All-aqueous Synthesis of Fibrillated Protein Microcapsules for Membrane-bounded Culture of Tumor Spheroids

27 February 2023, Version 1
This content is a preprint and has not undergone peer review at the time of posting.


Water-in-water (W/W) emulsion provides cytocompatible compartments for cell encapsulation and 3D printing of tissues. Formation of semi-permeable membrane at the W/W interface is critical to entrapment of cells in the droplet phase and supply of nutrients from the continuous phase. However, the adsorption of colloidal particles at the W/W interface is dynamic and reversible, which represents an inherent limitation for fabrication of mechanically robust cell-laden microcapsules. Here we demonstrate the preparation of thin, inflatable and semi-permeable microcapsules by using clusters of protein fibrils as building blocks and control their assembly at W/W interface. These fibril clusters are prepared by cross-linking lysozyme fibrils with multi-arm polyethylene glycol (PEG) through click chemistry. Compared to linear-structured fibrils, fibril clusters can strongly adsorb at W/W interface, and they packed into an interconnected meshwork to stabilize W/W emulsion. Moreover, when fibril clusters are complexed with calcium alginate, the hybrid microcapsules can bear a high osmotic gradient of 60 mOsm/Kg induced by dextran (Mw=500,000), and their surface area expand by over 100% without rupture. This all-aqueous biomaterial synthesis approach allows fabrication of mechanically robust capsules for long-time culture of SGC-996 tumor spheroids, with great potentials to be used in anti-tumor drug-screening and tissue transplantation.


water-in-water emulsion
fibril cluster
protein microcapsules
tumor spheroids

Supplementary materials

Supporting Information
The supporting information includes the fluorescence microscopy images and CLSM images of emulsion droplets, rupture of microcapsules in diluted trypsin-EDTA, and the cell-laden microcapsules produced through an electrospray approach with different cell densities.


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