Abstract
A sequential injection analysis system (SIA) using a small column of immobilized concanavalin A (Con A) reacting with horseradish peroxidase (HRP) as a model system was developed to demonstrate the feasibility of flow binding assay for bioprocess monitoring and control. The selection of liquid drive and immobilized supports is justified. System performance was evaluated on three assay formats: a direct binding assay, competitive binding assay, and sandwich-binding assay. In direct binding assay, immobilized Con A mounted in the SIA system was used to determine the concentration of HRP. The competitive binding assay using a free carbohydrate, methyl-D-mannopyranoside to provide competitive binding situation gives more complex calibration curves and decreased precision at higher concentrations. The sandwich-binding assay did not work well. Direct binding assay is recommended to be the best choice based on sensitivity and precision viewpoints. The total time for one assay cycle ranges from 5 ∼ 8 min, depending on the conditions. The current SIA system is fully computerized and yields rapid and reliable system performance. The potential of using SIA-based flow binding assay on the area of bioprocess monitoring is promising.
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