Proteolytic digestion of cardiac troponin I

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.