[Show abstract][Hide abstract] ABSTRACT: Picogram levels of antibodies against the protective antigen (PA) of Bacillus anthracis were detected in an automated electrochemical sandwich-type enzyme-linked immunosorbant assay. The antibodies were captured and detected using an 8 x 3 array of 50-microm-diameter cavities. The reagent and sample volumes were as low as 200 nL in a less than 25-min assay from capture to signal generation. The electrochemical detection of the antibodies was demonstrated at 0.05-10 microg/mL containing only 10-5,000 pg antibodies. The limit of detection is 10 fg for a 200-nL sample. Detection of anti-PA immunoglobulin G performed in spiked normal human serum and fresh whole human blood did not show a significant difference from detection in a buffer. The initial automation of the assay involved the use of a digital syringe pump for the delivery of reagents to the capture surface.
No preview · Article · Oct 2007 · Analytical and Bioanalytical Chemistry
[Show abstract][Hide abstract] ABSTRACT: Studies toward the development of automated, rapid, sensitive, low-cost, and reliable biomedical and environmental assays using arrays of microcavities with three individually addressable electrodes is reported. Each microcavity in the array contains a bottom electrode called recessed microdisk electrode (or RMD), a middle layer electrode found along the wall called tubular nanoband electrode (or TNB), and a rim/top electrode. The microelectrode TNB was used to detect the signal from a microwell assay for as low as 100 pg of Plasmodium falciparum CSP gene over 5-min enzyme substrate incubation. A self-contained microelectrochemical assay for Cryptosporidium parvum was generated using the RMD for capture, the TNB as the detecting electrode, and the rim/top as the pseudoreference/auxiliary electrodes. The 4-μm distance of the capture surface to the TNB detecting electrode provided close proximity to the assay components which allowed for a rapid response where the first signal was recorded within 30 s of enzyme substrate incubation. The self-contained nature of the microcavity provided low cost assays with the use of reagents down to 200 nL. The self-contained microelectrochemical microarrays were compatible with automation that was demonstrated in the detection of 25 pg Bacillus anthracis anti-PA IgG in < 25 min total assay time.