Summary of VHH characterization data.

AVHH epitopes are named arbitrarily based on their inability to compete with the binding of VHHs recognizing other epitopes.BSubunit recognition was assessed by ELISA with purified BoNT light chain (Lc) or heavy chain (Hc). VHHs recognizing BoNT holotoxin without recognition of purified Lc or Hc are indicated as none. RBD indicates recognition of the 50 kDa carboxyl end receptor binding domain of Hc.CVHH neutralization was determined by the ability of the VHH to prevent intoxication of primary neurons by 10 pM BoNT/A (Figure 1). ‘Strong’ indicates that the presence of ≤0.1 nM VHH led to obvious toxin neutralization in primary neuron assays (see Figure 1). ‘Weak’ indicates detectable toxin neutralization when the medium contained ≤1 nM VHH. None indicates no toxin neutralization was detected when the medium contained ≤10 nM VHH.DSurface plasmon resonance (SPR) studies were performed using chips coated with ciBoNTA for BoNT/A VHHs and ciBoNTB for BoNT/B VHHs as described in Methods and Materials.