PEGMA-Fl) composites display high water dispersibility and stable fluorescence. The results from cell viability examination and confocal laser scanning microscopy also demonstrated that HNTs/poly(PEGMA-Fl) composites could be internalized by L929 cells with bright fluorescence and low cytotoxicity. Taken together, we developed a novel photo-initiated RAFT polymerization method for the fabrication of HNTs based fluorescent polymeric composites with great potential for biomedical applications. More importantly, many other multifunctional HNTs based polymer composites could also be fabricated through a similar strategy owing to good designability of RAFT polymerization.

Cell encapsulation spatially alters crosslink density of poly(ethylene glycol) hydrogels formed from free-radical polymerizations.

Photopolymerizable poly(ethylene glycol) (PEG) hydrogels are a promising platform for chondrocyte encapsulation and cartilage tissue engineering. This study demonstrates that during the process of encapsulation, chondrocytes alter the formation of PEG hydrogels leading to a reduction in the bulk and local hydrogel crosslink density. Freshly isolated chondrocytes were shown to interact with hydrogel precursors, in part through thiol-mediated events between dithiol crosslinkers and cell surface free thiols, depleting crosslinker concentration and causing a reduction in the bulk hydrogel crosslink density. This effect was more pronounced with increasing cell density at the time of encapsulation. Encapsulation of chondrocytes in fluorescently labeled hydrogels exhibited a gradient in hydrogel density around the cell, which was abrogated by treatment of the cells with the antioxidant estradiol prior to encapsulation. This gradient led to spatial variations in the degradation behavior of a hydrolytically degradable PEG hydrogel, creating regions devoid of hydrogel surrounding cells. Collectively, findings from this study indicate that the antioxidant defense mechanisms in chondrocytes alter the resultant properties of PEG hydrogels formed by free-radical polymerizations. These interactions will have a significant impact on tissue engineering, affecting the local microenvironment around cells and how tissue grows within the hydrogels. STATEMENT OF SIGNIFICANCE: Cell encapsulations in synthetic hydrogels formed by free-radical polymerizations offer numerous benefits for tissue engineering. Herein, we studied cartilage cells and identified that during encapsulation, cells interfered with hydrogel formation through two distinct mechanisms. Thiol-mediated events between monomers led to monomer depletion and a lower crosslinked hydrogel. Cells' antioxidant defense mechanisms interfered with free-radicals and inhibited hydrogel formation near the cell. These cell-mediated effects led to softer hydrogels and created unique hydrogel degradations patterns causing rapid degradation around the cells. The latter has benefits for tissue engineering, where these regions provide space for tissue growth. Overall, this study demonstrates that cells play a key role in how the hydrogel structure forms when cells are present.

Lossless immunocytochemistry using photo-polymerized hydrogel thin-films.

Immunocytochemistry (ICC), or immunofluorescence microscopy, is an essential biological technique for phenotyping cells in both research and diagnostic applications. Standard ICC methods often do not work well when the cell sample contains a small number of cells (<10 000) because of the significant cell loss that occurs during washing, staining, and centrifugation steps. Cell loss is particularly relevant when working with rare cells, such as circulating tumor cells, where such losses could significantly bias experimental outcomes. In order to eliminate cell loss in ICC protocols, we present a method to encapsulate the cell sample in a photo-polymerized hydrogel thin-film. The hydrogel thin-film is permeable to antibodies and other ICC reagents, thereby allowing the use of standard ICC protocols without modification. The cell sample is physically constrained by the hydrogel at the bottom surface of a standard (unmodified) imaging microtiter plate, thereby enabling the acquisition of high-quality micrographs regardless of the properties of the cell sample or staining reagents. Furthermore, while standard ICC requires several centrifugation steps during staining and washing, our hydrogel encapsulation method requires only a single centrifugation step. This property greatly reduces the time required to perform ICC protocols and is more compatible with robotic platforms. In this study, we show that standard ICC and Cytospin protocols are extremely lossy (>70% loss) when the sample contains less than 10 000 cells, while encapsulating the cells using a permeable hydrogel thin-film results in a lossless ICC process.