Enzyme immunosensor for diagnosis of myocardial infarction

Abstract

Amperometric enzyme immunosensors have been developed for the detection of the early myocardial infarction marker heart fatty acid binding protein (H-FABP) in blood plasma. They are based on anti-H-FABP antibodies immobilized on nitrocellulose or activated nylon membranes covering a modified Clark-type oxygen electrode. Two approaches have been tested using bovine H-FABP as a model: (i) the competition principle, where defined amounts of glucose oxidase (GOD)-labelled H-FABP compete with free H-FABP for binding to the immobilized capture antibodies; (ii) the sandwich principle, where the free H-FABP binds to the immobilized capture antibody and to a secondary GOD-labelled antibody, forming a sandwich. In both approaches, after addition of glucose, the activity of the enzyme label is determined with the amperometric electrode within minutes. The immunosensor using the sandwich principle on activated nylon membranes can be used repeatedly for at least 10 measurements after regeneration of the antibody membranes without loss of binding capacity of the capture antibodies. For the determination of human H-FABP, the sandwich principle with monoclonal anti-bovine-H-FABP antibodies as capture and GOD-labelled goat-anti-human-H-FABP antibodies as secondary antibodies is chosen. Initial experiments demonstrate that the sensitivity of the immunosensor is sufficient for the determination of clinical levels of H-FABP in plasma after myocardial infarction (100–300 ng ml −1).