Rapid Immunoassay for Acute Myocardial Injury Detection

Phase I is directed at the development of a rapid and quantitative diagnostic immunoassay for acuteinjury. This immunoassay will utilize a proven dry chemistry test system incorporating a cardiac-spechemically conjugated to active-site inactivated coagulation factor Xa. The test principle involvespeptide- Xa and blood troponin for an insolublized anti-troponin monoclonal antibody in the first stfollowed by UV irradiation to reactivate the factor Xa that subsequently converts prothrombin to thrpresence of fibrinogen results in a fibrin clot. The clotting time in this assay will be inversely poscillation of paramagnetic iron oxide particles in the dry chemistry test matrix. This proposal hasfirstly, Xa- troponin T peptide conjugates will be prepared and characterized. Conjugation will be agroups on Xa and a C- terminal sulfhydryl group on the peptide using a heterobifunctional crosslinkiof reacting with a free sulfhydryl group at one end and a free amino group at the other. In the secoApplicant intends to develop standard calibration curves for the troponin immunoassay using blood saknown amounts of peptide or troponin. Next, the Applicant will develop a test card-based assay formaperforming the troponin immunoassay on a single drop of blood using a modified TAS instrument retroflight source. Finally, the Applicant intends to validate the measurements against an established refblood samples obtained from myocardial infarction patients.