High-sensitive and fast detection of C-Reactive Protein and Troponin biomarkers using liquid-gated single silicon nanowire biosensors.

Fast, precise, and reliable detection of cardiac biomarkers is essentially important for early diagnosis and prognosis of cardiovascular diseases, the most common life-threatening illnesses. However, the current commercially available diagnostic methods either lack of the sensitivity needed for the proper detection of cardiac biomarkers at the very early disease development stage, or still suffer from the long-term process of enzymatic labeling. In this work, we designed and fabricated liquid-gated single silicon nanowire field-effect transistors (NW FETs) for label-free sensing of cardiac biomolecules. We demonstrate selective and direct detection of C-reactive protein (CRP) and cardiac troponin I (cTnI) proteins as gold standard biomarkers for prediction and detection of cardiovascular diseases at early stage. A detailed electrical characterization of fabricated nanosensors as well as the proof-of-principle experiments such as pH response were performed before sensing measurements. Sensor characteristics reflect high working stability and nearly ideal Nernstian sensitivity confirming the high-quality of fabricated structures. High specific recognition elements like aptamers and antibodies were immobilized on functionalized nanowire surfaces. A novel attachment approach was shown by using dibenzocyclooctyne (DBCO)-linked aptamers for cardiac troponin to azide-functionalized nanowire surface. The approach has several advantages: fast covalent and directed attachment of aptamers to azide-terminated surface of nanowires, efficient linkage and high-yield covering of nanowire surface with cTnI-specific aptamers. In particular, the DBCO-linked aptamers do not react with amines or hydroxyls presented in most of biomolecules, which makes functionalization of nanowires more efficient and specific. After covalent immobilization of cTnI-specific aptamers and CRP-specific antibodies on the silicon nanowires surface, the highly sensitive and fast electrical detection are performed. A 1 fg/ml detection limit concentration was achieved for both CRP and cTnI biomolecules. The target biomolecules on the Si NW surface are confirmed by atomic force microscopy (AFM) characterization. Moreover, application of nanowire structures with different sizes allowed us to detect biomarkers in a wide concentration range (from 1 fg/ml to 1 µg/ml). This range covers the dynamic concentration range of CRP and cTnI biomarkers release during the early disease stage and progression. Thus, we demonstrate that our specifically functionalized silicon NW FET biosensors integrated with polydimethylsiloxane (PDMS) microfluidic channels can be effectively used as highly sensitive diagnostic tools for early prediction and prevention of cardiovascular diseases