# Scale-down optimization of a robust, parallelizable human induced pluripotent stem cell bioprocess for high-throughput research

## TL;DR

Researchers developed a cost-effective, scalable method to grow human induced pluripotent stem cells in small volumes for efficient high-throughput studies.

## Contribution

A novel scale-down bioprocess for hiPSC expansion that maintains pluripotency and enables high-throughput research.

## Key findings

- Scale-down systems using preformed aggregates achieve comparable cell expansion to commercial systems.
- Pluripotency markers and differentiation capacity are preserved in scale-down cultures.
- The protocol enables rapid, cost-effective research at less than 20 mL scales.

## Abstract

•Preformation of aggregates tuned by cell density enable cultivation of hiPSCs in scale-down shear environments.•Scale-down systems utilizing preformation protocols achieve comparable fold expansion with commercial systems.•Expression of pluripotency markers and functional differentiation capacity is maintained following passage in scale-down culture.•Successful application of hiPSC protocols at < 20 mL scales enable rapid and cost-effective research into cell phenotype under dynamic conditions.

Preformation of aggregates tuned by cell density enable cultivation of hiPSCs in scale-down shear environments.

Scale-down systems utilizing preformation protocols achieve comparable fold expansion with commercial systems.

Expression of pluripotency markers and functional differentiation capacity is maintained following passage in scale-down culture.

Successful application of hiPSC protocols at < 20 mL scales enable rapid and cost-effective research into cell phenotype under dynamic conditions.

Human induced pluripotent stem cell (hiPSC) derived therapeutics require clinically relevant quantities of high-quality cell populations for applications in regenerative medicine. The lack of efficacy exhibited across clinical trials suggests deeper understanding of the networks governing phenotype is needed. Further, costs limit study throughput in characterizing the artificial niche relative to outcomes. We present herein an optimized strategy to enable high-throughput hiPSC expansion at <20 mL research scale. We assessed viability of single cell inoculation and aggregate preformation to facilitate proliferation. We modeled aggregate characteristics against agitation rate. Our results demonstrate tunable control with fold expansion comparable to commercial systems. Marker quantification and teratoma assay confirm functional pluripotency. This approach constitutes a scalable protocol to accelerate hiPSC research, and a significant step in advancing the rate of progress in elucidating links to derivative functionality. This work will enable statistically rigorous studies targeting hiPSC and downstream phenotype for clinical manufacturing.

Implementation of adapted protocols enable scale-down systems as a tool for high-throughput iPSC biomanufacturing research, in platforms conducive to scale-up for clinical manufacturing.Image, graphical abstract