Cell culture substrate and its production method

Abstract

A cell culture substrate which is durable and which can be readily produced in commercial scale at a low cost, and its production method are provided. The cell culture substrate comprises a substrate and a layer formed by surface modification, which comprises a polymer containing amino group produced by reacting a polymer represented by the following formula (II): (wherein n is 0 or a positive integer, and m is a positive integer, the n and m representing degree of polymerization) formed by chemical vapor deposition of formyl[2.2]paracyclophane represented by the following formula (I): (wherein k is 0 or 1) with a polymer having at least one amino group (—NH 2 ) capable of forming Schiff base in its monomer. The production method of the cell culture substrate comprises the step of synthesizing such polymer on the substrate.

1 . A cell culture substrate comprising a substrate and a layer formed by surface modification, said layer comprising a polymer containing amino group produced by reacting a polymer represented by the following formula (II): 
     
       
         
         
             
             
         
       
     
     (wherein n is 0 or a positive integer, and m is a positive integer, the n and m representing degree of polymerization) formed by chemical vapor deposition of formyl[2.2]paracyclophane represented by the following formula (I): 
     
       
         
         
             
             
         
       
     
     (wherein k is 0 or 1) with a polymer having at least one amino group (—NH 2 ) capable of forming Schiff base in its monomer. 
   
   
       2 . The cell culture substrate according to  claim 1  wherein the polymer containing amino group constituting the layer formed by surface modification is represented by the following formula (III): 
     
       
         
         
             
             
         
       
     
     (wherein n is 0 or a positive integer, m is a positive integer, X represents a group having primary amine wherein amino group is bonded directly to the carbon atom in the polymer backbone or via an intervening alkylene group containing 1 to 4 carbon atoms to carbon or nitrogen in the polymer backbone). 
   
   
       3 . The cell culture substrate according to  claim 1  wherein the polymer having amino group constituting the surface modified layer is represented by the following formula (IV): 
     
       
         
         
             
             
         
       
     
     (wherein n is 0 or a positive integer, m is a positive integer, X represents a group having primary amine wherein amino group is bonded directly to the carbon atom in the polymer backbone or via an intervening alkylene group containing 1 to 4 carbon atoms to carbon or nitrogen in the polymer backbone). 
   
   
       4 . The cell culture substrate according to  claim 2  wherein the polymer backbone in X in the formula (III) or (IV) is polyethylene, polyethyleneimine, or polymethylene amide. 
   
   
       5 . The cell culture substrate according to  claim 2  wherein, in the formula (III) or (IV), X is bonded to —CH 2  via carbon or nitrogen. 
   
   
       6 . The cell culture substrate according to  claim 1  wherein the polymer represented by the formula (II) is formed by chemical vapor deposition of formyl[2.2]paracyclophane represented by the formula (I) on the underlying layer of chloro-substituted [2.2]paracyclophane. 
   
   
       7 . The cell culture substrate according to  claim 1  wherein the substrate is a three dimensional structure. 
   
   
       8 . A method for producing a cell culture substrate comprising a substrate and a layer formed by surface modification, comprising the steps of
 forming a layer of a polymer represented by formula (II):   
     
       
         
         
             
             
         
       
     
     (wherein n is 0 or a positive integer, and m is a positive integer, the n and m representing degree of polymerization)
 on the substrate by conducting chemical vapor deposition of formyl[2.2]paracyclophane represented by the following formula (I): 
 
     
       
         
         
             
             
         
       
     
     (wherein k is 0 or 1); and
 reacting the layer of the polymer represented by the formula (II) with a polymer having at least one amino group (—NH 2 ) capable of forming Schiff base in its monomer to obtain a layer of a polymer represented by the formula (III): 
 
     
       
         
         
             
             
         
       
     
     (wherein n is 0 or a positive integer, m is a positive integer, X represents a group having primary amine wherein amino group is bonded directly to the carbon atom in the polymer backbone or bonded via an intervening alkylene group containing 1 to 4 carbon atoms to carbon or nitrogen in the polymer backbone), this polymer layer constituting the-layer formed by surface modification. 
   
   
       9 . The method for producing a cell culture substrate according to  claim 8  wherein said reaction is conducted by immersing the polymer in a solution containing the polymer containing amino group (—NH 2 ). 
   
   
       10 . The method for producing a cell culture substrate according to  claim 9  wherein the solution containing the polymer containing amino group (—NH 2 ) has the polymer concentration of 0.1 to 0.00001%. 
   
   
       11 . The method for producing a cell culture substrate according to  claim 8  further comprising the step of
 reducing the polymer represented by the formula (III) to obtain the layer of a polymer represented by the formula (IV)   
     
       
         
         
             
             
         
       
     
     (wherein n is 0 or a positive integer, m is a positive integer, X represents a group having primary amine wherein amino group is bonded directly to the carbon atom in the polymer backbone or bonded via an intervening alkylene group containing 1 to 4 carbon atoms to carbon or nitrogen in the polymer backbone), this polymer layer constituting the layer formed by surface modification.