# US 10,309,971 B2 · Thyroglobulin quantitation by mass spectrometry

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- Number: US 10,309,971 B2
- Document ID: US10309971B2
- Status: Granted
- Kind: B2
- Application number: 14/726,957
- Filed: 2015-06-01
- Published: 2016-02-11
- Granted: 2019-06-04
- Earliest claimed: 2007-12-06
- Assignees: Quest Diagnostics Investments Incorporated
- Applicants: Quest Diagnostics Investments Incorporated
- Inventors: Yanni Zhang; Nigel J. Clarke; Richard E. Reitz
- Web page: https://patentgrep.com/patent/US10309971B2

## CPC

- G01N 33/6848: {Methods of protein analysis involving mass spectrometry}
- G01N 33/68: involving proteins, peptides or amino acids {(involving lipoproteins G01N33/92)}
- G01N 33/6893: {related to diseases not provided for elsewhere}
- G01N 33/78: Thyroid gland hormones {, e.g. T3, T4, TBH, TBG or their receptors}
- G01N 2800/52: Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis
- G01N 2800/7028: Cancer
- Y10T 436/24: Nuclear magnetic resonance, electron spin resonance or other spin effects or mass spectrometry
- 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)
- Y10T: TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION

## Abstract

Provided are methods for determining the amount of thyroglobulin in a sample using various purification steps followed by mass spectrometry. The methods generally involve purifying thyroglobulin in a test sample, digesting thyroglobulin to form peptide T129, purifying peptide T129, ionizing peptide T129, detecting the amount of peptide T129 ion generated, and relating the amount of peptide T129 ion to the amount of thyroglobulin originally present in the sample.

## Claims

### Claim 1

A method for determining the amount of thyroglobulin in a test sample, comprising:
(a) enriching a digested thyroglobultin (Tg) peptide consisting of an amino acid sequence VIFDANAPVAVR (SEQ. ID NO: 4);
(b) ionizing the Tg peptide to generate one or more ions detectable by mass spectrometry
(c) quantifying the amount of the one or more Tg peptide ions from step (b) by mass spectrometry, wherein the amount of the one or more Tg peptide ions is related to the amount of thyroglobulin in said test sample.

### Claim 2 (depends on 1)

The method of claim 1, wherein the test sample comprises a body fluid.

### Claim 3 (depends on 2)

The method of claim 2, wherein the body fluid comprises serum or plasma.

### Claim 4 (depends on 1)

The method of claim 1, wherein the test sample comprises a thyroglobulin autoantibody.

### Claim 5 (depends on 1)

The method of claim 1, wherein the test sample is taken from a patient suffering from thyroid cancer.

### Claim 6 (depends on 1)

The method of claim 1, further comprising a digestion step that is carried out in the presence of trypsin.

### Claim 7 (depends on 1)

The method of claim 1, wherein the mass spectrometry is tandem mass spectrometry.

### Claim 8 (depends on 1)

The method of claim 1, wherein the ionizing step comprises ionizing the Tg peptide to generate a multiply charged ions detectable by mass spectrometry.

### Claim 9 (depends on 1)

The method of claim 1, further comprising purifying thyroglobulin from a body fluid or tissue sample to generate a thyroglobulin containing test sample.

### Claim 10 (depends on 1)

The method of claim 1, wherein the mass spectrometry is performed in positive ion mode.

### Claim 11 (depends on 1)

The method of claim 1, wherein the mass spectrometry is quadrupole or time-of-flight (TOF) mass spectrometry.

### Claim 12 (depends on 11)

The method of claim 11, wherein the TOF mass spectrometry is matrix-assisted laser desorption ionization coupled with TOF (MALDI-TOF).

### Claim 13 (depends on 1)

The method of claim 1 wherein the mass spectrometry comprises electrospray ionization.

### Claim 14 (depends on 1)

The method of claim 1, further comprising liquid chromatography.

### Claim 15 (depends on 14)

The method of claim 14, wherein the liquid chromatography is high performance liquid chromatography (HPLC).

### Claim 16 (depends on 1)

The method of claim 1, further comprising using a C18 column.

### Claim 17 (depends on 1)

The method of claim 1, further comprising protein precipitation.