Aptamer-Based Electrochemical Microfluidic Biosensor for the Detection of Cryptosporidium parvum

Abstract

Cryptosporidium parvum is a high-risk and opportunistic waterborne parasitic pathogen with highly infectious oocysts that can survive harsh environmental conditions for long periods. Current state-of-the-art methods are limited to lengthy imaging and antibody-based detection techniques that are slow, labor-intensive, and demand trained personnel. Therefore, the development of new sensing platforms for rapid and accurate identification at the point-of-care (POC) is essential to improve public health. Herein, we propose a novel electrochemical microfluidic aptasensor based on hierarchical 3D gold nano-/ microislands (NMIs), functionalized with aptamers specific to C. parvum. We used aptamers as robust synthetic biorecognition elements with a remarkable ability to bind and discriminate among molecules to develop a highly selective biosensor. Also, the 3D gold NMIs feature a large active surface area that provides high sensitivity and a low limit of detection (LOD), especially when they are combined with aptamers,. The performance of the NMI aptasensor was assessed by testing the biosensor's ability to detect different concentrations of C. parvum oocysts spiked in different sample matrices, i.e., buffer, tap water, and stool, within 40 min detection time. The electrochemical measurements showed an acceptable LOD of 5 oocysts mL −1 in buffer medium, as well as 10 oocysts mL −1 in stool and tap water media, over a wide linear range of 10−100,000 oocysts mL −1 . Moreover, the NMI aptasensor recognized C. parvum oocysts with high selectivity while exhibiting no significant cross-reactivity to other related coccidian parasites. The specific feasibility of the aptasensor was further demonstrated by the detection of the target C. parvum in patient stool samples. Our assay showed coherent results with microscopy and real-time quantitative polymerase chain reaction, achieving high sensitivity and specificity with a significant signal difference (p < 0.001). Therefore, the proposed microfluidic electrochemical biosensor platform could be a stepping stone for the development of rapid and accurate detection of parasites at the POC.

Unraveling the Possibilities: Recent Progress in DNA Biosensing

“…Even after 21 days of freezing, the sensor’s current reached 85% of the initial current effect. The sensor shows remarkable stability and is expected to be a promising candidate for point-of-care testing (POCT) application in parasite detection [ 19 ].…”

Microfluidic Platforms for Real‐Time In Situ Monitoring of Biomarkers for Cellular Processes

“…In Figure 3b, Mahshid et al report a microfluidic electrochemical biosensor based on layered 3D gold nanoislands, functionalized with an aptamer specific to Cryptosporidium parvum. [63] The 3D gold nanoisland demonstrates a large active surface area, which can significantly improve the sensitivity and low detection limits of microfluidic electrochemical biosensors, and nucleic acid aptamers further enhance the performance of the microfluidic platform. The biosensor detects Cryptosporidium parvum oocysts at 40 minutes.…”

“…Copyright 2023, American Chemical Society. [ 63 ] c) Copper ions in sweat detection. Reproduced with permission.…”

“…Reproduced with permission. Copyright 2023, American Chemical Society [63]. c) Copper ions in sweat detection.…”