Abstract
Microarray printing of components of an enzyme-linked immunosorbant assay (ELISA) is attractive for constructing miniaturized platforms to simultaneously and quantitatively analyze multiple chemical species in clinical, environmental, and forensic applications. A truncated version of an electrochemical ELISA was used to address a major challenge of this approach, namely the effect of evaporation on assay activity. An antibody-alkaline phosphatase conjugate (Ab-AP) was printed with a microarrayer onto an N-hydroxysuccinimide-ester-terminated self-assembled monolayer on gold. The activity of the enzyme label AP to convert the substrate p-aminophenyl phosphate to electroactive p-aminophenol was followed as a function of different conditions via detection by cyclic voltammetry at microband electrodes. It was found that enzyme activity decreased by 60% from 30 min to 4 h after evaporation. However, printing in ambient humidity ("dry") as opposed to printing in a humidified chamber, resulted in the highest assay signals observed and low nonspecific adsorption.
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