van Zee, M. | bioRxiv Science

bioRxiv2021

High-throughput selection of microalgae based on biomass accumulation rates in production environments using PicoShell Particles

Production of high-energy lipids by microalgae may provide a sustainable, renewable energy source that can help tackle climate change. However, microalgae engineered to produce more lipids usually grow slowly, leading to reduced overall yields. Unfortunately, tools that enable the selection of cells based on growth while maintaining high biomass production, such as well-plates, water-in-oil droplet emulsions, and nanowell arrays do not provide production-relevant environments that cells experience in scaled-up cultures (e.g. bioreactors or outdoor cultivation farms). As a result, strains that are developed in the lab often do not exhibit the same beneficial phenotypic behavior when transferred to industrial production. Here we introduce PicoShells, picoliter-scale porous hydrogel compartments, that can enable >100,000 individual cells to be compartmentalized, cultured in production-relevant environments, and selected based on growth and biomass accumulation traits using standard flow cytometers. PicoShells consist of a hollow inner cavity where cells are encapsulated, and a porous outer shell that allows for continuous solution exchange with the external environment so that nutrients, cell-communication factors, and cytotoxic cellular byproducts can transport freely in and out of the inner cavity. PicoShells can also be placed directly into shaking flasks, bioreactors, or other production-relevant environments. We experimentally demonstrate that Chlorella sp. and Saccharomyces cerevisiae grow to significantly larger colony sizes in PicoShells than in water-in-oil droplet emulsions (P < 0.05). We have also demonstrated that PicoShells containing faster biomass accumulating Chlorella clonal colonies can be selected using a fluorescence-activated cell sorter and re-grown. Using the PicoShell process, we select a Chlorella population that accumulates biomass 8% faster than does an un-selected population after a single selection cycle.

bioengineering↗

bioRxiv2020

Massively parallel encapsulation of single cells with structured microparticles and secretion-based flow sorting

Techniques to analyze and sort single cells based on functional outputs, such as secreted products, have the potential to transform our understanding of cellular biology, as well as accelerate the development of next generation cell and antibody therapies. However, secreted molecules rapidly diffuse away from cells, and analysis of these products requires specialized equipment and expertise to compartmentalize individual cells and capture their secretions. Herein we demonstrate the use of suspendable microcontainers to sort single viable cells based on their secreted products at high-throughput using only commonly accessible laboratory infrastructure. Our microparticles act as solid supports which facilitate cell attachment, partition uniform aqueous compartments, and capture secreted proteins. Using this platform, we demonstrate high-throughput screening of stably- and transiently-transfected producer cells based on relative IgG production as well as screening of B lymphocytes and hybridomas based on antigen-specific antibody production using commercially available flow sorters. Leveraging the high-speed sorting capabilities of standard sorters, we sorted >1,000,000 events in less than an hour. The reported microparticles can be easily stored, and distributed as a consumable reagent amongst researchers, democratizing access to high-throughput functional cell screening.

bioengineering↗