Microwell array articles and methods of use

Abstract

The disclosure provides microstructured articles and methods useful for detecting an analyte in a sample. The articles include microwell arrays. The articles can be used with an optical system component in methods to detect or characterize an analyte

1 . An article, comprising
 a microstructured layer with upper and lower major surfaces, comprising a plurality of optically-isolated microwells extending between the upper and lower major surfaces; and   an optically-transmissive flexible layer coupled to the lower major surface of the microstructured layer;   wherein each microwell in the microstructured layer comprises a top opening, a bottom opening, and at least one side wall extending between the top opening and the bottom opening;   wherein the optically-transmissive flexible layer has an average thickness of about 2 μm to about 50 μm.   
     
     
         2 . An article, comprising
 a microstructured layer with upper and lower major surfaces, comprising a plurality of optically-isolated microwells extending below the upper major surface; and   an optically-transmissive flexible layer coupled to the lower major surface of the microstructured layer;   wherein each microwell in the microstructured layer comprises an opening, an optically-transmissive bottom wall, and at least one side wall extending between the opening and the bottom wall;   wherein the bottom wall has a thickness of about 0.1 μm to about 5 μm.   
     
     
         3 . An article, comprising
 a microstructured layer with upper and lower major surfaces, comprising a plurality of optically-isolated microwells extending below the upper major surface; and   an optically-transmissive flexible layer coupled to the lower major surface of the microstructured layer;   wherein each microwell in the microstructured layer comprises an opening, an optically-transmissive bottom wall, and at least one side wall extending between the opening and the bottom wall;   wherein a thickness (t) is defined by a thickness of the bottom wall plus a thickness of the optically-transmissive flexible layer;   wherein t is about 2 μm to about 55 μm.   
     
     
         4 . The article of  claim 1 , wherein the microstructured layer comprises a colorant. 
     
     
         5 . The article of  claim 4 , wherein the colorant is selected from the group consisting of carbon black, fuchsin, carbazole violet, and Foron Brilliant Blue. 
     
     
         6 . The article of  claim 1 , wherein the optically-transmissive flexible layer is transmissive to a selected wavelength of light. 
     
     
         7 . The article of  claim 2 , wherein the bottom wall is substantially transmissive to the selected wavelength of light. 
     
     
         8 . The article of  claim 6 , wherein the at least one sidewall is substantially nontransmissive to the selected wavelength of light. 
     
     
         9 . The article of  claim 8 , wherein the at least one sidewall is at least 50% less transmissive of a selected wavelength of light than the bottom wall. 
     
     
         10 . The article of  claim 8 , wherein a sidewall is at least 90% less transmissive of a selected wavelength of light than the bottom wall. 
     
     
         11 . The article of  claim 1 , wherein the bottom wall and/or the at least one sidewall of a microwell further comprise a coating. 
     
     
         12 . The article of  claim 11 , wherein the bottom wall and/or the at least one side wall of a microwell further comprises a plurality of coatings. 
     
     
         13 . The article of  claim 11 , wherein at least one coating comprises SiO 2 . 
     
     
         14 . The article of  claim 11 , wherein at least one coating comprises a reflective coating. 
     
     
         15 . The article of  claim 1 , wherein the microstructured layer is a cured polymer derived from a resin. 
     
     
         16 . The article of  claim 15 , wherein the resin is selected from the group consisting of acrylic-based resins derived from epoxies, polyesters, polyethers, and urethanes; ethylenically unsaturated compounds; aminoplast derivatives having at least one pendant acrylate group; polyurethanes (polyureas) derived from an isocyanate and a polyol (or polyamine); isocyanate derivatives having at least one pendant acrylate group; epoxy resins other than acrylated epoxies; and mixtures and combinations thereof. 
     
     
         17 . The article of  claim 16 , wherein the optically transmissive layer comprises polyethylene terephthalate, polyethylene naphthalate, high density polyethylene, low density polyethylene, or linear low density polyethylene. 
     
     
         18 . The article of  claim 16 , wherein the film comprises a multi-layer film. 
     
     
         19 . The article of  claim 1 , wherein the optically-transmissive flexible layer further comprises an adhesive. 
     
     
         20 . The article of  claim 1 , wherein the microstructured layer further comprises a tab region that is substantially free of microwells. 
     
     
         21 . The article of  claim 20 , wherein the tab region comprises a detachable portion. 
     
     
         22 . The article of  claim 1 , further comprising a cover layer coupled to the upper major surface of the microstructured layer. 
     
     
         23 . The article of  claim 22 , wherein the cover layer is removably coupled to the microstructured layer. 
     
     
         24 . The article of  claim 1 , further comprising a cover layer coupled to the optically-transmissive flexible layer on a surface opposite the microstructured layer. 
     
     
         25 . The article of  claim 24 , wherein the cover layer is removably coupled to the optically-transmissive flexible layer. 
     
     
         26 . The article of  claim 1 , further comprising an optical detection system comprising an optical device, wherein the optical device is optically coupled to a substrate. 
     
     
         27 . The article of  claim 26 , wherein the optical device comprises a fiber optic face plate. 
     
     
         28 . The article of  claim 26 , wherein the optical device comprises a CCD image sensor, a CMOS image sensor, or a photomultiplier tube. 
     
     
         29 . The article of  claim 26 , wherein the optical system further comprises a processor. 
     
     
         30 . The article of  claim 1 , wherein at least one microwell further comprises a polynucleotide. 
     
     
         31 . The article of  claim 30 , wherein the polynucleotide is coupled to a particle. 
     
     
         32 . A process of manufacturing a microwell array article, comprising the steps of:
 providing
 a tool having a molding surface with a plurality of projections extending therefrom suitable for forming the microstructure elements; 
 a flowable, curable resin composition; 
 an optically-transmissive flexible layer having first and second major surfaces; 
   applying to the molding surface a volume of the flowable curable resin composition suitable for forming desired microstructure elements;   contacting the resin composition with the first major surface of the optically-transmissive flexible layer;
 wherein the first major surface of the optically-transmissive flexible layer is surface-treated to promote adhesion to a cured resin composition; 
 wherein the thickness of the optically-transmissive flexible layer is about 50 μm or less; and 
   curing the resin composition while in contact with the flexible layer to form a microwell array article comprising a cured microstructured layer including a plurality of microwells bonded to the optically-transmissive flexible layer, and   removing the microwell array article from the tool.   
     
     
         33 . The process of  claim 32 , wherein contacting the resin composition with the first major surface of the optically-transmissive flexible layer comprises applying pressure to the resin composition to substantially displace the resin between the tops of the projections in the tool and the surface of the optically-transmissive layer. 
     
     
         34 . The process of  claim 32 , wherein the optically-transmissive flexible layer is surface-treated with a process selected from the group consisting of radiation treatment, corona discharge treatment, flame treatment, plasma treatment, high energy UV treatment, and chemical priming treatment. 
     
     
         35 . The process of  claim 32 , wherein the optically-transmissive flexible layer is coupled to a carrier. 
     
     
         36 . The process of  claim 32 , wherein the resin composition comprises a colorant. 
     
     
         37 . The process of  claim 34 , wherein said curing comprises exposing the resin composition to at least one curing treatment selected from the group consisting of actinic radiation from a radiation source, an electron beam, and a chemical curing agent. 
     
     
         38 . The process of  claim 32 , wherein the flexible layer has a thickness of about 2 μm to about 48 μm. 
     
     
         39 . The process of  claim 32 , further comprising the step of removing a portion of the microstructured layer. 
     
     
         40 . The process of  claim 32 , further comprising the step of removing a portion of the optically-transmissive flexible layer. 
     
     
         41 . The process of  claim 32 , further comprising the step of disposing a reagent in a microwell. 
     
     
         42 . A method of detecting an analyte in a microwell array, comprising:
 providing
 a sample suspected of containing an analyte; 
 a reagent for the optical detection of the analyte; 
 an optical detection system; and 
 an article according to  claim 1 ; 
   contacting the sample and the reagent in at least one microwells under conditions suitable to detect the analyte, if present, in the at least one microwells; and   using the optical detection system to detect the presence or absence of the analyte in a microwell.   
     
     
         43 . The method of  claim 42 , wherein the optical system is optically coupled to the substrate. 
     
     
         44 . The method of  claim 42 , wherein the optical system comprises a fiber optic face plate and wherein using the optical detection system comprises passing a signal through the fiber optic face plate. 
     
     
         45 . The method of  claim 42 , wherein the optical system comprises a CCD image sensor, a CMOS image sensor, or a photomultiplier tube. 
     
     
         46 . The method of  claim 42 , wherein the optical system further comprises a processor. 
     
     
         47 . The method of  claim 42 , wherein detecting the presence or absence of an analyte comprises detecting light that is indicative of the presence of the analyte. 
     
     
         48 . The method of  claim 47 , wherein detecting light comprises detecting light by absorbance, reflectance, or fluorescence. 
     
     
         49 . The method of  claim 48 , wherein detecting light comprises detecting light from a lumigenic reaction. 
     
     
         50 . The method of  claim 42 , wherein detecting the presence or absence of the analyte comprises obtaining an image of a microwell. 
     
     
         51 . The method of  claim 50 , wherein detecting the presence or absence of the analyte comprises displaying, analyzing, or printing the image of a microwell. 
     
     
         52 . The method of  claim 42 , wherein contacting the sample and the reagent in a plurality of microwells under conditions suitable to detect the analyte comprises an enzyme and an enzyme substrate. 
     
     
         53 . The method of  claim 42 , wherein contacting the sample and the reagent in a plurality of microwells under conditions suitable to detect the analyte comprises forming a hybrid between two polynucleotides. 
     
     
         54 . An assay system, comprising:
 an article according to  claim 1 ;   an imaging device optically coupled thereto; and   a processor.   
     
     
         55 . The assay system of  claim 54 , wherein optically coupled comprises adhesively coupling the microwell array article to a solid interface. 
     
     
         56 . A composition, comprising a compound selected from the group consisting of 1-(3-methyl-n-butylamino)-9,10-anthracenedione; 1-(3-methyl-2-butylamino)-9,10-anthracenedione; 1-(2-heptylamino)-9,10-anthracenedione; 1,1,3,3-tetramethylbutyl-9,10-anthracenedione; 1,10-decamethylene-bis-(-1-amino-9,10-anthracenedione); 1,1-dimethylethylamino-9,10-anthracenedione; and 1-(n-butoxypropylamino)-9,10-anthracenedione. 
     
     
         57 . The composition of  claim 56 , further comprising a cured polymer. 
     
     
         58 . The composition of  claim 57 , where the cured polymer is derived from a resin selected from the group consisting of acrylate resins, acrylic resins, acrylic-based resins derived from epoxies, polyesters, polyethers, and urethanes; ethylenically unsaturated compounds; aminoplast derivatives having at least one pendant acrylate group; polyurethanes (polyureas) derived from an isocyanate and a polyol (or polyamine); isocyanate derivatives having at least one pendant acrylate group; epoxy resins other than acrylated epoxies; and mixtures and combinations thereof.