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Laser-induced cell injury in closed micro physiological systems: A novel method to study regeneration processes in vitro

: Schmieder, Florian; Förster, Deborah; Rösler, Mechthild; Sradnick, Jan; Klotzbach, Udo; Hohenstein, Bernd; Sonntag, Frank

Postprint urn:nbn:de:0011-n-5821996 (680 KByte PDF)
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Created on: 27.3.2020

Klotzbach, Udo (Ed.) ; Society of Photo-Optical Instrumentation Engineers -SPIE-, Bellingham/Wash.:
Laser-based Micro- and Nanoprocessing XIV : 1-6 February 2020, San Francisco, California
Bellingham, WA: SPIE, 2020 (Proceedings of SPIE 11268)
Paper 1126804, 7 pp.
Conference "Laser-Based Micro- and Nanoprocessing" <14, 2020, San Francisco/Calif.>
Bundesministerium fur Wirtschaft und Energie BMWi (Deutschland)
IGF; 19175 BR/1; ZEBRA
Conference Paper, Electronic Publication
Fraunhofer IWS ()
cell damage; microfluidic; Lab-on-a-Chip; MPS; micro physiological systems; regeneration; Laser; cell injury

Closed micro-physiological systems (MPS) are miniaturized, chip-sized platforms that can be used as cellularized organoid systems to study cellular processes like migration, regeneration or proliferation in vitro. Due to the limited accessibility of the cells inside of closed MPS, the establishment of a well-defined mechanism to induce specific cell damage is difficult. Here we present a novel laser based method to induce well-defined lesions in closed cell layers. This could be a novel tool to study cellular mechanisms of different cell types after injury. The present project aimed to establish well-defined lesion in cellular layers without removing the dead cells and the molecular signaling that is caused by apoptosis. Considering that, we constructed a MPS that was produced by layer laminate manufacturing. According to the experimental needs, the MPS contains two fluidic circuits which include reservoirs, channels, and an integrated micro pump. To establish the method, blood endothelial outgrowth cells (BOEC) were seeded into the MPS previously coated with collagen (5µg/cm²) at a density of approximately 7,5x104 cells/cm². After 3 hours of attachment, a pulsatile flow was applied to the channels. When the whole channel was covered with a BOEC monolayer, laser ablation took place between day 3 and 6 after seeding. To induce the selective cell injury we used a JenLas® laser that was optically integrated into an inverted microscope. The irradiation took several seconds with a wavelength of 532 nm. The damage and the following regeneration processes were observed by fluorescence microscopy using LIVE/DEAD® Viability/Cytotoxicity Kit and Time Lapse recording.