Selection and characterization of alpha-cobrotoxin specific llama single-domain antibodies

Stewart, Christine S.
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University of Guelph

Snake bite is a serious global medical concern for which the only effective treatment is the administration of specific antivenom. Conventional antivenoms, however, often elicit adverse reactions and have mismatched pharmacokinetic profiles with their target toxins due to antibody/toxin size discrepancies. VHHs, the smallest antigen binding fragment, are highly diffusible, stable under physiological conditions, non-immunogenic, and are easily produced ' in vitro' using bacterial and yeast expression systems. Three [alpha]-cobrotoxin specific VHHs were isolated from a nai?ve llama phage display library. [alpha]-Cobrotoxin binding specificity was determined via phage-displayed VHH ELISA. Surface plasmon resonance analysis determined the K D values to be 2, 3 and 3 [mu]M; however, these affinities were too low for [alpha]-cobrotoxin detection by soluble VHH ELISA. A pentamerization strategy, linking the VHH to the verotoxin 1 B subunit, improved the VHH functional affinities for ELISA development. VHH-based antivenoms may improve antivenom biodistribution and neutralization while also reducing adverse effects.

alpha-cobrotoxin, llama single-domain antibodies, VHH-based antivenoms, isolation, detection, VHH ELISA