Click‐Chemistry Based Multi‐Component Microarrays by Quill‐Like Pens

We demonstrate the generation of multi‐component spot microarrays by blotting different ink solutions via quill‐like pens. The obtained arrays are immobilized by click‐chemistry in form of the copper(I)‐catalyzed azide‐alkyne cycloaddition and remain stable against washing and immersion in aqueous solution. The average spot radius ranges from 10 to 20 μm and is about an order of magnitude smaller than in currently commercially applied arraying techniques, effectively bridging the gap to high resolution methods as dip‐pen nanolithography and polymer pen lithography. The use of the quill‐like‐pen‐generated spot microarrays as binding assay is demonstrated by capturing of streptavidin from solution and by bioactive sandwich structures from neutravidin and biotin‐labeled fibronectin. Thus, our multi‐component spot microarrays have ideal dimensions and biochemical properties to accommodate (single) cells. Additionally, the building up of the cell‐recruiting protein sandwich structure on top of the basic spot microarray allows for the highly selective adhesion of fibroblasts. This results then in ordered (single) cell arrays, demonstrating the bio‐compatibility and high throughput of this multi‐component spot microarray platform.     

Protein‐Corona‐by‐Design in 2D: A Reliable Platform to Decode Bio–Nano Interactions for the Next‐Generation Quality‐by‐Design Nanomedicines

Hard corona (HC) protein, i.e., the environmental proteins of the biological medium that are bound to a nanosurface, is known to affect the biological fate of a nanomedicine. Due to the size, curvature, and specific surface area (SSA) 3‐factor interactions inherited in the traditional 3D nanoparticle, HC‐dependent bio–nano interactions are often poorly probed and interpreted. Here, the first HC‐by‐design case study in 2D is demonstrated that sequentially and linearly changes the HC quantity using functionalized graphene oxide (GO) nanosheets. The HC quantity and HC quality are analyzed using NanoDrop and label‐free liquid chromatography–mass spectrometry (LC‐MS) followed by principal component analysis (PCA). Cellular responses (uptake and cytotoxicity in J774 cell model) are compared using imaging cytometry and the modified lactate dehydrogenase assays, respectively. Cellular uptake linearly and solely correlates with HC quantity (R² = 0.99634). The nanotoxicity, analyzed by retrospective design of experiment (DoE), is found to be dependent on the nanomaterial uptake (primary), HC composition (secondary), and nanomaterial exposure dose (tertiary). This unique 2D design eliminates the size–curvature–SSA multifactor interactions and can serve as a reliable screening platform to uncover HC‐dependent bio–nano interactions to enable the next‐generation quality‐by‐design (QbD) nanomedicines for better clinical translation.     

基于点击化学的壳聚糖水凝胶的制备及性能

本研究首先在甲磺酸中用丙烯酰氯对壳聚糖(CS)进行接枝改性,通过一步法合成了水溶性丙烯酰基壳聚糖(CS-AC).之后,在光引发剂I2959和紫外光(UV)照下,以二硫苏糖醇(DTT)为交联剂制备了基于巯基-烯点击化学的CS-AC/DTT快速交联水凝胶.红外光谱(FTIR)和核磁共振氢谱(1 H-NMR)结果定性和定量地...     

Bio‐Orthogonal T Cell Targeting Strategy for Robustly Enhancing Cytotoxicity against Tumor Cells

T cells can kill tumor cells by cell surface immunological recognition, but low affinity for tumor‐associated antigens could lead to T cell off‐target effects. Herein, a universal T cell targeting strategy based on bio‐orthogonal chemistry and glycol‐metabolic engineering is introduced to enhance recognition and cytotoxicity of T cells in tumor immunotherapy. Three kinds of bicycle [6.1.0] nonyne (BCN)‐modified sugars are designed and synthesized, in which Ac₄ManN‐BCN shows efficient incorporation into wide tumor cells with a BCN motif on surface glycans. Meanwhile, activated T cells are treated with Ac₄GalNAz to introduce azide (N₃) on the cell surface, initiating specific tumor targeting through a bio‐orthogonal click reaction between N₃ and BCN. This artificial targeting strategy remarkably enhances recognition and migration of T cells to tumor cells, and increases the cytotoxicity 2 to 4 times for T cells against different kinds of tumor cells. Surprisingly, based on this strategy, the T cells even exhibit similar cytotoxicity with the chimeric antigen receptor T‐cell against Raji cells in vitro at the effector: target cell ratios (E:T) of 1:1. Such a universal bio‐orthogonal T cell‐targeting strategy might further broaden applications of T cell therapy against tumors and provide a new strategy for T cell modification.     

Microscale Electro‐Hydrodynamic Cell Printing with High Viability

Cell printing has gained extensive attentions for the controlled fabrication of living cellular constructs in vitro. Various cell printing techniques are now being explored and developed for improved cell viability and printing resolution. Here an electro‐hydrodynamic cell printing strategy is developed with microscale resolution (<100 µm) and high cellular viability (>95%). Unlike the existing electro‐hydrodynamic cell jetting or printing explorations, insulating substrate is used to replace conventional semiconductive substrate as the collecting surface which significantly reduces the electrical current in the electro‐hydrodynamic printing process from milliamperes (>0.5 mA) to microamperes (<10 µA). Additionally, the nozzle‐to‐collector distance is fixed as small as 100 µm for better control over filament deposition. These features ensure high cellular viability and normal postproliferative capability of the electro‐hydrodynamically printed cells. The smallest width of the electro‐hydrodynamically printed hydrogel filament is 82.4 ± 14.3 µm by optimizing process parameters. Multiple hydrogels or multilayer cell‐laden constructs can be flexibly printed under cell‐friendly conditions. The printed cells in multilayer hydrogels kept alive and gradually spread during 7‐days culture in vitro. This exploration offers a novel and promising cell printing strategy which might benefit future biomedical innovations such as microscale tissue engineering, organ‐on‐a‐chip systems, and nanomedicine.     

Metal‐Free Click Chemistry Reactions on Surfaces

In the last decade, interest in the functionalization of surfaces and materials has increased dramatically. In this regard, click chemistry deserves a central focus because of its mild reaction conditions, high efficiency, and easy post‐treatment. Among such novel click reactions, those that do not require any metal catalyst are of special interest, as metals may have undesirable effects in many fields. In this Review, the backgrounds and application of such metal‐free click reactions for the modification of surfaces are highlighted.     

3D Scaffolds to Study Basic Cell Biology

Mimicking the properties of the extracellular matrix is crucial for developing in vitro models of the physiological microenvironment of living cells. Among other techniques, 3D direct laser writing (DLW) has emerged as a promising technology for realizing tailored 3D scaffolds for cell biology studies. Here, results based on DLW addressing basic biological issues, e.g., cell‐force measurements and selective 3D cell spreading on functionalized structures are reviewed. Continuous future progress in DLW materials engineering and innovative approaches for scaffold fabrication will enable further applications of DLW in applied biomedical research and tissue engineering.