An Automation Workflow for High‐Throughput Manufacturing and Analysis of Scaffold‐Supported 3D Tissue Arrays

Abstract

Patient‐derived organoids have emerged as a useful tool to model tumour heterogeneity. Scaling these complex culture models while enabling stratified analysis of different cellular sub‐populations, however, remains a challenge. One strategy to enable higher throughput organoid cultures is the scaffold‐supported platform for organoid‐based tissues (SPOT). SPOT allows the generation of flat, thin, and dimensionally‐defined microtissues in both 96‐ and 384‐well plate footprints that are compatible with longitudinal image‐based readouts. SPOT is currently manufactured manually, however, limiting scalability. In this study, an automation approach to engineer tumour‐mimetic 3D microtissues in SPOT using a liquid handler is optimized and comparable within‐ and between‐sample variation to standard manual manufacturing is shown. Further, a liquid handler‐supported cell extraction protocol to support single‐cell‐based end‐point analysis using high‐throughput flow cytometry and multiplexed cytometry by time of flight is developed. As a proof‐of‐value demonstration, 3D complex tissues containing different proportions of tumour and stromal cells are generated to probe the reciprocal impact of co‐culture. It is also demonstrated that primary patient‐derived organoids can be incorporated into the pipeline to capture patient‐level tumour heterogeneity. It is envisioned that this automated 96/384‐SPOT workflow will provide opportunities for future applications in high‐throughput screening for novel personalized therapeutic targets.

An Engineered 3D Co-culture Model of Primary Macrophages and Patient-Derived Tumour Cells to Explore Cellular Responses in the Graded Hypoxic Microenvironment of Pancreatic Cancer

“…However, because macrophages experienced some activation from the TRACER scaffold, culturing macrophages as monocultures in TRACER without any exposure to external stimuli (e.g., activating cytokines, co-culture with another cell type) should be interpreted cautiously. Nevertheless, here we show that culturing primary macrophages in scaffold-supported cultures such as TRACER is possible and importantly, could be integrated into other similar models with different applications, such as the SPOT model developed by our group for high-throughput cell characterization 117,118 or those developed by the Whitesides and Lockett group 119,120 .…”

Optimization of liquid handling parameters for viscous liquid transfers with pipetting robots, a “sticky situation”

“…that currently rely on manual optimization of the liquid handling parameters. 15,16 Our protocol would benefit by increasing the level of automation of these tasks and improving the repeatability of the assays.…”

Digital light processing-based bioprinting of microtissue hydrogel arrays using dextran-induced aqueous emulsion ink

“…[2][3][4] Fortunately, advancements in tissue engineering technology have created more sophisticated in vitro 3D cellular models, such as microtissue arrays and 3D printing, which are becoming more commonly used. [5][6][7][8][9] The widespread use of these 3D models provides a strong foundation for preclinical drug research. Among them, the microtissue array is a microscale three-dimensional bionic model carrying cells, 10,11 which can replicate the function of an organ at its most fundamental level.…”