Labels for electronic detection of individual molecules and methods for their detection

Abstract

The invention provides labels for electronic detection of individual molecules. The labels are comprised of elements with different electrical properties that affect the electric current flowing through a nanoelectrode. The labels are of polymeric or filamentous structure where the elements are arranged linearly along their length. The arrangement of the elements is predetermined and combinatorial, so that a high diversity of labels can be generated in a manner that resembles barcoding on a nanoscale level. Methods for the synthesis of said barcode labels and for the binding of the barcode labels to individual molecules, their movement past a nanoelectrode, and their detection are also provided.

1 . A label for the electronic detection of individual target analyte molecules comprising a target specific probe capable of binding the target analyte molecule attached to a polymeric molecule or filament, wherein the polymeric molecule or filament comprises one or more elements with different electrical properties as detected by a nanoelectrode and the elements with different electrical properties are not detected by changes in fluorescence. 
     
     
         2 . The label according to  claim 1  wherein the target analyte molecule is a nucleic acid, a protein, a polysaccharide, a polypeptide, a carbohydrate or a small molecule. 
     
     
         3 . The label according to  claim 2  wherein the target analyte molecule is a nucleic acid. 
     
     
         4 . The label according to  claim 1  wherein the target specific probe is a nucleic acid molecule complementary to the target analyte molecule, a nucleic acid mimetic, a locked nucleic acid, a nucleic acid aptamer, a peptide, a protein, a receptor, a ligand, an antibody, an antibody fragment or a lecitin molecule. 
     
     
         5 . The label according to  claim 4  wherein the target specific probe is a nucleic acid molecule complementary to the target analyte molecule. 
     
     
         6 . The label according to  claim 1  wherein the polymeric molecule is a nucleic acid molecule, a protein, a peptide or a polyacrylic copolymer. 
     
     
         7 . The label according to  claim 8  wherein the polymeric molecule is a nucleic acid molecule. 
     
     
         8 . The label according to  claim 1  wherein the polymeric material or filament comprises from two to six elements with different electrical properties as detected by a nanoelectrode. 
     
     
         9 . The label according to  claim 8  wherein the polymeric material or filament comprises two alternating elements with different electrical properties as detected by a nanoelectrode. 
     
     
         10 . The label according to  claim 9  wherein the first of the two elements causes a greater decrease in current in the nanoelectrode than the second of the two elements. 
     
     
         11 . The label according to  claim 10  wherein the polymeric material is a nucleic acid molecule which has been modified to contain one or more biotin residues, the first of the two elements comprises a strepavidin-coated nanoparticle and the second of the two elements comprises an unmodified portion of the nucleic acid molecule. 
     
     
         12 . The label according to  claim 1  further comprising one or more additional elements capable of providing sufficient stretching of the label to allow each of the elements with different electrical properties to be separately detected by the nanoelectrode. 
     
     
         13 . The label according to  claim 12  having one additional element comprising a magnetic nanosphere. 
     
     
         14 . The label according to  claim 12  having a first additional element comprising a superparamagnetic nanosphere and a second additional element comprising a diamagnetic nanosphere. 
     
     
         15 . The label according to  claim 12  having a first additional element comprising a magnetic nanosphere and a second additional element comprising a hydrophobic moiety. 
     
     
         16 . A composition comprising two or more labels, each of which comprises a target specific probe capable of binding a target analyte molecule attached to a polymeric molecule or filament wherein the polymeric molecule or filament comprises one or more elements with different electrical properties as detected by a nanoelectrode, and wherein the elements with different electrical properties are not detected by changes in fluorescence and the target specific probe of each label is different from each of the other labels. 
     
     
         17 . A kit for the electronic detection of individual target analyte molecules comprising:
 (a) one or more labels each of which comprises a target specific probe capable of binding the target analyte molecule attached to a polymeric molecule or filament which comprises at least two elements with different electrical properties; and   (b) a nanoelectrode capable of detecting the elements with different electrical properties.   
     
     
         18 . The kit according to  claim 17  wherein the target specific probe is a nucleic acid molecule complementary to the target analyte molecule and the polymeric molecule is a nucleic acid molecule. 
     
     
         19 . The kit according to  claim 17  comprising two or more labels wherein the target specific probe of each label is different from each of the other labels. 
     
     
         20 . The kit according to  claim 17  wherein the polymeric material or filament comprises two elements with different electrical properties as detected by a nanoelectrode. 
     
     
         21 . The kit according to  claim 17  wherein the label further comprises one or more additional elements capable of providing sufficient stretching of the label to allow each of the elements with different electrical properties to be separately detected. 
     
     
         22 . The kit according to  claim 21  having one additional element comprising a magnetic nanosphere. 
     
     
         23 . The kit according to  claim 21  having a first additional element comprising a magnetic nanosphere and a second additional element comprising a hydrophobic moiety. 
     
     
         24 . The kit according to  claim 17  wherein the nanoelectrode has a critical dimension of about 20 nm to about 100 nm. 
     
     
         25 . The kit according to  claim 17  wherein the nanoelectrode has a critical dimension of less than about 100 nm. 
     
     
         26 . The kit according to  claim 17  wherein the nanoelectrode is fabricated from gold, platinum, copper or carbon nanotubes. 
     
     
         27 . The kit according to  claim 17  further comprising one or more additional nanoelectrodes fabricated to form an array. 
     
     
         28 . The kit according to  claim 48  wherein the redox mediator is 1,4-benzoquinone, ferrocene carboxylic acid, phenazine methosulphate, tetrathiofulvalene, tetracyanoquinodimethane, ferrocyanides, dimethylviologen, 1,1′-dimethylferrocene, phenazine ethosulfate or ferrocenylmethanol. 
     
     
         29 . The kit according to  claim 17  wherein the nanoelectrode is located at the bottom of a channel in a silicon, glass or plastic chip. 
     
     
         30 .- 47 . (canceled) 
     
     
         48 . The kit according to  claim 17  further comprising a redox mediator capable of undergoing redox reactions at a surface of the nanoelectrode.