Myocardial infarction (MI) is a severe cardiovascular event that can lead to death. Cardiac troponin I (cTnI) is an MI biomarker in the circulation system; however, methods for detecting cTnI protein require substantial time, tedious operations, an expensive reader for translating signals, and a lot of reagents. This study aims to create a cTnI protein test kit with results easily distinguished by color differences, explicitly focusing on the resolution between different concentrations that eyes can discern. These results will aid in creating a commercial, portable, convenient, daily-use rapid-test kit. This study proposes a cTnI biosensor that the naked eye can perceive, performs diagnoses based on pattern color, does not require a reader machine, is easy to operate, and is portable. Our device shortens diagnosis time, has a 0.32−200 ng/mL quantitative analysis range in the human serum matrix, achieves a 0.32 ng/mL limit of detection, and exhibits many advantages compared to a traditional cTnI ELISA plate.