Myocardial infarction (MI) is highly related to cardiac arrest leading to death and organ damage. Radiological techniques and electrocardiography have been used as preliminary tests to diagnose MI; however, these techniques are not sensitive enough for early‐stage detection. A blood biomarker‐based diagnosis is an immediate solution, and due to the high correlation of troponin with MI, it has been considered to be a gold‐standard biomarker. In the present research, the cardiac biomarker troponin I (cTnI) was detected on an interdigitated electrode sensor with various surface interfaces. To detect cTnI, a capture aptamer‐conjugated gold nanoparticle probe and detection antibody probe were utilized and compared through an alternating sandwich pattern. The surface metal oxide morphology of the developed sensor was proven by microscopic assessments. The limit of detection with the aptamer‐gold‐cTnI‐antibody sandwich pattern was 100 aM, while it was 1 fM with antibody‐gold‐cTnI‐aptamer, representing 10‐fold differences. Further, the high performance of the sensor was confirmed by selective cTnI determination in serum, exhibiting superior nonfouling. These methods of determination provide options for generating novel assays for diagnosing MI.