Design Considerations for DNA-based Lateral Flow Strip Biosensors for Possible Cd(II) Detection
Date of Award
Master of Science in Chemistry
Erwin P. Enriquez, PhD
Cadmium (Cd) is a naturally occurring element and is classified as an extremely toxic heavy metal. It is still nonetheless used in industrial workplaces that could lead to the contamination of water, air and soil and eventual accumulation in crops and animals that are grown for human consumption. Atomic absorption spectroscopy and inductively coupled plasma-mass spectrometry offer high sensitivity and accuracy for Cd detection though they are not amenable to on-site, field-based testing being lab- based instruments in general. There is a need for a field-ready test for Cd(II) for rapid, cost-effective, and extensive screening of Cd(II) in the environment and water effluents from human settlements. In this study, the lateral flow strip biosensor was investigated as a possible platform for Cd(II) detection mainly for its ease-of-use and affordability. The study tested different approaches to detect Cd(II) by DNA aptamers as biorecognition element, which are conjugated to gold nanoparticles (AuNPs) to serve as visual colorimetric labels. The first sensor design studied was based on the target-induced desorption of DNA aptamers in AuNP surface, thereby allowing the bare AuNPs to be captured in the detection zone. Preliminary colorimetric studies showed that increasing Cd(II) caused the aggregation of AuNPs with a linear working range of 300-3000 ppb and detection and quantification limit of 290 ppb and 965 ppb, respectively. Polyelectrolyte assembly and bovine serum albumin were used as the components of the control and test lines in the detection zones, respectively. The sensor probes used were DNA-coated AuNPs that showed tolerance against aggregation in highly ionic environment compared to the bare AuNPs. However, in the lateral flow strip biosensor, the polyelectrolyte assemblies indiscriminately captured (by AuNP aggregation) the bare AuNPs and DNA-coated AuNPs at the test line, whereas bovine serum albumin was not able to capture bare AuNPs or the DNA-coated AuNPs even up to 100 μM of bovine serum albumin. The second design tested utilized a competitive binding mechanism wherein the Cd(II) in the migrating sample competed against the immobilized DNA capture probe for binding to the Cd-specific DNA aptamer in the sensor probes. For this competitive- version of lateral flow strip biosensor, appearance of a line on the test line would be positive detection of Cd(II). Two DNA capture probes were designed with varying theoretical free energy of binding (ΔG37) values using an online software. After the optimization of the parameters such as concentration and volume of DNA capture probes and DNA-AuNP ratios, the lateral flow strip biosensor/s yielded false negative results from 1 ppb to 10 ppm of Cd(II). Further, high Cd(II) concentrations (100 ppm and 500 ppm) caused the aggregation of the sensor probes in the conjugate pad, thus no lines were observed in the detection zone. It is possible that the DNA hybridization is favored to Cd(II)-DNA aptamer binding, which attributed to the formation of the test lines even at high Cd(II) concentrations. Despite the negative results, these two test cases point to other directions for lateral flow strip biosensor design for a DNA aptamer-based lateral flow strip biosensor design for Cd(II) detection.
(2019). Design Considerations for DNA-based Lateral Flow Strip Biosensors for Possible Cd(II) Detection. Ateneo de Manila University.