Adherent cell culture method

Abstract

Provided are a method capable of evaluating adherent cells under an environment similar to an in vivo environment by a culture method similar to a two-dimensional culture, and applications thereof. An adherent cell culture method uses, as a culture chamber ( 10 ), a chamber in which two or more culture spaces each having an equivalent diameter (D) that is 1 to 5 times the diameter of a desired spheroid and each having a height (H) that is 0.3 to 5 times the equivalent diameter are arranged and a surface of each of the culture spaces has a water contact angle of 45 degrees or less. Spheroids of adherent cells are cultured in the respective culture spaces ( 11 ) arranged in the culture chamber ( 10 ).

1 . A method of culturing adherent cells comprising:
 culturing adherent cells in a culture chamber to form a plurality of spheroids, the culture chamber comprising a plurality of culture spaces with each culture space comprising a wall partitioning adjacent culture spaces, and wherein a spheroid is cultured in each culture space;   wherein the spheroid cultured in a culture space is contained within that culture space and does not come into contact with a spheroid in an adjacent culture space during culturing,   wherein each culture space has an equivalent diameter that is 1 to 5 times the diameter of a desired spheroid,   wherein each culture space has a height that is 0.3 to 5 times the equivalent diameter, and   wherein a surface of each culture space has a water contact angle of 45 degrees or less.   
     
     
         2 . The method of  claim 1 , wherein each culture space has a cylindrical shape or rectangular prism shape. 
     
     
         3 . The method of  claim 2 , wherein each culture space has a hemispherical bottom. 
     
     
         4 . The method of  claim 2 , wherein each culture space has an inverted cone-shaped bottom. 
     
     
         5 . The method of  claim 1 , wherein each culture space has an equivalent diameter in a range of 100 μm to 5000 μm. 
     
     
         6 . The method of  claim 1 , wherein a height of the wall is equal to a height of the culture space. 
     
     
         7 . The method of  claim 1 , wherein a thickness of the wall is in a range of 5 μm to 50 μm. 
     
     
         8 . The method of  claim 1 , wherein an angle formed between an upper surface and a side surface of the wall partitioning the adjacent culture spaces is in a range of 90 degrees to 135 degrees. 
     
     
         9 . The method of  claim 1 , wherein 60% or more of all cultured spheroids have a diameter within a range of plus-minus 5% of an average value of diameters of the spheroids obtained after the culturing. 
     
     
         10 . The method of  claim 1 , wherein the spheroids are cancer cells. 
     
     
         11 . The method of  claim 1 , wherein a thickness of a bottom portion of the culture chamber is 1 mm or less. 
     
     
         12 . The method of  claim 1 , wherein the culture chamber is formed of a resin molded product that is made of one selected from the group consisting of acrylic resin, polylactic acid, polyglycolic acid, styrene resin, acrylic-styrene copolymer resin, polycarbonate resin, polyester resin, polyvinyl alcohol resin, ethylene-vinylalcohol copolymer resin, thermoplastic elastomer, vinyl chloride resin, and silicon resin, or a combination thereof. 
     
     
         13 . The method of  claim 1 , wherein the surface of each culture space has been processed by glass processing so that the surface has a water contact angle of 45 degrees or less. 
     
     
         14 . The method of  claim 1 , wherein the surface of each culture space has been processed by forming a functional group thereon by plasma treatment so that the surface has a water contact angle of 45 degrees or less. 
     
     
         15 . The method of  claim 1 , wherein a polymer having hydrophilic and hydrophobic properties that vary depending on temperature or light has been coated on the surface of each culture space. 
     
     
         16 . The method of  claim 1 , wherein a hydrophilic polymer chain that inhibits cell adhesion has been immobilized on the surface of each culture space. 
     
     
         17 . The method of  claim 1 , wherein a phospholipid or a phospholipid-polymer complex has been immobilized on the surface of each culture space. 
     
     
         18 . The method of  claim 1 , wherein the surface of each culture space has been processed by glass processing so that the surface has a water contact angle of 45 degrees or less, and thereafter a polymer having hydrophilic and hydrophobic properties that vary depending on temperature or light, a hydrophilic polymer chain that inhibits cell adhesion, or one of a phospholipid and a phospholipid-polymer complex is immobilized on the surface of the culture chamber. 
     
     
         19 . The method of  claim 1 , wherein the surface of each culture space has been processed by forming a functional group thereon by plasma treatment so that the surface has a water contact angle of 45 degrees or less, and thereafter a polymer having hydrophilic and hydrophobic properties that vary depending on temperature or light, a hydrophilic polymer chain that inhibits cell adhesion, or one of a phospholipid and a phospholipid-polymer complex is immobilized on the surface of the culture chamber. 
     
     
         20 . A culture chamber for culturing adherent cells to form spheroids according to the method of  claim 1 .