High-Throughput Kinetic Characterization of Ricin Toxin B Chain and Ovalbumin Antibodies Using Surface Plasmon Resonance.

摘要

Hybridoma cell lines were developed against ricin toxin B (RTB) chain and ovalbumin. These cells were then grown in culture and purified for characterization and possible placement in the Critical Reagents Program repository. From the cell culture medium, antibodies were purified using protein A purification, desalted in 1 phosphate buffered saline using Sephadex G-25 columns, and then filtered with a 0.2 m filter. Twenty-seven purified clones were characterized against their respective immunogens using a Bio-Rad ProteOn XPR36 high-throughput surface plasmon resonance instrument. RTB clones were also characterized against ricin holotoxin. Using an indirect assay format, we estimated kinetic data and toxin reactivity in 14 h. Data were collected and analyzed using Scrubber Pro software. Two clones were found to bind ricin holotoxin in addition to the RTB. Kinetic constants for the ovalbumin clones were also obtained. In the future, this highthroughput screening method can be applied to other purified antibody clones to rank their utility in downstream field assay applications.