Label Free Electrochemical cDNA-Biosensor for Selective and Sensitive Detection of Canine Distemper Virus in Biological Samples

Abstract

<title>Abstract</title> Canine distemper, a viral disease with a global impact on various animals including dogs, foxes, wolves, lions, and leopards, requires early diagnosis for effective treatment and outbreak prevention. Common laboratory methods, such as enzyme-linked immunosorbent assay, polymerase chain reaction, and viral isolation, face challenges such as extended turnaround time, high costs, and the expertise required. This study has developed a field-based biosensor for canine distemper detection, utilizing a screen-printed carbon electrode and a computer-assisted portable potentiostat. A 30-mer oligonucleotide capture probe, designed using the Primer3 Plus software version: 3.3.0, detects hybridization with the canine distemper virus complementary strand through electrochemical analysis via differential pulsed voltammetry. The developed biosensor exhibited good linearity in quantifying the target analyte concentration (0.1 µM to 12.8 µM) with a detection limit of 0.05 µM. Specificity tests using complementary and non-complementary sequences confirm the biosensor’s accuracy. The screen-printed electrode can be reused up to eight times with a residual capacity of 93.72 ± 5.45% after regeneration using a 50 mM NaOH solution. The developed biosensor was also used to detect the canine distemper virus from biological samples after the extraction of RNA and amplification. Results aligned with reverse transcriptase polymerase chain reaction findings, showing 100% agreement. These findings support the potential development of a field-deployable portable device for effectively diagnosing canine distemper in biological samples.