C-reactive protein (CRP) and cardiac troponin I (cTnI) biomolecules represent the earliest enzymes that appear in the blood when a cardiac injury occurs. Real-time and selective detection of these biomarkers is essential for the prediction and detection of cardiovascular diseases at an early stage. Here we report on the label-free specific detection of both proteins at picomolar concentrations using fabricated nanowire-based biosensors. We demonstrate a novel functionalization technique based on the attachment of dibenzocyclooctyne (DBCO)-linked troponin-specific aptamers to azide-functionalized silicon (Si) nanowire (NW) surface. Due to the fast and reliable immobilization of cTnI-specific aptamers and CRP-specific antibodies on the Si NWs, the fabricated devices can rapidly detect target biomolecules demonstrating high sensitivity. We confirm the attachment of proteins to the surface of Si NWs by atomic force microscopy (AFM). Moreover, we demonstrate that nanowire structures of different sizes enable the detection of biomarkers in a wide concentration range (from 1 pg/ml to 1 µg/ml), corresponding to CRP and cTnI elevation levels during the early stage of disease formation.