(293f) Identification of Troponin I Binding Peptides Using Phage Display for Biosensor Development

Acute myocardial infarction (AMI) is the leading cause of morbidity and mortality in most industrialized nations throughout the world. Several clinical biomarkers are commonly used for the early detection of heart disease including the MB isoenzyme of creatine kinase (CK-MB), and myoglobin, however, there are some limitations to their use due to low specificity, selectivity and sensitivity. Troponin I has rapidly become the preferred diagnostic biomarker due to its high specificity and selectivity for myocardial injury.

We have used polyvalent phage display to isolate unique linear peptide motifs that recognize both the human and rat homologues of troponin I. The peptide specific for human troponin I has a sequence of FYSHSFHENWPS and the peptide specific for the rat troponin I has a sequence of FHSSWPVNGSTI. Enzyme-linked immunosorbent assays (ELISAs) were used to evaluate the binding interactions, and the two peptides exhibited some cross-reactivity, but they were both more specific for the troponin I homologue they were selected against. The binding affinities of the peptides were impacted in the presence of complex tissue culture media (MEM). And the addition of 10% calf serum further interfered with the binding of the target proteins. Kinetic indirect ELISAs revealed that both troponin I binding peptides were found to be nanomolar binders while attached to the phage particles. Specifically, the K_d value of peptide selected against rat troponin I was 0.95 ± 0.09 nM. The peptide obtained through selection against human troponin I had a K_d value of 2.5 ± 0.12 nM. These new peptides will potentially have utility in the development of new clinical assays for cardiac injury as well as in monitoring of cardiac cells in tissue culture applications.