Proteolytic digestion of cardiac troponin I

Patent metadata

Field	Value
Publication number	US-10472403-B2
Application number	US-201515535964-A
Country	US
Kind code	B2
Filing date	Dec 23, 2015
Priority date	Dec 23, 2014
Publication date	Nov 12, 2019
Grant date	Nov 12, 2019

Abstract

A fluid sample container comprising a protease, when a sample fluid is placed in the fluid sample container the protease breaking a target analyte in the sample fluid into at least two peptides, the at least two peptides being smaller than the original target analyte.

First claim

What is claimed is:

1. A diagnostic assay for determining a level of cardiac Troponin I (cTnI) in a whole blood sample fluid, the assay comprising the steps of:
(a) receiving the whole blood sample fluid comprising cTnI, wherein the whole blood sample fluid further contains heparin, wherein the heparin prevents red blood cells (RBCs) in the whole blood sample from clotting, wherein the heparin binds to individual RBCs, and wherein the heparin binds to cTnI;
(b) mixing the whole blood sample fluid of (a) with chymotrypsin to create a sample fluid mixture, wherein the chymotrypsin breaks the cTnI present in the whole blood sample fluid into at least three peptides, the at least three peptides comprising at least an N-terminal peptide fragment, a C-terminal peptide fragment, and a middle peptide fragment, wherein the heparin present in the whole blood sample fluid binds to the C-terminal peptide fragment of cTnI;
(c) filtering the sample fluid mixture through a sample fluid filter such that the sample fluid filter prevents the heparin-bound RBCs and the heparin-bound C-terminal peptide fragment of cTnI from flowing there through and outputs the N-terminal peptide fragment of cTnI and the middle peptide fragment of cTnI that are not bound by heparin to flow there through to produce a filtered sample fluid mixture; and
(d) determining an amount of the middle peptide fragment of cTnI in the filtered sample fluid mixture.

2. The diagnostic assay of claim 1 , wherein the middle peptide fragment of cTnI has the amino acid sequence represented by SEQ ID NO:3.

3. The diagnostic assay of claim 1 , wherein the C-terminal peptide fragment of cTnI has the amino acid sequence represented by SEQ ID NO:4.

4. The diagnostic assay of claim 1 , wherein the step of determining an amount of the middle peptide fragment of cTnI uses a detection technology selected from fluorescence, chemiluminescence, electrochemical luminescence, electrochemically, and luminescent oxygen channeling immunoassay (LOCI).

5. The diagnostic assay of claim 1 , wherein the N-terminal peptide fragment of cTnI has the amino acid sequence represented by SEQ ID NO:2.

Assignees

• Siemens Healthcare Diagnostics Inc

Inventors

• Ledden David
• Cowden Eric

Classifications

• G01N33/6887

  from muscle, cartilage or connective tissue · CPC title

• G01N2333/4712

  Muscle proteins, e.g. myosin, actin, protein · CPC title

• C07K14/475 Primary

  Growth factors; Growth regulators · CPC title

• C12Q1/37 Primary

  involving peptidase or proteinase · CPC title

• G01N2333/976

  Trypsin; Chymotrypsin · CPC title