Very large scale immobilized polymer synthesis

Abstract

A synthetic strategy for the creation of large scale chemical diversity. Solid-phase chemistry, photolabile protecting groups, and photolithography are used to achieve light-directed spatially-addressable parallel chemical synthesis. Binary masking techniques are utilized in one embodiment. A reactor system, photoremovable protecting groups, and improved data collection and handling techniques are also disclosed. A technique for screening linker molecules is also provided.

1 . A method of detecting hybridization between biological polymers, comprising the acts of: 
 providing a substrate having a surface including at least one biological polymer and at least one fluorescent label associated with the biological polymer;    generating an excitation laser beam;    scanning said laser beam relative to said surface; collecting fluorescent radiation responsive to said laser beam using optics;    detecting said collected fluorescent radiation; and    autofocusing to bring into focus with respect to said optics at least a portion of said surface including the biological polymer.    
     
     
         2 - 62 . (canceled)  
     
     
         63 . A method of interrogating an addressable array unit having a transparent substrate with a back surface, and an array with a plurality of different chemical features on a front surface, the method comprising: (a) illuminating the features while the array is dry, with an interrogating light which is directed through the substrate from the back surface and onto the chemical features on the front surface; and (b) detecting light emitted from respective features in response to the interrogating light, which detected light has passed from the front surface, through the substrate and out the back surface; wherein the light is emitted from locations of the features which are spaced from the front surface a distance of less than one-eighth of the wavelength of the illuminating light in a gas or a vacuum which is in contact with the dry array.  
     
     
         64 . A method according to  claim 63  wherein the light emitting locations of the chemical features are spaced from the front surface a distance of less than one-tenth of the wavelength of the illuminating light.  
     
     
         65 . A method according to  claim 63  wherein the light emitting locations of the chemical features are spaced from the front surface a distance of less than one-twentieth of the wavelength of the illuminating light.  
     
     
         66 . A method according to  claim 63  wherein the light emitting locations of the chemical features are spaced from the front surface a distance of less than one-fiftieth of the wavelength of the illuminating light.  
     
     
         67 . A method according to  claim 63  wherein the interrogating light is directed toward the back surface at an angle of between 0 and 45 degrees to a normal to the back surface.  
     
     
         68 . A method according to  claim 67  wherein the angle is less than 25 degrees.  
     
     
         69 . A method according to  claim 67  wherein the angle is less than 10 degrees.  
     
     
         70 . A method according to  claim 63  wherein the chemical features are polynucleotides.  
     
     
         71 . A method according to  claim 63  wherein the chemical features are amino acid polymers.  
     
     
         72 . A method of interrogating an addressable array unit having a transparent substrate with a back surface, and an array with a plurality of different chemical features on a front surface, the method comprising: (a) illuminating the features while the array is dry, with an interrogating light which is directed through the substrate from the back surface and onto the chemical features on the front surface; and (b) detecting light emitted from respective features in response to the interrogating light, which detected light has passed from the front surface, through the substrate and out the back surface; wherein the light is emitted from locations of the features which are spaced from the front surface a distance of less than one-eighth of the wavelength of the emitted light in a gas or a vacuum which is in contact with the dry array.  
     
     
         73 . A method according to  claim 72  wherein the light is emitted from locations of the features which are spaced from the front surface a distance of less than one-tenth of the emitted light wavelength.  
     
     
         74 . A method according to  claim 72  wherein the light is emitted from locations of the features which are spaced from the front surface a distance of less than one-fiftieth of the emitted light wavelength.  
     
     
         75 . A method of interrogating an addressable array unit having a substrate with a back surface, and an array with a plurality of different chemical features on a front surface, the method comprising: (a) illuminating the features while the array is dry, with an interrogating light which is directed through the substrate from the back surface and onto the chemical features on the front surface; and (b) detecting light emitted from respective features in response to the interrogating light, which detected light has passed from the front surface, through the substrate and out the back surface; wherein the light is emitted from locations of the features which are spaced from the front surface a distance such that the average detected signal from the dry array is at least 10% greater than would be detected under the same conditions except with the interrogating light and detected emitted light not passing through the substrate.  
     
     
         76 . A method according to  claim 75  wherein the average detected signal from the array is at least 40% greater.  
     
     
         77 . A method according to  claim 75  wherein the average detected signal from the array is at least 80% greater.  
     
     
         78 . A package comprising: an addressable array unit having a transparent substrate with a back surface, and an array with a plurality of different chemical features on a front surface, the chemical features having a thickness of less than 100 nm; and instructions to: (i) interrogate the array with an interrogating light which is directed through the substrate from the back surface and onto the chemical features on the front surface; and (ii) detect light emitted from respective features in response to the interrogating light, which detected light has passed from the front surface, through the substrate and out the back surface.  
     
     
         79 . A package according to  claim 78  wherein the features have a thickness of less than 50 nm.  
     
     
         80 . A package according to  claim 78  wherein the features have a thickness of less than 10 nm.  
     
     
         81 . A method according to  claim 63  additionally comprising, prior to the illuminating and detecting: exposing the array to a sample in a liquid; and washing and drying the array.  
     
     
         82 . A method for use with an interrogating an addressable array unit having a transparent substrate with a back surface, and an array with a plurality of different chemical features on a front surface, the method comprising: (a) machine reading an identifier associated with the array unit; (b) based on the read identifier, retrieving by a processor an instruction that the array should be interrogated and read through the substrate from the back surface.  
     
     
         83 . A method according to  claim 82  wherein the identifier is on the array substrate or a housing carrying the array substrate.  
     
     
         84 . A method according to  claim 82  wherein the instruction is retrieved from the read identifier.  
     
     
         85 . A method according to  claim 82  wherein the instruction is retrieved from a memory using data from the read identifier.  
     
     
         86 . A method according to  claim 82  additionally comprising checking that the array is oriented within an array reader such that the array can be interrogated and read by the reader through the substrate from the back surface.