Lateral flow assays using dna dendrimers

Abstract

The present invention relates to lateral flow devices and kits using DNA dendrimers, and to methods for detecting an analyte using the lateral flow devices and kits. More specifically, the DNA dendrimers may be dried on a location on the test device upstream from said test zone, before said test device is used.

1 . A test device for detecting an analyte in a liquid sample, which device comprises a porous matrix that comprises a test zone on said porous matrix, said test zone comprising a test reagent that binds to an analyte or another binding reagent that binds to said analyte, or is an analyte or an analyte analog that competes with an analyte in said sample for binding to a binding reagent for said analyte,
 wherein a liquid sample flows laterally along said test device and passes said test zone to form a detectable signal to indicate presence, absence and/or amount of said analyte in said liquid sample, the formation of said detectable signal requires the use of a detectable label and a DNA dendrimer, said DNA dendrimer comprises a first component that links said DNA dendrimer to a binding reagent linkable to said DNA dendrimer, an analyte linkable to said DNA dendrimer, or an analyte analog linkable to said DNA dendrimer, and a second component that links said DNA dendrimer to said detectable label linkable to said DNA dendrimer, and wherein:   a) said analyte is not a polynucleotide; or   b) said DNA dendrimer comprises from about 1 to about 324, preferably from about 60 to about 324, from about 80 to about 300 or from about 100 to about 200, said first components; or   c) said DNA dendrimer, before said test device is used, is dried on a location on said test device upstream from said test zone.   
     
     
         2 - 7 . (canceled) 
     
     
         8 . A method for detecting an analyte in a liquid sample, which method comprises:
 a) contacting a liquid sample with the test device of  claim 1 , wherein the liquid sample is applied to a site of the test device upstream of the test zone;   b) transporting an analyte, if present in the liquid sample, a detectable label and a DNA dendrimer to the test zone; and   c) assessing the presence, absence, and/or amount of a signal generated by the detectable label at the test zone to determining the presence, absence and/or amount of the analyte in the liquid sample.   
     
     
         9 - 12 . (canceled) 
     
     
         13 . A test device for detecting an analyte in a liquid sample, which device comprises a porous matrix that comprises a test zone on said porous matrix, said test zone comprising a test reagent that binds to an analyte or another binding reagent that binds to said analyte, or is an analyte or an analyte analog that competes with an analyte in said sample for binding to a binding reagent for said analyte,
 wherein a liquid sample flows laterally along said test device and passes said test zone to form a detectable signal to indicate presence, absence and/or amount of said analyte in said liquid sample, the formation of said detectable signal requires the use of a DNA dendrimer linked to a detectable label non-covalently, and said DNA dendrimer comprises a component that links said DNA dendrimer to a binding reagent linkable to said DNA dendrimer, an analyte linkable to said DNA dendrimer, or an analyte analog linkable to said DNA dendrimer, and wherein:   a) said analyte is not a polynucleotide; or   b) said DNA dendrimer comprises from about 1 to about 324, preferably from about 60 to about 324, from about 80 to about 300 or from about 100 to about 200, said first components; or   c) said DNA dendrimer, before said test device is used, is dried on a location on said test device upstream from said test zone.   
     
     
         14 - 21 . (canceled) 
     
     
         22 . A method for detecting an analyte in a liquid sample, which method comprises:
 a) contacting a liquid sample with the test device of  claim 13 , wherein the liquid sample is applied to a site of the test device upstream of the test zone;   b) transporting an analyte, if present in the liquid sample, a detectable label and a DNA dendrimer to the test zone; and   c) assessing the presence, absence, and/or amount of a signal generated by the detectable label at the test zone to determining the presence, absence and/or amount of the analyte in the liquid sample.   
     
     
         23 - 25 . (canceled) 
     
     
         26 . The test device of  claim 1 , wherein the DNA dendrimer, before the test device is used, is dried on a location on the test device upstream from the test zone, and at least about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% of the dried DNA dendrimer is capable of being moved to the test zone by a liquid. 
     
     
         27 - 32 . (canceled) 
     
     
         33 . The test device of  claim 26 , wherein the liquid has a volume of at least about 20 ul, 25 ul, 30 ul, 35 ul, 40 ul, 45 ul, 50 ul, 55 ul, 60 ul, 65 ul, 70 ul, 75 ul, 80 ul, 85 ul, 90 ul, 95 ul, 100 ul, 110 ul, 120 ul, 150 ul, 200 ul, 300 ul, 400 ul, 500 ul, or a volume larger than 500 ul. 
     
     
         34 . The test device of  claim 1 , wherein the DNA dendrimer comprises:
 a) an one-layer DNA dendrimer that comprises from about 1-2, 2-4 or 4-8 of the first and/or second component(s);   b) a two-layer DNA dendrimer that comprises from about 1-4, 4-8 or 8-18 of the first and/or second component(s);   c) a three-layer DNA dendrimer that comprises from about 2-6, 6-10, 10-18, 18-28, 28-38, 38-50 or more than 50 of the first and/or second components; and/or   d) a four-layer DNA dendrimer that comprises from about 15-25, 25-45, 45-60, 60-80, 85-105, 105-130 or more than 130 of the first and/or second components.   
     
     
         35 . The test device of  claim 34 , wherein at least one of the first component(s) and the second component(s) is a polynucleotide and the other is a non-polynucleotide moiety. 
     
     
         36 . The test device of  claim 34 , wherein the DNA dendrimer comprises:
 a) an one-layer DNA dendrimer that comprises from about 1-2, 2-4 or 4-8 of the first component(s);   b) a two-layer DNA dendrimer that comprises from about 1-4, 4-8 or 8-18 of the first component(s);   c) a three-layer DNA dendrimer that comprises from about 2-6, 6-10, 10-18, 18-28, 28-38, 38-50 or more than 50 of the first components; and/or   d) a four-layer DNA dendrimer that comprises from about 15-25, 25-45, 45-60, 60-80, 85-105, 105-130 or more than 130 of the first components.   
     
     
         37 . The test device of  claim 36 , wherein the first component(s) comprise polynucleotide(s). 
     
     
         38 . The test device of  claim 37 , wherein the polynucleotide(s) are DNA strand(s). 
     
     
         39 . A method for detecting an analyte in a liquid sample, which method comprises:
 a) contacting a liquid sample with the test device of  claim 26 , wherein the liquid sample is applied to a site of the test device upstream of the test zone;   b) transporting an analyte, if present in the liquid sample, a detectable label and a DNA dendrimer to the test zone; and   c) assessing the presence, absence, and/or amount of a signal generated by the detectable label at the test zone to determining the presence, absence and/or amount of the analyte in the liquid sample.   
     
     
         40 . The method of  claim 39 , wherein the signal generated by the detectable label at the test zone is enhanced by:
 a) at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, or 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 folds or higher relative to a signal generated by the detectable label at the test zone using a DNA dendrimer comprising a smaller number of the first and/or second component(s); or   b) at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, or 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200 folds or higher relative to a signal generated by the detectable label at the test zone without using a DNA dendrimer.   
     
     
         41 . A test device for detecting an analyte in a liquid sample, which device comprises a porous matrix that comprises a test zone on said porous matrix, said test zone comprising a test reagent that binds to an analyte or another binding reagent that binds to said analyte, or is an analyte or an analyte analog that competes with an analyte in said sample for binding to a binding reagent for said analyte,
 wherein a liquid sample is capable of flowing laterally along said test device and passing said test zone to form a detectable signal to indicate presence, absence and/or amount of said analyte in said liquid sample,   wherein:   a) the formation of said detectable signal requires the use of a detectable label and a DNA dendrimer, said DNA dendrimer comprises a first component that links said DNA dendrimer to a binding reagent linkable to said DNA dendrimer, an analyte linkable to said DNA dendrimer, or an analyte analog linkable to said DNA dendrimer, and a second component that links said DNA dendrimer to said detectable label linkable to said DNA dendrimer, or   b) the formation of said detectable signal requires the use of a DNA dendrimer linked to a detectable label covalently or non-covalently, and said DNA dendrimer comprises a third component that links said DNA dendrimer to a binding reagent linkable to said DNA dendrimer, an analyte linkable to said DNA dendrimer, or an analyte analog linkable to said DNA dendrimer,   and wherein:   c) said analyte is not a polynucleotide; and/or   d) said DNA dendrimer comprises from about 1 to about 324, preferably from about 60 to about 324, from about 80 to about 300 or from about 100 to about 200, said first components; and/or   e) said DNA dendrimer, before said test device is used, is dried on a location on said test device upstream from said test zone.   
     
     
         42 . A method for detecting an analyte in a liquid sample, which method comprises:
 a) contacting a liquid sample with the test device of  claim 41 , wherein the liquid sample is applied to a site of the test device upstream of the test zone;   b) transporting an analyte, if present in the liquid sample, a detectable label and a DNA dendrimer to the test zone; and   c) assessing the presence, absence, and/or amount of a signal generated by the detectable label at the test zone to determining the presence, absence and/or amount of the analyte in the liquid sample.   
     
     
         43 . A kit for detecting an analyte in a liquid sample, which kit comprises:
 a) a porous matrix that comprises a test zone on said porous matrix, said test zone comprising a test reagent that binds to an analyte or another binding reagent that binds to said analyte, or is an analyte or an analyte analog that competes with an analyte in said sample for binding to a binding reagent for said analyte; and   b) a DNA dendrimer, wherein:
 1) said DNA dendrimer comprises a first component that links said DNA dendrimer to a binding reagent linkable to said DNA dendrimer, an analyte linkable to said DNA dendrimer, or an analyte analog linkable to said DNA dendrimer, and a second component that links said DNA dendrimer to said detectable label linkable to said DNA dendrimer, or 
 2) said DNA dendrimer is linked to a detectable label covalently or non-covalently and comprises a third component that links said DNA dendrimer to a binding reagent linkable to said DNA dendrimer, an analyte linkable to said DNA dendrimer, or an analyte analog linkable to said DNA dendrimer. 
   
     
     
         44 . The kit of  claim 43 , wherein the DNA dendrimer comprises from about 1 to about 324, preferably from about 60 to about 324, from about 80 to about 300 or from about 100 to about 200, of the first or third components. 
     
     
         45 - 46 . (canceled) 
     
     
         47 . The kit of any  claim 43 , wherein at least two of:
 1) the DNA dendrimer;   2) the first binding reagent that binds to the analyte, the second binding reagent that binds to another binding reagent that binds to the analyte, the analyte or the analyte analog, each of the first binding reagent, second binding reagent, analyte or analyte analog being linkable to the DNA dendrimer; and   3) the detectable label linkable to the DNA dendrimer;   are covalently bound or crosslinked to each other.   
     
     
         48 . The kit of any of  claim 43 , wherein the DNA dendrimer comprises from about 10 to about 1,500, preferably from about 40 to about 1,500 or from about 900 to about 1,100, the detectable label. 
     
     
         49 . The kit of any of  claim 43 , wherein the DNA dendrimer comprises from about 400 to about 80,000, preferably from about 4,000 to about 80,000, or from about 60,000 to about 80,000, DNA nucleotides. 
     
     
         50 . The kit of any of  claim 43 , wherein the DNA dendrimer comprises a one-layer, a two-layer, a three-layer or a four-layer structure, or a DNA dendrimer structure comprising at least one monomeric unit as described in U.S. Pat. No. 6,274,723B1. 
     
     
         51 . The kit of any of  claim 43 , which further comprises one or more of an antibody-oligo conjugate, an analyte capture antibody, streptavidin-label, e.g., gold particle, conjugate or other label, buffers, reagents, instructions etc.