Microwell Array Chip and Its Manufacturing Method

Abstract

A microwell array chip having multiple microwells on a principal surface of a substrate, said microwells being of a shape and size permitting the storage of only a single organic cell in each microwell, wherein microwell markers are present on the same substrate surface as the openings of the microwells. A microwell array chip having multiple microwells on a principal surface of a substrate, said microwells being of a shape and size permitting the storage of only a single organic cell in each microwell, wherein protrusions are present in the openings of said microwells so as to narrow said openings. A method for manufacturing the microwell array chip. The method comprises the steps of: forming a film on at least one principal surface of a substrate; applying a resist coating on the film that has been formed; exposing the resist surface through a mask having a microwell pattern and removing uncured portions of resist; etching the exposed portions of said film and substrate to fabricate wells in the form of a microwell array; and removing the resist. A microwell array chip made of silicon and having multiple microwells, each microwell being used to store a single specimen organic cell, wherein each microwell is of a size and shape holding just one organic cell.

1 . A microwell array chip having multiple microwells on a principal surface of a substrate, said microwells being of a shape and size permitting the storage of only a single organic cell in each microwell,
 wherein microwell markers are present on the same substrate surface as the openings of the microwells.   
     
     
         2 . The microwell array chip according to  claim 1 , wherein said multiple microwells are arranged horizontally and vertically at identical spacing and markers are provided at a prescribed number of microwells. 
     
     
         3 . The microwell array chip according to  claim 1 , wherein said multiple microwells are positioned on a principal surface of the substrate by being divided into groups each comprising a prescribed number of microwells and markers are provided in a manner permitting determination of the position of each group. 
     
     
         4 . The microwell array chip according to  claim 3 , wherein the number of microwells in each group falls within a range of from 10 to 10,000. 
     
     
         5 . The microwell array chip according to  claim 1 , wherein said marker is comprised of a fluorescent material or a reflective material. 
     
     
         6 . The microwell array chip according to  claim 1 , wherein said marker is a positioning marker. 
     
     
         7 . The microwell array chip according to  claim 1 , wherein said substrate is made of silicon, metal, or resin. 
     
     
         8 . The microwell array chip according to  claim 1 , wherein the shape of said microwells is cylindrical, polyhedral comprised of multiple surfaces, inversely conical, inversely pyramidal, or a combination of two or more of the above. 
     
     
         9 . The microwell array chip according to  claim 1 , wherein the diameter of the largest circle that can be inscribed within the planar shape of the microwells falls within a range of from 0.5 to 2-fold the diameter of the organic cell that is to be contained in the microwells, and the depth of the microwells falls within a range of from 0.5 to 4-fold the diameter of the organic cell that is to be contained in the microwells. 
     
     
         10 . The microwell array chip according to  claim 1 , wherein the organic cell is a lymphocyte and the microwell array chip is employed to detect single antigen-specific lymphocytes. 
     
     
         11 . The microwell array chip according to  claim 1 , wherein a hydrophobic region is provided in a manner surrounding said multiple microwells on said principal surface. 
     
     
         12 . The microwell array chip according to  claim 11 , wherein said hydrophobic region is comprised of a silicon surface or fluorine-containing surface. 
     
     
         13 . A microwell array chip having multiple microwells on a principal surface of a substrate, said microwells being of a shape and size permitting the storage of only a single organic cell in each microwell,
 wherein protrusions are present in the openings of said microwells so as to narrow said openings.   
     
     
         14 . The microwell array chip according to  claim 13 , wherein said protrusions are formed by providing a film on the substrate surface that protrudes into the openings. 
     
     
         15 . The microwell array chip according to  claim 13 , wherein the size of the openings formed by said protrusions permits the passage of the organic cell to be stored in the microwell. 
     
     
         16 . The microwell array chip according to  claim 13 , wherein said substrate is made of silicon, metal, or resin. 
     
     
         17 . The microwell array chip according to  claim 14 , wherein the film provided on said substrate surface is an oxide film, nitride film, impurity diffusion film, metal film, or resin film. 
     
     
         18 . A method for manufacturing the microwell array chip according to  claim 13  comprising the steps of:
 forming a film on at least one principal surface of a substrate;   applying a resist coating on the film that has been formed;   exposing the resist surface through a mask having a microwell pattern and removing uncured portions of resist;   etching the exposed portions of said film and substrate to fabricate wells in the form of a microwell array; and   removing the resist.   
     
     
         19 . The method for manufacturing a microwell array chip according to  claim 18 , wherein said substrate is made of silicon, metal, or resin. 
     
     
         20 . The method for manufacturing a microwell array chip according to  claim 18 , wherein said film provided on said substrate surface is an oxide film, nitride film, impurity diffusion film, metal film, or resin film. 
     
     
         21 . The microwell array chip according to  claim 13 , wherein a hydrophobic region is provided in a manner surrounding said multiple microwells on said principal surface. 
     
     
         22 . The microwell array chip according to  claim 21 , wherein said hydrophobic region has a silicon surface or fluorine-containing surface. 
     
     
         23 . A microwell array chip made of silicon and having multiple microwells, each microwell being used to store a single specimen organic cell,
 wherein each microwell is of a size and shape holding just one organic cell.   
     
     
         24 . The microwell array chip according to  claim 23 , wherein each of said microwells is cylindrical, polyhedral comprised of multiple surfaces, inversely conical, inversely pyramidal, or a combination of two or more of the above. 
     
     
         25 . The microwell array chip according to  claim 23 , wherein the diameter of the largest circle that can be inscribed within the planar shape of the microwells falls within a range of from 0.5 to 2-fold the diameter of the organic cell that is to be contained in the microwells, and the depth of the microwells falls within a range of from 0.5 to 4-fold the diameter of the organic cell that is to be contained in the microwells. 
     
     
         26 . The microwell array chip according to  claim 23 , wherein said organic cell is a lymphocyte and the microwell array chip is used to detect single antigen-specific lymphocytes. 
     
     
         27 . The microwell array chip according to  claim 23 , wherein the interior surface of said microwells is coated with a fluorocarbon film or a silicon oxide film. 
     
     
         28 . The microwell array chip according to  claim 27 , employed so that a single organic cell stored in a single microwell is recovered from the microwell. 
     
     
         29 . The microwell array chip according to  claim 23 , wherein a hydrophobic region is provided in a manner surrounding said multiple microwells on said principal surface. 
     
     
         30 . The microwell array chip according to  claim 29 , wherein said hydrophobic region has a silicon surface or fluorine-containing surface. 
     
     
         31 . A microwell array chip having a microwell on a principal surface of a substrate, wherein a hydrophobic surface region is provided in a manner surrounding said microwell on said principal surface. 
     
     
         32 . The microwell array chip according to  claim 31 , wherein said hydrophobic region has a silicon surface or fluorine surface.