Professor University of California, Los Angeles, United States
Introduction: Microparticles with defined 3D shapes and spatially-tailored chemical functionality enable new opportunities in biotechnology at the scale of cells and tissues. The current approach to manufacture these shaped particles requires precise injection of multiple polymer precursors into a flow focusing microfluidic device which has a limited scalability. Further, these previous approaches lacked the ability to spatially pattern different chemical functionalities, limiting the flexibility of the platform. In this report, we introduce a new induced-phase separation concept to overcome tradeoffs between particle complexity and fabrication throughput for the manufacture of microparticles with tunable localized surface chemistry and shape (Figure 1).
Materials and
Methods: Phase separation of polyethylene glycol (PEG) and gelatin is dependent on the temperature of the system due to the conformational change of gelatin upon cooling. Using a microfluidic droplet generator, we constructed two isothermal binodal curves, corresponding to which PEG and gelatin undergo phase transitions at 4 and 22 °C respectively. By using compositions of the PEG/gelatin solutions located at points between the two binodal curves a transition is enabled from a miscible solution to a phase-separated state induced by the temperature change (Figure 2). Using the conditional phase separation of PEG and gelatin, photocrosslinkable PEG and gelatin ATPS droplets were generated and crosslinked with UV light to form uniform microparticles. The morphology of the droplets and resulting particles were controlled by changing the composition of PEG/gelatin solution. We used fluorescein isothiocyanate (FITC)-conjugated gelatin to demonstrate the particles were selectively functionalized on their inner cavities with higher densities of gelatin, which we showed was beneficial for their application as a platform for cell-carriers and reaction vessels to perform single-cell assays (Figure 3A).
Results, Conclusions, and Discussions: We manufactured hydrogel microparticles at rates of greater than 40 million/hour with localized surface chemistry using a parallelized step emulsification device and temperature-induced phase-separation. The localized gelatin on the cavity promoted deterministic attachment of cells only within the cavities via integrin binding (Figure 3B). Cells adhered to nanovials could be sorted using standard fluorescence activated cell sorting (FACS) and higher cell viability was achieved for cells attached to nanovials compared to unbound cells, implying that these nanovials provide protection from fluid shear stresses during the sorting process (Figure 3C). Localized gelatin was modified with different biomolecules to capture secretions from encapsulated cells. Secretion assay with human IgG producing chinese hamster ovary (CHO) cells demonstrate the ability to capture secretions on particles containing cells without crosstalk to neighboring particles (Figure 4).
We overcome previous barriers to achieve scalable production of hydrogel microparticles with localized surface chemistry using a parallelized step emulsification device and temperature-induced phase-separation. The engineered particles was utilized as individual cell carriers modified to run assays on single cells, opening of numerous applications in functional screening of cells at throughputs of 100,000s of single cells.
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