# US 9,039,888 B2 · Methods and apparatus for detecting molecular interactions using FET arrays

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- Number: US 9,039,888 B2
- Document ID: US9039888B2
- Status: Granted
- Kind: B2
- Application number: 14/033,934
- Filed: 2013-09-23
- Published: 2014-03-20
- Granted: 2015-05-26
- Earliest claimed: 2006-12-14
- Assignees: Life Technologies Corporation
- Applicants: Life Technologies Corporation
- Inventors: Jonathan Rothberg; Wolfgang Hinz
- Web page: https://patentgrep.com/patent/US9039888B2

## CPC

- C12Q 1/6825: Nucleic acid detection involving sensors
- C12Q 1/6874: involving nucleic acid arrays, e.g. sequencing by hybridisation
- G01N 27/26: by investigating electrochemical variables; by using electrolysis or electrophoresis
- G01N 27/4145: {specially adapted for biomolecules, e.g. gate electrode with immobilised receptors}
- G01N 27/4148: {Integrated circuits therefor, e.g. fabricated by CMOS processing}
- G01N 33/54373: {involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings}
- G01N 33/6818: {Sequencing of polypeptides}
- H10D 30/60: Insulated-gate field-effect transistors [IGFET] (H10D30/40 takes precedence)
- H10D 30/68: Floating-gate IGFETs
- C12Q 1/6869: Methods for sequencing
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- H10D: INORGANIC ELECTRIC SEMICONDUCTOR DEVICES

## Abstract

Methods and apparatuses relating to large scale FET arrays for analyte detection and measurement are provided. ChemFET (e.g., ISFET) arrays may be fabricated using conventional CMOS processing techniques based on improved FET pixel and array designs that increase measurement sensitivity and accuracy, and at the same time facilitate significantly small pixel sizes and dense arrays. Improved array control techniques provide for rapid data acquisition from large and dense arrays. Such arrays may be employed to detect a presence and/or concentration changes of various analyte types in a wide variety of chemical and/or biological processes.

## Claims

### Claim 1

A method for sequencing a polypeptide, comprising:
disposing a plurality of polypeptides into each of a plurality of reaction chambers, at least one reaction chamber of the plurality of reaction chambers coupled to at least one chemical-sensitive field effect transistor (chemFET) having a floating gate;
sequentially introducing amino acyl synthetases into the at least one reaction chamber, the amino acyl synthetase operable to bind to a specific amino acid when present at a free end of the polypeptide;
detecting the presence of an amino acyl synthetase bound to the free end of the polypeptide; and
cleaving the amino acid from the free end of the polypeptide.

### Claim 2 (depends on 1)

The method of claim 1, wherein detecting the presence of the amino acyl synthetase includes detecting a detection moiety conjugated to the amino acyl synthetase.

### Claim 3 (depends on 2)

The method of claim 2, wherein the detection moiety includes an enzyme.

### Claim 4 (depends on 3)

The method of claim 3, wherein the detecting the detection moiety includes measuring a change in the electrical properties of the chemFET in response to a reaction catalyzed by the enzyme.

### Claim 5 (depends on 1)

The method of claim 1, wherein the plurality of polypeptides are bound to a bead, and at least one bead is disposed into each of the plurality of reaction chambers.

### Claim 6 (depends on 1)

The method of claim 1, further comprising denaturing the polypeptide.

### Claim 7 (depends on 1)

The method of claim 1, wherein cleaving further includes cleaving the amino acid using a protease conjugated to the amino acyl synthetase.

### Claim 8 (depends on 1)

The method of claim 1, wherein cleaving further includes cleaving the amino acid using a limited Edmond degradation or a proximity based radical cleavage.

### Claim 9 (depends on 1)

The method of claim 1, wherein the floating gate of the at least one chemFET is coupled to the at least one reaction chamber via a passivation layer sensitive to hydrogen ions.

### Claim 10

A method for sequencing a polypeptide, comprising:
disposing a plurality of polypeptides into at least one of a plurality of reaction chambers, at least one reaction chamber of the plurality of reaction chambers coupled to at least one chemical-sensitive field effect transistor (chemFET) having a floating gate;
sequentially cleaving amino acids from an end of the polypeptide;
sequentially detecting the identity of the amino acids by a change in an electrical property of the at least one chemFET having the floating gate.

### Claim 11 (depends on 10)

The method of claim 10, wherein a center-to-center distance between adjacent reaction chambers is selected from a range of about 1 μm to about 10 μm.

### Claim 12 (depends on 10)

The method of claim 10, wherein the plurality of reaction chambers includes at least 105 reaction chambers.

### Claim 13 (depends on 10)

The method of claim 10, wherein the floating gate of the at least one chemFET is coupled to the at least one reaction chamber via a passivation layer sensitive to hydrogen ions.

### Claim 14 (depends on 10)

The method of claim 10, wherein the change in the electrical property is due to a change in threshold voltage of the at least one chemFET resulting from a change in ion concentration related to a chemical reaction catalyzed by a detection moiety.

### Claim 15 (depends on 10)

The method of claim 10, wherein the detection moiety is conjugated to an amino acyl synthetase specific for the amino acid being detected.

### Claim 16 (depends on 15)

The method of claim 15, wherein the detection moiety is a hydrogen peroxidase.

### Claim 17

A method for sequencing a polypeptide, comprising:
disposing a plurality of polypeptides into at least one of a plurality of reaction chambers, at least one reaction chamber of the plurality of reaction chambers coupled to at least one chemical-sensitive field effect transistor (chemFET) having a floating gate;
sequentially cleaving amino acids from an end of the polypeptide;
detecting a change in the level of a reaction product of a detection moiety conjugated to an amino acyl synthetase as an indicator of the identity of the amino acid.

### Claim 18 (depends on 17)

The method of claim 17, wherein the amino acyl synthetase is bound to the amino acid and is specific for the amino acid.

### Claim 19 (depends on 17)

The method of claim 17, wherein the change in the level of the reaction product comprises detecting a threshold voltage of the at least one chemFET.

### Claim 20 (depends on 17)

The method of claim 17, wherein cleaving amino acids from the end of the polypeptide involves a protease conjugated to the amino acyl synthetase.