Transitioning from the bench to the real world: Adaptation of uranyl specific antibodies for use in field-based environmental analysis
Description
The ability to obtain quantifiable data about a specific environmental contaminant while at a field-based research site has been a need for many researchers working outside the laboratory. This report describes a progression of studies aimed to transition a traditional laboratory-based immunoassay for uranium into a portable field-based format that operates on the principle of kinetic exclusion and is capable of uranium measurement in near real-time. Non-competitive kinetic exclusion-based immunoassays were developed for low molecular weight haptens using a variety of monoclonal antibodies. These non-competitive assays were compared to tradition competitive ELISA's using the same antibodies and hapten-protein conjugates. In most cases the competitive ELISA's showed less sensitivity than the respective non-competitive immunoassay. A trend was observed correlating the relative insensitivity of competitive ELISA to the degree of difference in equilibrium dissociation constants between binding to the free hapten and hapten-protein conjugates. These studies led to the adaptation of a kinetic exclusion-based immunoassay for uranium to a prototype Field Portable Sensor and InLine Immunosensor for use at field site during an ongoing large-scale bioremediation experiment. Samples of groundwater were taken, processed, and measured on-site. Aliquots of the same samples were measured off site using a different technique for uranium determination. Our measurements showed a high degree of agreement with this laboratory-based assay. While these field-based immunoassays worked well there was still room for improvement, particularly regarding the antibody that was used. Final experiments in this study were directed toward engineering a uranium antibody with better binding characteristics. The variable regions of 12F6 were amplified and used to construct a single-chain format of this antibody, which could subsequently be used for phage display experiments. While this project focused on the analysis of uranium in environmental samples, the studies detailed here show a proof of concept towards developing a field-based immunoassay for any low molecular weight contaminant for which an antibody is available