Automated Adherent Cell Elimination by a High-Speed Laser Mediated by a Light-Responsive Polymer

Abstract

Conventional cell handling and sorting methods require manual dissociation, which decreases cell quality and quantity and is not well suited for monitoring applications. To purify adherent cultured cells, in situ cell purification technologies that are high throughput and can be utilized in an on-demand manner are expected. Previous demonstrations using direct laser-mediated cell elimination revealed only limited success in terms of their usability and throughput. Here, we developed a Laser-Induced, Light-responsive-polymer-Activated, Cell Killing (LILACK) system that enables high-speed and on-demand adherent cell sectioning and purification. This system employs a visible laser beam, which does not kill cells directly, but induces local heat production through the trans-cis-trans photo-isomerization of azobenzene moieties in only the irradiated area of a light-responsive thin layer. Using this system in each passage for sectioning, human induced pluripotent stem cells (hiPSCs) were maintained their pluripotency and self-renewal during long-term culture. Furthermore, combined with deep machine-learning analysis on fluorescent and phase contrast images, a label-free and automatic cell processing system was developed by eliminating unwanted spontaneously differentiated cells in undifferentiated hiPSC culture conditions.

Downstream bioprocessing of human pluripotent stem cell‐derived therapeutics

“…In addition, it was found that committed cells can be eliminated by a high‐speed laser irradiating specific areas of light‐responsive polymers (i.e. poly[(methylmethacrylate)‐co‐(Disperse Yellow 7 methacrylate)] layers) [26]. Alternatively, laminins of different isoforms promote different adhesion strength of various differentiated cell types.…”

“…Because cell/adhesive surface interactions are regulated upon differentiation, this has led to the development of novel biomaterials for the cell isolation of particular differentiated phenotype [25,26]. At the undifferentiated stage, iPSC colonies can be selectively detached from thermosensitive polymer (PNIPAAm (poly(N-isopropylacrylamide)), at low temperature (i.e.…”