Abstract
In the present study, we fabricated a dual-mode cardiac troponin I (cTnI) biosensor comprised of multi-functional DNA (MF-DNA) on Au nanocrystal (AuNC) using an electrochemical method (EC) and a localized surface plasmon resonance (LSPR) method. To construct a cTnI bioprobe, a DNA 3 way-junction (3WJ) was prepared to introduce multi-functionality. Each DNA 3WJ arm was modified to possess a recognition region (Troponin I detection aptamer), an EC-LSPR signal generation region (methylene blue: MB), and an anchoring region (Thiol group), respectively. After an annealing step, the multi-functional DNA 3WJ was assembled, and its configuration was confirmed by Native-TBM PAGE for subsequent use in biosensor construction. cTnI was also expressed and purified for use in biosensor experiments. To construct an EC-LSPR dual-mode biosensor, AuNCs were prepared on an indium-tin-oxide (ITO) substrate using an electrodeposition method. The prepared multi-functional (MF)-DNA was then immobilized onto AuNCs by covalent bonding. Field emission scanning electron microscope (FE-SEM) and atomic force microscopy (AFM) were used to analyze the surface morphology. LSPR and electrochemical impedance spectroscopy (EIS) experiments were performed to confirm the binding between the target and the bioprobe. The results indicated that cTnI could be effectively detected in the buffer solution and in diluted-human serum. Based on the results of these experiments, the loss on drying (LOD) was determined to be 1.0 pM in HEPES solution and 1.0 pM in 10% diluted human serum. Additionally, the selectivity assay was successfully tested using a number of different proteins. Taken together, the results of our study indicate that the proposed dual-mode biosensor is applicable for use in field-ready cTnI diagnosis systems for emergency situations.
Highlights
As the obesity rate and the overall age of the population are both increasing worldwide, concerns of higher incidences of cardiovascular diseases (CVDs) are gradually growing [1]
The atomic force microscopy (AFM) experiments and surface roughness analysis were performed to confirm the formation of Au nanocrystal (AuNC) onto the ITO substrate and the immobilization process of cardiac troponin I (cTnI) onto the multi-functional DNA 3 way-junction (3WJ) on AuNC-modified substrate, respectively. (Figure 2a) indicates the topography of cleaned ITO substrate by AFM
The present study proposed a highly sensitive, label-free cTnI detection biosensor composed of multi-functional DNA on AuNC-modified ITO substrate that functions through a electrochemical method (EC)-localized surface plasmon resonance (LSPR) dual detection method
Summary
As the obesity rate and the overall age of the population are both increasing worldwide, concerns of higher incidences of cardiovascular diseases (CVDs) are gradually growing [1]. Among CVDs, acute myocardial infarction (AMI) in the aging society is one of the most serious diseases due to its high mortality rate and low recovery rate [3,4]. Upon experiencing AMI, the patient should be treated and transported to the hospital within 1 h for the highest chance of recovery. After 1 h with lack of proper treatment, the patient can experience multi-organ failure that can lead to death or loss of brain function. Rapid and appropriate treatment of AMI is critical for patient survival [5]
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