Native tissues provide cells with complex, three-dimensional (3D) environments comprised of hydrated networks of extracellular matrix proteins and sugars. By mimicking the dimensionality of native tissue while deconstructing the effects of environmental parameters, protein-based hydrogels serve as attractive, in vitro platforms to investigate cell–matrix interactions. For cell encapsulation, the process of hydrogel formation through physical or covalent cross-linking must be mild and cell compatible. While many chemical cross-linkers are commercially available for hydrogel formation, only a subset are cytocompatible; therefore, the identification of new and reliable cytocompatible cross-linkers allows for greater flexibility of hydrogel design for cell encapsulation applications. Here, we introduce tetrakis(hydroxymethyl) phosphonium chloride (THPC) as an inexpensive, amine-reactive, aqueous cross-linker for 3D cell encapsulation in protein-based hydrogels. We characterize the THPC-amine reaction by demonstrating THPC's ability to react with primary and secondary amines of various amino acids. In addition, we demonstrate the utility of THPC to tune hydrogel gelation time (6.7 ± 0.2 to 27 ± 1.2 min) and mechanical properties (storage moduli ∼250 Pa to ∼2200 Pa) with a recombinant elastin-like protein. Lastly, we show cytocompatibility of THPC for cell encapsulation with two cell types, embryonic stem cells and neuronal cells, where cells exhibited the ability to differentiate and grow in elastin-like protein hydrogels. The primary goal of this communication is to report the identification and utility of tetrakis(hydroxymethyl) phosphonium chloride (THPC) as an inexpensive but widely applicable cross-linker for protein-based materials.

中文翻译：

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四（羟甲基）氯化鏻作为共价交联剂用于蛋白质基水凝胶中的细胞包封

天然组织为细胞提供了复杂的三维 (3D) 环境，该环境由细胞外基质蛋白和糖的水合网络组成。通过模拟天然组织的维度，同时解构环境参数的影响，基于蛋白质的水凝胶可作为有吸引力的体外平台来研究细胞-基质相互作用。对于细胞封装，通过物理或共价交联形成水凝胶的过程必须温和且与细胞相容。虽然许多化学交联剂可用于水凝胶形成，但只有一部分是细胞相容的；因此，新的和可靠的细胞相容性交联剂的鉴定为细胞封装应用的水凝胶设计提供了更大的灵活性。这里，我们介绍了四（羟甲基）氯化鏻（THPC）作为一种廉价的、胺反应性的、水性交联剂，用于在基于蛋白质的水凝胶中封装 3D 细胞。我们通过证明 THPC 与各种氨基酸的伯胺和仲胺反应的能力来表征 THPC-胺反应。此外，我们证明了 THPC 用重组弹性蛋白样蛋白调节水凝胶凝胶时间（6.7 ± 0.2 至 27 ± 1.2 分钟）和机械性能（储存模量~250 Pa 至~2200 Pa）的效用。最后，我们展示了 THPC 与两种细胞类型（胚胎干细胞和神经元细胞）的细胞封装的细胞相容性，其中细胞表现出在弹性蛋白样蛋白水凝胶中分化和生长的能力。