Binding Kinetics of Antiricin Single Domain Antibodies and Improved Detection Using a B Chain Specific Binder

摘要

Single domain antibodies are the recombinantly expressednbinding fragments derived from heavy chain antibodiesnfound in camels and llamas. These unique binding elementsnoffer many desirable properties such as their smallnsize (∼15 kDa) and thermal stability, which makes themnattractive alternatives to conventional monoclonal antibodies.nWe created a phage display library from llamasnimmunized with ricin toxoid and selected a number ofnsingle domain antibodies. Phage selected on ricin werenfound to bind to either ricin A chain or the intact molecule;nno ricin B chain binders were identified. By panning onnB chain, we identified binders and have characterizedntheir binding to the ricin B chain. While they have a poorernaffinity than the previously described A chain binders, itnwas found that they performed dramatically better asncapture reagents for the detection of ricin, providing anlimit of detection in enzyme linked immunosorbent assayn(ELISA) below 100 pg/mL and excellent specificity fornricin versus the highly related RCA 120 (1 to 10 000).nWe also reevaluated the previously isolated antiricin singlendomain antibody binding kinetics using surface plasmonnresonance and found their Kds matched closely to thosenpreviously obtained under equilibrium binding conditionsnmeasured using the Luminex flow cytometer.