Nanosensor chip with compound nanopores

Abstract

A nanosensor chip for detecting and/or quantifying target molecules in a liquid sample includes a semiconductor or other substrate and one or more electrode structures. The substrate has one or more compound nanopores, referred to as “compores.” Each compore is an aperture formed in the substrate and comprises a plurality of nanopores. Each of the nanopores is functionalized with immobilized probe molecules for detecting the target molecules. For each compore, a corresponding electrode structure is laid out on the substrate. The electrode structure has a shape and a position relative to the compore to apply an electric field across all of the nanopores in the compore and to provide a conductive path for an aggregate current through all of the nanopores in the compore. The aggregate current changes in response to target molecules in the liquid sample binding to the probe molecules as a function of the electric field.

1 . A nanosensor chip for detecting and/or quantifying target molecules in a liquid sample, the nanosensor chip comprising:
 a substrate with one or more compound nanopores (“compores”), wherein each compore comprises an aperture formed in the substrate and further comprises a central region and a sloped sidewall, wherein the central region is thinner than the surrounding substrate and comprises therein a plurality of nanopores with a height that is the same as the central region, further wherein each of the nanopores has a larger opening and a smaller opening and a sidewall with a slope between 6 and 60 degrees between the two openings, and each of the nanopores is functionalized with a plurality of immobilized probe molecules affixed to the sidewall of the nanopore, the probe molecules for binding the target molecules; and   for each compore, a corresponding electrode that surrounds the plurality of nanopores in the central region, wherein an aggregate current conducted by the electrode through all of the nanopores in the compore changes in response to target molecules in the liquid sample binding to the probe molecules as a function of an electric field applied by the electrode across all of the nanopores in the compore.   
     
     
         2 . The nanosensor chip of  claim 1 , wherein the substrate is a semiconductor substrate. 
     
     
         3 . The nanosensor chip of  claim 2 , wherein the aperture for each compore includes a region of the semiconductor substrate thinned to less than 1 micron, and the thinned region contains all of the nanopores for that compore. 
     
     
         4 . The nanosensor chip of  claim 2 , wherein the electrode for each compore is laid out on one side of the semiconductor substrate. 
     
     
         5 . The nanosensor chip of  claim 1 , wherein each compore has a maximum width between 1 micron and 1000 microns. 
     
     
         6 . The nanosensor chip of  claim 1 , wherein each nanopore has a maximum width between 1 nanometer and 300 nanometers. 
     
     
         7 . The nanosensor chip of  claim 2 , wherein the nanosensor chip comprises a plurality of compores and corresponding electrodes laid out on the semiconductor substrate, and the electrodes are configured so that the aggregate current through all of the nanopores in each compore may be individually measured. 
     
     
         8 . The nanosensor chip of  claim 2 , wherein the nanosensor chip comprises a plurality of compores and corresponding electrodes laid out on the semiconductor substrate, and the electrodes are configured so that different electric fields may be either individually applied across each compore or collectively applied across all compores simultaneously. 
     
     
         9 . The nanosensor chip of  claim 1 , wherein the probe molecules have a binding affinity to the target molecules, the binding of the target molecules to the probe molecules causes a change in the aggregate current through all of the nanopores in the compore as a function of the electric field applied across the compore, and the change in the aggregate current indicates a presence and concentration of the target molecules in the liquid sample. 
     
     
         10 . The nanosensor chip of  claim 1 , wherein, for at least one of the compores, each of the plurality of nanopores in that compore are functionalized with a same type of probe molecule. 
     
     
         11 . The nanosensor chip of  claim 1 , wherein at least one compore contains nanopores of two or more different sizes. 
     
     
         12 . The nanosensor chip of  claim 1 , wherein the nanosensor chip comprises a plurality of compores all functionalized with a same type of probe molecule. 
     
     
         13 . The nanosensor chip of  claim 1 , wherein the nanosensor chip comprises a plurality of compores and at least two of the compores are functionalized with a different type of probe molecule. 
     
     
         14 . The nanosensor chip of  claim 1 , wherein, for at least one of the compores, the nanopores in that compore are functionalized with two or more different types of probe molecule. 
     
     
         15 . The nanosensor chip of  claim 1 , wherein at least one of the nanopores is functionalized with two or more different types of probe molecule. 
     
     
         16 . The nanosensor chip of  claim 1 , wherein the probe molecules comprise antibodies or antibody analogs. 
     
     
         17 . The nanosensor chip of  claim 1 , wherein the probe molecules comprise nanobodies. 
     
     
         18 . The nanosensor chip of  claim 1 , wherein the probe molecules comprise proteins. 
     
     
         19 . The nanosensor chip of  claim 1 , wherein the probe molecules comprise aptamers. 
     
     
         20 . The nanosensor chip of  claim 1 , wherein the probe molecules comprise polymers. 
     
     
         21 . The nanosensor chip of  claim 1 , wherein the probe molecules comprise oligonucleotides. 
     
     
         22 . The nanosensor chip of  claim 1 , wherein each of the nanopores is functionalized with at least two types of probe molecules selected from a group consisting of antibodies, antibody analogs, proteins, aptamers, polymers, oligonucleotides, and nanobodies. 
     
     
         23 . The nanosensor chip of  claim 1 , wherein the probe molecules are attached to the nanopores by covalent binding, non-covalent binding, or physisorption. 
     
     
         24 . (canceled) 
     
     
         25 . The nanosensor chip of  claim 1 , wherein at least one of the plurality of nanopores is blocked and yet the aggregate current through all of the nanopores changes in response to the target molecules in the liquid sample binding to the probe molecules in the presence of the electric field even if one of the plurality of nanopores is blocked. 
     
     
         26 . The nanosensor chip of  claim 25 , wherein the plurality of nanopores are not uniformly functionalized by the probe molecules and yet the aggregate current through all of the nanopores changes in response to the target molecules in the liquid sample binding to the probe molecules in the presence of the electric field even if the plurality of nanopores are not uniformly functionalized by the probe molecules. 
     
     
         27 . The nanosensor chip of  claim 1 , wherein the target molecules bind to the probe molecules at a specific voltage applied across the compore, and the target molecules and the probe molecules, when subjected to an application of a varying voltage that includes the specific voltage, continuously bind and release. 
     
     
         28 . The nanosensor chip of  claim 27 , wherein upon application of the specific voltage, if the target molecules are present in the liquid sample, the target molecules flow inside the plurality of nanopores and bind to the probe molecules, and the binding of the target molecules to the probe molecules results in an ionic current change. 
     
     
         29 . (canceled) 
     
     
         30 . The nanosensor chip of  claim 1 , wherein the probe molecules remain affixed to the sidewalls of the nanopores after use, and the compore is configured to be used multiple times. 
     
     
         31 . A detection system for detecting and/or quantifying target molecules in a liquid sample, the detection system comprising:
 a nanosensor chip comprising:
 a substrate with one or more compound nanopores (“compores”), wherein each compore comprises an aperture formed in the substrate and further comprises a central region and a sloped sidewall, wherein the central region is thinner than the surrounding substrate and comprises a plurality of nanopores with a height that is the same as the central region, further wherein each of the nanopores has a larger opening and a smaller opening and a sidewall with a slope between 6 and 60 degrees between the two openings, and each of the nanopores is functionalized with a plurality of immobilized probe molecules affixed to the sidewall of the nanopore for detecting the target molecules; and 
 for each compore, a corresponding electrode attached to the compore and that surrounds the plurality of nanopores in the central region, wherein an aggregate current conducted by the electrode through all of the nanopores in the compore changes in response to target molecules in the liquid sample binding to the probe molecules as a function of an electric field applied by the electrode across all of the nanopores in the compore; 
   a variable voltage source coupled to a circuit between an upper electrode on a top surface of the compore and a lower electrode on a bottom side of the compore;   a current detector coupled to the circuit; and   a controller coupled to control the variable voltage source and current detector, the controller configured to:
 control the variable voltage source to apply a range of different voltages across all of the plurality of nanopores in the compore, wherein binding of target molecules to probe molecules is a function of the electric fields across the compore caused by the different voltages; and 
 control the current detector to detect an aggregate current through all of the plurality of nanopores in the compore, wherein binding of target molecules to probe molecules causes changes in the aggregate current, and the changes in the aggregate current as a function of the applied electric field indicate a presence and/or concentration of the target molecules in the liquid sample.