Event Abstract Back to Event Construction of in situ cell separation system by visible light irradiation Chie Kojima1*, Yusuke Nakajima1*, Naoya Oeda1*, Takeshi Kawano2* and Yusuke Taki2* 1 Osaka Prefecture University, Dept. Applied Chemistry, Japan 2 Nikon, Japan Introduction: Separation of functional cells from cultured cells is important for cell engineering and regenerative medicine. We designed a visible light-induced in situ single cell detachment system as an advanced photolithography technology. Gold nanoparticle (AuNP) was embedded in a collagen gel to produce a visible light-responsive cell scaffold. When green light is irradiated at the target cells cultured on the AuNP/collagen hybrid gel, the photo-irradiated collagen gel is thermally denatured into the gelatin sol due to the photothermogenic property of the AuNP. Consequently, the irradiated cells are possibly detached from the scaffold (Fig. 1). In this paper, we prepared AuNP/collagen hybrid gels and performed the in situ single cell detachment by irradiating visible light. Experiments: AuNPs with different diameters were prepared by a seeded growth method using hydroquinone [1]. The collagen sol (Cell matrix IA (Nitta Gelatin)) and the AuNPs were mixed and then induced the gel formation according to the manufacturer’s instruction (Collagen 0.08% and [Au]=0.25mM). Several model cells were cultured on the hybrid gel. Laser (532 nm and 50 mW) was irradiated at the center of target cells for 10 s, 30 s or 60 s, followed by the collection of the medium at the above of the gel. Live/dead assay was performed by using calcein-AM and propidium iodide (PI). Results and Discussion: The average sizes of the AuNPs were estimated by TEM. The most effective photochemical property was observed from the 50 nm-sized AuNP, which was consistent with our previous report [2]. The AuNP with 50 nm was embedded in the collagen gel. The hybrid gel exhibited the photochemical properties due to the AuNP, and dissolved around 50 oC. Some kinds of cells (HeLa cells, MCF-7 cells, MDCK cells, GFP-expressing colon26 cells and so on) were cultured on the gel. No significant cytotoxicity was observed before and after the photoirradiation, indicating that our system was biocompatible. The target cells were surveyed, and the green laser was irradiated by using microscopy equipped with the laser irradiation system. The photo-irradiated cells were detached after the collection of the medium. We have attempted to culture the collected cells further. Conclusion: We succeeded in construction of visible light-induced in situ cell detachment system by using the AuNP and the collagen gel. The materials and the separation system are cell-friendly. This technology is useful for cell separation system of desired cells from the cultured cells, which will become a key technology in a wide range of biomedical studies.
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