Photosensitive Crosslinking Research Articles

An Optimized Protocol for Multiple Immunohistochemical Staining of Fragile Tissue Samples.

Owing to the high occurrence of tissue detachment during the sample preparation process, the application of multiplex immunohistochemistry (mIHC) technology is limited in the field of fragile tissue samples, such as tendons, ligaments, and bones. To optimize a method for preparing sections for mIHC on fragile tissue samples, taking the human anterior cruciate ligament as an example, paraffin-embedded continuous sections with a thickness of 4 μm were divided into two groups: baking groups underwent routine section processing, and after being mounted on glass slides, they were baked at 65°C for 4 h, 8 h, or 24 h; ultraviolet (UV) photosensitive cross-linking groups used adhesive-coated slides for mounting and were directly subjected to UV light-induced cross-linking, with the cross-linking time set at 0 s, 20 s, 40 s, 1 min, 2 min, 3 min, 4 min, and 5 min, respectively. After deparaffinization and rehydration, we simulated the microwave step, which was most likely to cause tissue detachment during the mIHC experimental procedure, and then, the sections were stained with eosin. Finally, using the optimal cross-linking time selected from the UV cross-linking groups, mIHC staining of tendon and bone tissues was performed. After deparaffinization and rehydration, both groups were able to maintain the integrity of the tissue structure, except for the slides from the UV-sensitive cross-linking 0 s group, which showed complete tissue detachment. Following the seventh microwave treatment, the baking groups presented significant tissue detachment. The UV cross-linking groups were affected by the cross-linking time, and severe tissue detachment occurred with cross-linking times of 20 s, 40 s, and 5 min, whereas the tissues cross-linked for 1 min, 2 min, 3 min, and 4 min all maintained complete tissue morphology and structure. Finally, after 2 min of cross-linking, the results of spectral imaging revealed that the tissue morphology and structure were intact. During the process of mIHC staining, photocrosslinking with UV irradiation for 1-4 min effectively preserves the integrity of the tissue morphological structure.

Chiral Cell Nanomechanics Originated in Clockwise/Counterclockwise Biofunctional Microarrays to Govern the Nuclear Mechanotransduction of Mesenchymal Stem Cells.

Cell chirality is extremely important for the evolution of cell morphogenesis to manipulate cell performance due to left-right asymmetry. Although chiral micro- and nanoscale biomaterials have been developed to regulate cell functions, how cell chirality affects cell nanomechanics to command nuclear mechanotransduction was ambiguous. In this study, chiral engineered microcircle arrays were prepared by photosensitive cross-linking synthesis on cell culture plates to control the clockwise/counterclockwise geometric topology of stem cells. Asymmetric focal adhesion and cytoskeleton structures could induce chiral cell nanomechanics measured by atomic force microscopy (AFM) nanoindentation in left-/right-handed stem cells. Cell nanomechanics could be enhanced when the construction of mature focal adhesion and the assembly of actin and myosin cytoskeletons were well organized in chiral engineered stem cells. Curvature angles had a negative effect on cell nanomechanics, while cell chirality did not change cytoskeletal mechanics. The biased cytoskeleton tension would engender different nuclear mechanotransductions by yes-associated protein (YAP) evaluation. The chiral stimuli were delivered into the nuclei to oversee nuclear behaviors. A strong cell modulus could activate high nuclear DNA synthesis activity by mechanotransduction. The results will bring the possibility of understanding the interplay of chiral cell nanomechanics and mechanotransduction in nanomedicines and biomaterials.

Photoregulated supramolecular hydrogels driven by polyradical interactions

Organic radical as a powerful tool has been extensively applied in synthetic chemistry. However, harnessing radical-mediated noncovalent interactions to fabricate soft materials remains elusive. Here we report a new category of supramolecular hydrogel system held by multiple radical-radical (polyradical) interactions, and its photosensitive cross-linking structure. A simple polyacrylamide with triarylamine (TAA) pendants is designed as the precursor. The TAA units in polymer can be converted into active TAA•⁺ radical cations with light and further associate each other via TAA•⁺‒TAA•⁺ stacking interactions to form stable supramolecular network. Temporal control of the light irradiation dictates the degree of radical stacks, thus regulating the mechanical performance of the resulting hydrogel materials on-demand. Moreover, the reversible collapse of this hydrogels can be promoted by adding radical scavenger or exerting reduction voltage.

Graphenoxide Cross-Linker Based Potentiometric Biosensor Design for Sarcosine Determination.

Sarcosine, also known as N-methyl glycine, is a natural amino acid that is an intermediate and by product in glycine synthesis and degradation. Recently found in many peptides, sarcosine has been researched as a newly accepted prostate cancer marker. The increased concentration of sarcosine in blood serum and the urine showed that malignancy of measured prostate cancer cells is active. In this article, we aimed to design a potentiometric biosensor for detection of sarcosine with a low detection limit, high selectivity, short response time, wide linear range, and satisfactory long-term stability. In this article, we developed a new Graphene oxide (GFOX) photosensitive cross-linker based potentiometric biosensor based on the AmiNoAcid (monomer) Decorated and Light Underpinning Conjugation Approach (ANADOLUCA) method. The functional groups determined using Raman, FT-IR, XPS analyzes, and surface characterization, the morphology of synthesized GFOX photosensitive cross-linker were determined by TEM and AFM studies. Then, the performance of the GFOX based potentiometric biosensor has been evaluated. When the usage of the developed GFOX doped potentiometric biosensor against sarcosine determination, it was found that 10-4 mM sarcosine was determined in 60 seconds in the solution. In addition, the detection limit of the GFOX doped potentiometric biosensor was found to be 9.45x10-7 mM, and the linear potentiometric biosensor was found to be in the concentration range of 10-1 to 10-5 mM. The selectivity studies of the developed potentiometric biosensor were investigated using glycine solutions, and it was determined that GFOX doped potentiometric biosensor was more selective against sarcosine. Besides this, a reusability test using 10-3 mM sarcosine solution showed that reproducible studies were performed without the loss of potential of designed potentiometric biosensor and no loss of sensitivity. After applying the framework, we get a new potentiometric biosensor for sarcosine determination. GFOX photosensitive cross-linker was used in designing potentiometric biosensors, and this increased the stability and efficiency of the biosensor. Therefore, the developed potentiometric biosensor for sarcosine determination could be easily used for the early diagnosis of prostate cancer.

Affinity-based protein profiling to reveal targets of puerarin involved in its protective effect on cardiomyocytes

Natural products are an important source of new drugs. Some of them may be used directly in clinical settings without further structural modification. One of these directly used natural products is puerarin (Pue), which protects cardiomyocytes against oxidative stress and high glucose stress. Although Pue has been used in clinics for many years, its direct binding targets involved in the protection of cardiomyocytes are not yet fully understood. Here, we reported that Pue could prevent cardiomyocytes from apoptosis under H2O2 and high glucose conditions. Based on affinity-based protein profiling methods, we synthesized an active Pue probe (Pue-DA) with a photosensitive crosslinker to initiate a biological orthogonal reaction. Because of the steric hindrance of Pue-DA, two conformational isomers (syn and anti) unequivocally existed in the probe, and these transformed into one isomer when the probe was heated at 60 °C. We confirmed that the alkylation was on the 7-position phenol group of Pue. Mass spectroscopy revealed that Pue-DA can bind with three proteins, namely CHAF1B, UBE2C, and UBE2T. Finally, cellular thermal shift assay showed that Pue has the ability to stabilize CHAF1B stabilization. The knock-down of CHAF1B reduced the protective effect of Pue on cardiomyocytes. In conclusion, Pue protects cardiomyocytes from apoptosis through binding with CHAF1B.

Multistimuli-Responsive Optical Hydrogel Nanomembranes to Construct Planar Color Display Boards for Detecting Local Environmental Changes.

Planar metal-insulator-metal (MIM) optical cavities are attractive for biochemical and environmental sensing applications, as they offer a cost-effective cavity platform with acceptable performances. However, localized detection and scope of expansion of applicable analytes are still challenging. Here, we report a stimuli-responsive color display board that can exhibit local spectral footprints, for locally applied heat and alcohol presence. A thermoresponsive, optically applicable, and patternable copolymer, poly(N-isopropylacrylamide-r-glycidyl methacrylate), is synthesized and used with a photosensitive cross-linker to produce a responsive insulating layer. This layer is then sandwiched between two nanoporous silver membranes to yield a thermoresponsive MIM cavity. The resonant spectral peak is blue-shifted as the environmental temperature increases, and the dynamic range of the resonant peak is largely affected by the composition and structure of the cross-linker and the copolymer. The localized temperature increase of silk particles with gold nanoparticles by laser heating can be measured by reading the spectral shift. In addition, a free-standing color board can be transferred onto a curved biological tissue sample, allowing us to simultaneously read the temperature of the tissue sample and the concentration of ethanol. The stimuli-responsive MIM provides a new way to optically sense localized environmental temperature and ethanol concentration fluctuations.

Photoreactive Carboxybetaine Copolymers Impart Biocompatibility and Inhibit Plasticizer Leaching on Polyvinyl Chloride.

Protein and cell interactions on implanted, blood-contacting medical device surfaces can lead to adverse biological reactions. Medical-grade poly(vinyl chloride) (PVC) materials have been used for decades, particularly as blood-contacting tubes and containers. However, there are numerous concerns with their performance including platelet activation, complement activation, and thrombin generation and also leaching of plasticizers, particularly in clinical applications. Here, we report a surface modification method that can dramatically prevent blood protein adsorption, human platelet activation, and complement activation on commercial medical-grade PVC materials under various test conditions. The surface modification can be accomplished through simple dip-coating followed by light illumination utilizing biocompatible polymers comprising zwitterionic carboxybetaine (CB) moieties and photosensitive cross-linking moieties. This surface treatment can be manufactured routinely at small or large scales and can impart to commercial PVC materials superhydrophilicity and nonfouling capability. Furthermore, the polymer effectively prevented leaching of plasticizers out from commercial medical-grade PVC materials. This coating technique is readily applicable to many other polymers and medical devices requiring surfaces that will enhance performance in clinical settings.

Concanavalin A photocross-linked affinity cryogels for the purification of horseradish peroxidase

The present study describes an easy and efficient procedure for the purification of horseradish peroxidase from horseradish roots. For this purpose, supermacroporous cryogels having Concanavalin A were prepared by photosensitive cross-linking polymerization. Horseradish peroxidase binding and elution from the prepared cryogels were carried out changing various parameters such as initial peroxidase concentration and pH. The best binding performance was obtained at pH 7.0. The maximum horseradish peroxidase binding of the cryogels was found to be 3.85 mg g−1cryogel. Horseradish peroxidase purification from crude extract resulted in 115.1-fold. SDS-PAGE analysis and circular dichroism measurements indicated that the horseradish peroxidase purification from horseradish roots was successfully carried out.

Ferritin based bionanocages as novel biomemory device concept

Ferritin is an iron cage having protein, capable of extracting metal ions in their cages and a consequence of the electron transfer of metal ions in their cage by reduction and oxidation processes, electrochemical information storage devices can be designed. In this work, ferritin based protein biomemory substrate has been synthesized by using Amino Acid (monomer) Decorated and Light Underpinning Conjugation Approach (ANADOLUCA) method, which utilizes photosensitive electron transfer based microemulsion co-polymerization as nanobead form of ferritin. Protein substrate contains metal ions such as silver and copper or metal ion pairs namely, silver-copper (Janus bionanocage) and co-polymeric shell of the photosensitive crosslinker protein. The redox behavior of bionanocages differentiates electrochemical "writing" and "erase" states depending on these metal ions (silver or copper) or metal ion pairs. The bionanocages based biomemory substrates have been immobilized using graphene modified glassy carbon electrodes and the memory functions of ferritin based bionanocages have been confirmed by chronoamperometry (CA) and open circuit potential amperometry (OCPA). The stability and durability of multi-state memory devices represent promising properties for future bioelectronic information technologies.

Multistate proteinous biomemory device based on redox controllable hapten cross-linker

A multistate biomemory device consisting of cytochrome c (Cyt-c) photosensitively cross-linked by MACys-Ru(bipyr)2-MACys hapten molecules, which have memory effect through a charge transfer mechanism, has been developed. In this study, it has suggested a highly resolute surface-confined switch composed a signal-enhanced electro-active protein (Cyt-c) co-polymerized on the gold substrates that can be controlled by redox property through Ruthenium based cysteine monomer hapten, MACys-Ru(bipyr)2-MACys as an ANADOLUCA photosensitive cross-linker. The photosensitive cross-linking of the Cyt-c protein on the gold surface topography has been determined by the scanning electron microscopy (SEM). Two state memory functions, writing and erasing of the developed biomemory device, have been investigated by the chronoamperometry (CA) and open-circuit potential amperometry (OCPA). The polymeric proteinous memory device, p(MACys-Ru(bipyr)2-MACys-co-Cyt-c) layer, on the gold electrode is stable and repeatable up to with 104 times continuous cycle.

Simultaneous depletion of albumin and immunoglobulin G by using twin affinity magnetic nanotraps

ABSTRACTFor biomarker discovery, simultaneous removal of albumin and immunoglobulin G (IgG) become more important for both concentrating other species including potential biomarkers and getting rid of their masking effect. In this study, we have proposed a covalent and photosensitive cross-linking conjugation of the biomolecules on nanostructures for the depletion of target proteins from aqueous solution and serum. The effect of concentration, pH, temperature and ionic strength on the adsorption of twin affinity nanotraps based on Cibacron Blue F3GA and Protein A were investigated. The efficiency of albumin and IgG depletion from human serum was performed using sodium dodecylsulfate–polyacrylamide gel electrophoresis.

Layered Double Hydroxide Assemblies with Controllable Drug Loading Capacity and Release Behavior as well as Stabilized Layer-by-Layer Polymer Multilayers.

A stable drug release system with magnetic targeting is essential in a drug delivery system. In the present work, layered double hydroxide assemblies stabilized by layer-by-layer polymer multilayers were prepared by alternative deposition of poly(allylamine hydrochloride) and poly(acrylic acid) species on composite particles of Fe3O4 and ZnAl-LDH and then covalent cross-linkage of the polymer multilayers by photosensitive cross-linker. The successful fabrication was recorded by Zeta potential and Fourier transform infrared spectrum measurements. The formed assemblies were stable in high pH solutions (pH > 7). The drug loading capacity and release behavior of the assemblies could be controlled by treatment with appropriate acidic solution, and were confirmed by loading and release of a simulated drug, methylene blue. The formed assemblies possessed enough saturated magnetic strength and were sensitive to external magnetic field which was essential for targeting drug delivery. The formed assemblies were multifunctional assemblies with great potential as drug delivery system.

Controlled protein delivery from photosensitive nanoparticles

Light provides a powerful approach for delivery of cargos and study of important biological events. This article reports a series of photosensitive and biocompatible delivery of nanoparticles which released proteins upon light irradiation. The nanoparticles were synthesized by emulsion copolymerization of 2-(dimethylamino) ethyl methacrylate with functional monomers and a photoliable o-nitrobenzyl diacrylate crosslinker. Upon mild UV irradiation (λ = 365 nm, 10 mJ/cm(2)), the photosensitive crosslinker underwent light-induced degradation and the sizes of the nanoparticles increased dramatically. The nanoparticles were uptaken by the cells which is confirmed by flow cytometry analysis. Bovine serum albumin and green fluorescent protein were loaded as model proteins into the nanoparticles and accelerated photo-triggered release were achieved.

Photoreactive oligoethylene glycol polymers – versatile compounds for surface modification by thin hydrogel films

Solid surfaces are modified using photo-crosslinkable copolymers based on oligo(ethylene glycol) methacrylate (OEGMA) bearing 2-(4-benzoylphenoxy)ethyl methacrylate (BPEM) as a photosensitive crosslinking unit. Thin films of about 100 nm are formed by spin-coating these a priori highly biocompatible copolymers onto silicon substrates. Subsequent UV-irradiation assures immobilization and crosslinking of the hydrogel films. Their stability is controlled by the number of crosslinker units per chain and the molar mass of the copolymers. The swelling of the hydrogel layers, as investigated by ellipsometry, can be tuned by the crosslinker content in the copolymer. If films are built from the ternary copolymers of OEGMA, BPEM and 2-(2-methoxyethoxy) ethyl methacrylate (MEO2MA), the hydrogel films exhibit a swelling/deswelling transition of the lower critical solution temperature (LCST) type. The observed thermally induced hydrogel collapse is fully reversible and the onset temperature of the transition can be tuned at will by the copolymer composition. Different from analogously prepared thermo-responsive hydrogel films of photocrosslinked poly(N-isopropylacrylamide), the swelling–deswelling transition occurs more gradually, but shows no hysteresis.

Surface Modification of Natural Rubber Latex Film Using Photocrosslinkable Chitosan

Surface modifications of natural rubber latex films were carried out to improve the hydrophilicity by UV-induced immobilization of water-soluble butyryl derivative of chitosan using the photosensitive hetero-bifunctional crosslinking reagent, 4-azidobenzoic acid. The effects of the concentration of modified chitosan solution and UV irradiation time on the water contact angle values of natural rubber latex films were studied. ATR-FTIR analyses confirmed that the O-butylrylchitosan molecules were covalently bonded onto the surface of natural rubber latex film. The water contact angle value of natural rubber latex film which is about 95o reduces to 55o significantly with the introduction of O-butylrylchitosan. The results indicate that the hydrophilic behaviour of natural rubber latex film has been improved by immobilization of O-butylrylchitosan.

Mutual recognition of TNT using antibodies polymeric shell having CdS

Click chemistry is the latest strategy called upon in the development of state of the art exponents of bioconjugation. In this study, we have proposed a covalent and photosensitive crosslinking conjugation of the antibody on nano-structures. For this purpose, quantum dots (QDs) without affecting conformation and function of proteins through the ruthenium-chelate based aminoacid monomer linkages have been applied. The aminoacid-monomer linkages called ANADOLUCA (AmiNoAcid Decorated and Light Underpining Conjugation Approach) give reusable oriented and cross-linked anti 2,4,6-trinitrotoluene (TNT) conjugated QD for TNT detection. In this work, a new and simple method has improved to design and prepare high sensitive nanoconjugates for TNT determination. We have demonstrated the use of luminescent QDs conjugated to antibody for the specific detection of the explosive TNT in aqueous environments. The binding affinity of each nanoconjugates for TNT detection by using Langmuir adsorption methods has also been investigated.

Sunlight stimulated repeated self-healing

energy has been harnessed in many aspects. For example, plants live on photosynthesis induced by sunlight. With respect to the products made from polymers, however, sunlight is recognized to be an undesirable factor responsible for aging. Fortunately, anything which is negative can be turned to positive. In case photoreversible bonds are introduced to macromolecular chains, damages in polymers can be self-healed under sunlight stimulus. Because the use of light is clean, cheap and readily available, such an unconscious way of remending is obviously advantageous. In a recent proof-of-concept experiment of our laboratory [J. Mater. Chem., DOI: 10.1039/C1JM13467A], a novel coumarin derivative was synthesized and attached to a conventional covalently bonded polymer (i.e. polyurethane) as photosensitive crosslinker. Upon mechanical damage, the crosslinkages (i.e. coumarin dimers) of polyurethane network preferentially cleave returning to original coumarin moieties. Then, these coumarin moieties can be reconnected by illumination of 350 nm UV light (a major component of sunlight). The entire cycle is fully reversible. By using this habit, broken polyurethane are repeatedly re-bonded under sunlight, without the necessity of any catalyst, healing agent or surface treatment. The research on photochemical self-healing has only just begun and is still in infancy. In principle, reversible photochemical reaction is hard to proceed in solid polymer for structural application because molecular motion is restricted. The key issues seem to be that (i) the photosensitive segments on the fractured surface should be mobile enough, which favors wetting and diffusion of molecules at the solid-solid contact of the damaged parts to meet the molecular level movement requirement of crack healing; and (ii) macroscopic flow of the material should be prohibited during crack healing, maintaining load-bearing capacity of the material. Repeated damages are very common in daily life, like scratches on cars made by automatic brush washing and those on touch screens by fingernails. Although exterior paint that obscures scratches was developed by automotive industry, it works due to heating of the sun rather than photochemistry. Only fine scratches appearing in the form of plastic deformation can be recovered. In contrast, the sun stimulated photoreversible reaction is able to rebond completely damaged polymeric materials, despite the healing efficiency in terms of mechanical strength restoration is not that satisfactory for the moment. Evidently, the strategy is promising for developing a variety of next generation coatings, films and sheets to meet aesthetic, structural and functional requirements.

Photosensitive controlled release with polyethylene glycol–anthracene modified alginate

Covalent modification of alginate with polyethylene glycol–conjugated anthracene molecules has the potential to both stabilize the alginate and act as a photosensitive crosslinker. Release studies with Coomassie Blue show lengthy release times from the alginate photogels that extend past 70 days with, for example, 17% versus 27% release at 1750 h (73 days) for photogels with and without 365-nm UV light treatment for 30 min at 10 mW/cm 2 in the initial release period. Photocrosslinking of the photogels after loading effectively “locks” in drug compounds to control their release. Effective crosslinking densities and controls of polyethylene glycol-crosslinked alginate and physically crosslinked calcium alginate gels suggest strong interactions between Coomassie Blue and both alginate and anthracene. Photogels containing anthracene-capped star-polyethylene glycol show increased photosensitivity with modified release profiles. Ultimately, the covalent modification of alginate with photoactive crosslinkers has the potential to produce a long-term, photosensitive, controlled release system.

Coumarin imparts repeated photochemical remendability to polyurethane

Coumarin, a traditional perfume substance, serves as a photosensitive crosslinker to construct a polyurethane network. Upon mechanical damage or ultraviolet illumination at a wavelength of 254 nm, the coumarin dimers on the fractured surface that link the polyurethane backbones with one another are cleft, and can be reconnected via irradiation at 350 nm. By using this feature, cracks on the polymer are mended by ultraviolet exposure or even sunlight without the aid of any catalyst. Owing to the reversibility of the photocyclization of coumarin moieties, multiple cycles of crack mending are allowed, as demonstrated by both qualitative and quantitative characterization. This repeated photo-remendability enables the polyurethane to heal the same site in the material more than once, which is of value for practical applications.

Bold relief fabrication by means of electroconvection: basic properties of a suitable mixture

Electroconvection of liquid crystals does not only lead to a large variety of spontaneously formed patterns, but these patterns can also be used as a template for the fabrication of polymer surfaces exhibiting a periodic bold relief. Here, a useful room-temperature nematic mixture consisting of two photostable liquid crystals and a photosensitive cross-linker is presented. Its stability diagram in the frequency-voltage plane, the influence of monomer concentration and the topography of the resulting polymer surfaces are described.