Filer, Jessica, authorGeiss, Brian, advisorChen, Tom, advisorHenry, Charles, committee memberWilusz, Jeff, committee memberEbel, Greg, committee member2019-06-142020-06-102019https://hdl.handle.net/10217/195262Zika virus (ZIKV) emerged as a significant public health concern after the 2015-2016 outbreak in South and Central America. Severe neurological complications and birth defects in adults and children respectively underscore the need for quick and accurate diagnosis so that proper medical observation and intervention can be done. Electrochemical assays and biosensors are attractive as alternative diagnostic tools due to their sensitivity and ease of miniaturization. This dissertation describes three novel electrochemical assays and biosensors to detect ZIKV specific nucleic acid, antibodies, and virus particles. A nuclease protection ELISA (NP-ELISA) was developed for nucleic acid detection by enzymatic readout. The assay was validated using synthetic complementary oligos for absorbance, chemiluminescence, and electrochemical enzymatic readout. Two horseradish peroxidase substrates, 3,3',5,5'-Tetramethylbenzidine (TMB) and hydroquinone, were characterized electrochemically and compared for electrochemical assay use. Electrochemical TMB readout demonstrated better sensitivity compared to all tested detection modalities with a limit of detection of 3.72×103 molecules mL-1, which compares well to the amount of ZIKV RNA in clinical samples and to other approved assays like the CDC's Trioplex assay. For serological analysis, a capacitive microwire biosensor was developed and validated using immunized mouse sera to detect a ZIKV antibody response. Measurements were taken through a wide serial dilution range of 1:1018 to 1:103 and two dilutions (1:1012 and 1:106) were used for analysis for optimal sensitivity. A statistically significant immune response was detected four days after immunization at a 1:1012 dilution and was specific for ZIKV when compared with Chikungunya virus (CHIKV). These results indicate that serological analysis can be performed four days earlier with the wire sensor compared to ELISAs using ultra-dilute samples. The sensor also was used to differentiate between IgG and IgM antibodies and compared well with ELISA results. Lastly, an impedance array sensor was designed and validated for detection of ZIKV particles. The array allows for simultaneous handling of many electrodes, which increases throughput compared to other biosensor designs. The sensor demonstrated good sensitivity with an LOD of 22.4 focus forming units (FFU) which compares well to other reported sensors. In addition, it was optimized for specificity and tested using Sindbis virus (SINV) as a negative control. These novel platforms comprise new advancements in biosensor technology by simplifying existing assays, increasing sensitivity, and providing a new platform for handheld measurements.born digitaldoctoral dissertationsengCopyright and other restrictions may apply. User is responsible for compliance with all applicable laws. For information about copyright law, please see https://libguides.colostate.edu/copyright.diagnosticsimmunoassaybiosensorelectrochemicalZika virusText