Tetrakis(hydroxymethyl)
Phosphonium Chloride as a
Covalent Cross-Linking Agent for Cell Encapsulation within Protein-Based
Hydrogels
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Abstract
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