Abstract
A simple in vitro biosensor for the detection of β-amyloid 42 in phosphate-buffered saline (PBS) and undiluted human serum was fabricated and tested based on our platform sensor technology. The bio-recognition mechanism of this biosensor was based on the effect of the interaction between antibody and antigen of β-amyloid 42 to the redox couple probe of K4Fe(CN)6 and K3Fe(CN)6. Differential pulse voltammetry (DPV) served as the transduction mechanism measuring the current output derived from the redox coupling reaction. The biosensor was a three-electrode electrochemical system, and the working and counter electrodes were 50 nm thin gold film deposited by a sputtering technique. The reference electrode was a thick-film printed Ag/AgCl electrode. Laser ablation technique was used to define the size and structure of the biosensor. Cost-effective roll-to-roll manufacturing process was employed in the fabrication of the biosensor, making it simple and relatively inexpensive. Self-assembled monolayers (SAM) of 3-Mercaptopropionic acid (MPA) was employed to covalently immobilize the thiol group on the gold working electrode. A carbodiimide conjugation approach using N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC) and N–hydroxysuccinimide (NHS) was undertaken for cross-linking antibody of β-amyloid 42 to the carboxylic groups on one end of the MPA. The antibody concentration of β-amyloid 42 used was 18.75 µg/mL. The concentration range of β-amyloid 42 in this study was from 0.0675 µg/mL to 0.5 µg/mL for both PBS and undiluted human serum. DPV measurements showed excellent response in this antigen concentration range. Interference study of this biosensor was carried out in the presence of Tau protein antigen. Excellent specificity of this β-amyloid 42 biosensor was demonstrated without interference from other species, such as T-tau protein.
Highlights
Diagnosis of neuro-degenerative disorders, Alzheimer’s Disease (AD), is difficult and inconclusive
The cerebrospinal fluid (CSF) biomarkers include total tau (T-tau), hyperphosphorylated tau (P-tau), and the 42 amino acid isoforms of amyloid β (β-amyloid 42) [1,2,3]. β-amyloid 42 pathophysiology leads to plaque deposition and can accelerate antecedent limbic and brain tauopathy [4,5,6]
The examination of β-amyloid 42 accumulated in senile plagues and intracellular neurofibrillary tangles of P-tau is considered the hallmark of identification of AD [2,3]
Summary
Diagnosis of neuro-degenerative disorders, Alzheimer’s Disease (AD), is difficult and inconclusive. Researchers have explored the use of different modified electrodes for bio-sensing applications, such as graphene and graphene-based materials, graphene hybrids, chemical doping, as well as magnetic doping materials including Fe3 O4 nano-magnetic particles [22,23,24,25,26,27] These modifications are elegant and elaborate but with limited sensor uniformity and reproducibility; calibration of individual biosensor is often required and time-consuming. 0.1 M and human serum were used as the test media The selectivity of this biosensor was examined using an interference study by the T-Tau in the concentrations of 0.125 μg/mL and 0.5 μg/mL at corresponding β-amyloid 42 antigen concentrations. All measurements were carried out at room temperature (23 ◦ C)
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