A poly(N-isopropylacrylamide) (PNIPAAm) is known as a thermoresponsive polymer. An aqueous solution of PNIPAAm causes a phase separation due to a change in the temperature and at 31° C. or lower, it is dissolved in water and at a higher temperature, it is insolubilized and is separated out. N-isopropylacrylamide (NIPAAm) is easily polymerized with a radical initiator to obtain a PNIPAAm. Further, NIPAAm is known to be copolymerized with other functional monomers and the thus obtained polymer responds to various stimulation such as not only a change in the temperature, but also a light, an electric field, a change in pH and a solvent exchange.
A thermoresponsive polymer is known also to be used as a material for immobilizing biological materials (Japanese Patent Application Publication No. JP-A-2003-102466 (Patent Document 1) and Japanese Patent Application Publication No. JP-A-Hei 9-23876 (Patent Document 2)).
On the other hand, it has been attempted that an individual cell is specified, discriminated and a discriminated individual cell is used. For example, a study is made on that an antigen specificity of an individual lymphocyte is individually detected; a detected individual antigen-specific lymphocyte is collected; and using the collected individual antigen-specific lymphocyte, for example an antibody is produced (Tamiya et. al., “BIO INDUSTRY” Vol. 20, No. 7 (2003), pp. 60-67 (Non-Patent Document 1), Japanese Patent Application Publication No. JP-A-2004-173681 (Patent Document 3)).
However, an usual coated film of PNIPAAm is extremely easily dissolved in water or a polar organic solvent. Accordingly, when using a coated film of PNIPAAm, a biological material such as a cell is attempted to be immobilized, a part thereof which has been contacted with water is dissolved out on and on. Further, when for fine-processing a coated film of PNIPAAm, a resist is overcoated on an upper layer of the coated film of PNIPAAm, PNIPAAm is dissolved into a solvent of the resist, so that the both layers are mixed.
Thus, the present inventors not only have provided a material (NIPAAm) which is insoluble in water, an aqueous solution, and an organic solvent and has thermoresponsivity, but also have developed a chip utilizing a change in an adhering force to a cell which is caused by such a nature of a thermoresponsive material that the property of the material is changed from hydrophilic to hydrophobic corresponding to the change in the temperature, and have applied the patent (WO 2005/095510).
However, with respect to a chip utilizing a temperature-responsibility by which a material is changed from hydrophilic to hydrophobic (as a result, the adhering force to the cell is changed), there has been found such a drawback that when in a washing process, the cell is washed too strongly, all lymphocytes are washed away, on the contrary, when the cell is washed too weakly, lymphocytes are remained at a position where lymphocytes need not to be remained.
Thus, the object of the present invention is to provide a novel chip useful for treating cells and the like which has a mechanism and a structure wherein the size of a hole pattern is arbitrarily changed so that cells can easily move in and get out from the hole during scattering or collecting cells but can hardly get out from the hole during washing or antigen-stimulation.
USPTO Backgrounds
USPTO Backgrounds