Die Erfindung betrifft ein Verfahren und eine Vorrichtung zur Reprogrammierung von lebenden Zellen.

Die Aufgabe, eine neue Möglichkeit zur direkten, effizienten und schnellen Zell-Reprogrammierung ohne den Einsatz von Viren zu finden wird erfindungsgemäß beim Verfahren zur Zell-Reprogrammierung, bei dem ein Cocktail aus mindestens zwei Transkriptionsfaktoren und einer Mikro-RNS ins Innere mindestens einer Zelle eingeschleust werden soll, um diese in iPS-Zellen oder einen anderen Zelltyp umzuwandeln, gelöst durch Einlagern der zu reprogrammierenden Zellen in einer wässrigen Umgebung des Cocktails ohne viralen Träger und Richten eines Femtosekunden-Lasers mit einer Impulswiederholfrequenz im Bereich zwischen 50 MHz und 2 GHz, einer Wellenlänge im Bereich von 700 bis 1200 nm mittels eines Laser-Scanning-Mikroskops mit einer numerischen Apertur zwischen 0,9 und 1,5 auf eine Zellmembran der zu reprogrammierender Zelle in einen Fokus und Steuern der Position des Fokus, der Belichtungsdauer und der Laserleistung für die optische Bearbeitung der Zelle, sodass der Fokus in Abhängigkeit von der Impulsfolgefrequenz mit einer Leistung zwischen 7 mW und 100 mW eine transiente kleinporige Öffnung mit einer Größe im Bereich von bis zu 500 nm erzeugt, um eine Diffusion des Cocktails zur multiplen Reprogrammierung der Zelle durch optische multiple Reprogrammierung zu erreichen.

The present invention addresses the problem of providing a vascular progenitor cell sheet derived from induced pluripotent stem cells, which has the strength to tolerate practical applications and exhibits a high treatment effect. This vascular progenitor cell sheet derived from induced pluripotent stem cells is prepared by performing: (1) a step for preparing magnetically labeled Flk-1 positive cells derived from induced pluripotent stem cells; (2) a step for preparing a mixture of the Flk-1 positive cells and a gel material including type I collagen, laminin, type IV collagen and entactin as active ingredients, and then disseminating the mixture in a culture vessel; (3) a step for drawing the Flk-1 positive cells in the mixture to the culture surface of the culture vessel by application of a magnetic force to form a multi-layered cell layer; and (4) a step for gelling the gel material.

Provided are a culture method and a culture device for efficiently producing cells and/or cell tissue suitable for medication applications. A culture method of culturing cells adhered onto an electrode layer (30), the method comprising: a first step of adhering the cells onto the electrode layer and culturing the cells thereon, the electrode layer (30) being coated with an oligopeptide (10) including: a terminal amino acid (11) forming one end thereof to be bound to the electrode layer via thiolate; a cell adhesive sequence (14) forming another end thereof and having a cell-adhesive amino acid sequence; and an alternating sequence (13) to be bound to the one end side of the cell-adhesive sequence (14), the alternating sequence including a plurality of acidic amino acids (13a) and a plurality of basic amino acids (13b), being alternately bound to each other one by one; and a second step of applying, to the electrode layer (30) having the cells adhered thereonto, an electrical potential inducing reductive desorption of the oligopeptide (10), to thereby detach the cells from the electrode layer (30).

A new means of separately arranging individual animal cells on a substrate surface is provided. A method for preparing a substrate for arranging animal cells in an array, comprising steps (1) to (3): (1) preparing a substrate having adsorption surfaces in an array on an electrode substrate surface; (2) causing an extracellular matrix to adsorb to the electrode surface and the adsorption surfaces of the substrate; and (3) applying a weak potential to the electrode to cause the extracellular matrix on the electrode surface to separate to obtain a substrate with the extracellular matrix adhered to the adsorption surfaces thereof. A method for preparing a substrate on which animal cells have been arranged in an array, comprising steps (1) to (4): (1) preparing a substrate having adsorption surfaces in an array on an electrode substrate surface; (2) causing an extracellular matrix to adsorb to the electrode surface and the adsorption surfaces of the substrate; (3) applying a weak potential to the electrode to cause the extracellular matrix on the electrode surface to separate to obtain a substrate with extracellular matrix adhered to the adsorption surfaces thereof; and (4) culturing the animal cells on the surface of the substrate obtained in (3) to obtain a substrate on which the animal cells have adhered to the adsorption surface.