Microfluidic devices and methods

Abstract

Contemplated microfluidic devices and methods are drawn to protein arrays in which distinct and detergent-containing antigen preparations are deposited onto an optical contrast layer in a non-specific and non-covalent manner. Detection of binding a is carried out using a dye that precipitates or agglomerates to so form a visually detectable signal at a dynamic range of at least three orders of magnitude.

1 . A method of performing an analytic test, comprising:
 providing a carrier having a surface comprising an optical contrast layer;   wherein a plurality of detergent-containing non-purified distinct antigen preparations are non-covalently and non-specifically coupled to the optical contrast layer at respective predetermined locations to form an antigen array having a density of at least 10 distinct antigen preparations per cm 2 ;   wherein the optical contrast layer has a thickness and composition sufficient to prevent confluence of the antigen preparations when the distinct antigen preparations are deposited onto the optical contrast layer;   contacting the antigen array with a solution comprising an antibody under conditions to allow binding of the antibody to an antigen of at least one of the antigen preparations; and   detecting binding of the antibody using a visually detectable, and precipitating or agglomerating dye.   
     
     
         2 . The method of  claim 1  wherein the contrast layer and the antigen preparations have a composition such that the step of detecting binding of the antibody allows detection over a dynamic range of at least three orders of magnitude. 
     
     
         3 . The method of  claim 1  wherein the array comprises at least two distinct antigens from the same pathogen. 
     
     
         4 . The method of  claim 3  wherein the at least two distinct antigens have known quantified and known relative reactivities with respect to sera of a population infected with the pathogen. 
     
     
         5 . The method of  claim 1  wherein the carrier is enclosed in a chamber of a housing, wherein the chamber has a volume of 1 μl and 50 μl. 
     
     
         6 . The method of  claim 5  wherein the chamber has a plurality of cavities sized and positioned to allow resonance mixing of a fluid that is in contact with the array. 
     
     
         7 . The method of  claim 6  further comprising a step of mixing the fluid at between 3 and 4 kHz. 
     
     
         8 . The method of  claim 1  wherein the dye is selected from the group consisting of 3-amino-9-ethylcarbazole, 5-bromo-4-chloro-3-indolylphosphate, 3-3′-diaminobenzidine tetrachloride, 3,3′,5,5′-tetramethylbenzidine, and a colloidal metal. 
     
     
         9 . The method of  claim 1  wherein the step of detecting is performed using a scanner or CCD detector. 
     
     
         10 . The method of  claim 1  wherein the step of detecting is performed from opposite sides of the optical contrast layer. 
     
     
         11 . A microfluidic device, comprising:
 an enclosed reaction volume formed at least in part by a carrier material to which an optical contrast layer is coupled, wherein the optical contrast layer is disposed within the reaction volume and opposite to a cavity layer within the reaction volume;   wherein a plurality of distinct antigens is non-covalently and non-specifically coupled to the optical contrast layer in predetermined positions;   wherein the cavity layer has plurality of cavities that are sized and dimensioned to allow trapping of air in the plurality of cavities;   wherein at least one of the number and size of the cavities is selected such that hybridization of an antibody to at least one of the plurality of antigens is substantially complete within less than 60 minutes upon mixing; and   wherein the carrier material and optical contrast layer are configured to allow quantitative detection of a visually detectable, and precipitating or agglomerating dye.   
     
     
         12 . The microfluidic device of  claim 11  wherein the optical contrast layer comprises nitrocellulose. 
     
     
         13 . The microfluidic device of  claim 11  wherein the carrier material comprises at least one of glass and a transparent synthetic polymer. 
     
     
         14 . The microfluidic device of  claim 11  wherein the cavities in the cavity layer are circular cavities arranged at regular intervals in x- and y-coordinate. 
     
     
         15 . The microfluidic device of  claim 11  wherein a ratio between a number of the antigens to a number of cavities is at least 3:1, and wherein a ratio between an area of an antigen and a cavity diameter is at least 1:3. 
     
     
         16 . The microfluidic device of  claim 11  wherein the reaction volume is between 1 μl and 50 μl. 
     
     
         17 . The microfluidic device of  claim 11  wherein the carrier material and the optical contrast layer are configured to allow quantitative detection over a dynamic range of at least three orders of magnitude. 
     
     
         18 . The microfluidic device of  claim 11  wherein the plurality of distinct antigens are non-purified and further comprise a detergent. 
     
     
         19 . The microfluidic device of  claim 11  wherein the dye is selected from the group consisting of 3-amino-9-ethylcarbazole, 5-bromo-4-chloro-3-indolylphosphate, 3-3′-diaminobenzidine tetrachloride, 3,3′,5,5′-tetramethylbenzidine, and a colloidal metal. 
     
     
         20 . The microfluidic device of  claim 11  wherein the carrier material and the optical contrast layer are configured to allow optical detection from opposite sides of the optical contrast layer.