Tetrakis(hydroxymethyl) Phosphonium Chloride as a Covalent Cross-Linking Agent for Cell Encapsulation within Protein-Based Hydrogels
Open accessAbstract
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.
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Chung and colleagues generated a hydrogel using an elastin-like polypeptide matrix containing a fibronectin-derived RGD cell-binding domain with THPC as an amine-reactive crosslinker; this material transiently inhibited contractility of murine ESC-derived cardiomyocytes and enhanced survival of dorsal root ganglia cells from chick embryos.
“Chung and colleagues generated a hydrogel using an elastin-like polypeptide matrix containing a fibronectin-derived RGD cell-binding domain with tetrakis(hydroxymethyl)phosphonium chloride (THPC) as an amine-reactive crosslinker; this material transiently inhibited contractility of murine ESC-derived cardiomyocytes and enhanced survival of dorsal root ganglia cells from chick embryos189.”
From Towards organoid culture without Matrigel (2021) · cites this paper as [189]
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Towards organoid culture without Matrigel
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- Authors
- Chung, C., Lampe, K. J., & Heilshorn, S. C.
- Affiliations
- Stanford University (US)
- Corresponding author
- Heilshorn, Sarah C.
- Venue
- Biomacromolecules
- Publisher
- American Chemical Society (ACS)
- Published
- 2012-11-26
- Volume
- 13
- Issue
- 12
- Pages
- 3912-3916
- ISSN
- 1525-7797 (ISSN-L)
- 1526-4602
- PMC
- PMC3556456
- OpenAlex
- W2084513647
- Cited by
- 132
- References
- 23
- Citations by year
- 2026: 4 · 2025: 10 · 2024: 7 · 2023: 9 · 2022: 9 · 2021: 11 · 2020: 12 · 2019: 6 · 2018: 16 · 2017: 18 · 2016: 10 · 2015: 7 · 2014: 8 · 2013: 5
- FWCI
- 2.95
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Biomacromolecules (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- Funders
- American Heart Association
- Massachusetts Institute of Technology
- National Institutes of Health (5r21ar062359-02, 1dp2od006477-01)
- National Institute of Neurological Disorders and Stroke (F32NS076222)
- National Institute of Arthritis and Musculoskeletal and Skin Diseases (R21-AR062359)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Medicine › Hydrogels: synthesis, properties, applications
- Physical Sciences › Engineering › Biomedical Engineering › 3D Printing in Biomedical Research
- Physical Sciences › Materials Science › Biomaterials › Silk-based biomaterials and applications
- Keywords
- tetrakis(hydroxymethyl)phosphonium chloride, covalent cross-linking, protein hydrogels, cell encapsulation, elastin-like proteins, hydrogel mechanical properties, cytocompatibility, embryonic stem cells, neurons, cell differentiation, extracellular matrix mimicry
- MeSH terms
- Animals, Biocompatible Materials, Cell Line, Cross-Linking Reagents, Elastin, Organophosphorus Compounds, Recombinant Proteins, Cells, Immobilized, Hydrogels, Mice
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