Induction of Cell–Cell Connections by Using in situ Laser Lithography on a Perfluoroalkyl‐Coated Cultivation Platform

Abstract

This article describes a novel laser-directed microfabrication method carried out in aqueous solution for the organization of cell networks on a platform. A femtosecond (fs) laser was applied to a platform culturing PC12, HeLa, or normal human astrocyte (NHA) cells to manipulate them and to facilitate mutual connections. By applying an fs-laser-induced impulsive force, cells were detached from their original location on the plate, and translocated onto microfabricated cell-adhesive domains that were surrounded with a cell-repellent perfluoroalkyl (R(f)) polymer. Then the fs-laser pulse-train was applied to the R(f) polymer surface to modify the cell-repellent surface, and to make cell-adhesive channels of several μm in width between each cell-adhesive domain. PC12 cells elongated along the channels and made contact with others cells. HeLa and NHA cells also migrated along the channels and connected to the other cells. Surface analysis by X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM) confirmed that the R(f) polymer was partially decomposed. The method presented here could contribute not only to the study of developing networks of neuronal, glial, and capillary cells, but also to the quantitative analysis of nerve function.

Dynamic photopatterning of cellsin situby Q-switched neodymium-doped yttrium ortho-vanadate laser

“…However, this laser failed to induce photochemical ablation of used MPC polymer alone, whereas Ti:sapphire laser could. 27 Used gold coating absorbed laser photons induced a thermal effect to ablate the over-layered cytophobic MPC polymer. This work developed an in-situ engraving approach, which was easy to handle, faster in operation, and cost effective.…”

“…Laser applications occasionally cause hypertrophy of the cells and can damage the cell if the laser focal spot is too close to the cell. 27,31 Usually, application of an IR laser reduces the probability of cell damage in comparison with using the UV laser. [32][33][34] However, during in-situ engraving, the safe distance between the laser spot and the pre-existing cells had to be ensured.…”

“…26 Previously our group developed a femto-second laser induced in-situ micro-fabrication method to array cells on a platform with micrometer-scale cell-adhering domains on a cytophobic surface under culture conditions. 27,28 The method was based on photochemical ablation. 28 In terms of promoting the use of in-situ lithography in biological research, frequent tuning requirements and high costs of the femto-second laser are prohibitive.…”

“…28 In terms of promoting the use of in-situ lithography in biological research, frequent tuning requirements and high costs of the femto-second laser are prohibitive. Additionally, the engraving speed of Ti: sapphire is only around 20 μm∕s with a pulse frequency of 1 kHz, 27 which is too slow to make a pattern in a platform of 3 to 4 cm 2 . This work presents a novel dynamic in-situ ablation method for photopatterning of cells in a specially designed platform.…”

Surface Modification of Cell Scaffold in Aqueous Solution Using TiO₂Photocatalysis and Linker Protein L2 for Patterning Primary Neurons<i></i>

“…Such "in-situ"or "dynamic" patterning has been realized, for example, by using photoresponsive polymers [23,24] or laser ablation [25,26], just to name a few. We previously reported on the application of laser ablation phenomena to create growth pathways of neuronal processes (neurites) at sub-cellular resolution in situ [26].…”