A Highly Specific Monoclonal Antibody for Botulinum Neurotoxin Type A-Cleaved SNAP25

Botulinum neurotoxin type-A (BoNT/A), as onabotulinumtoxinA, is approved globally for 11 major therapeutic and cosmetic indications. While the mechanism of action for BoNT/A at the presynaptic nerve terminal has been established, questions remain regarding intracellular trafficking patterns and overall fate of the toxin. Resolving these questions partly depends on the ability to detect BoNT/A's location, distribution, and movement within a cell. Due to BoNT/A's high potency and extremely low concentrations within neurons, an alternative approach has been employed. This involves utilizing specific antibodies against the BoNT/A-cleaved SNAP25 substrate (SNAP25197) to track the enzymatic activity of toxin within cells. Using our highly specific mouse monoclonal antibody (mAb) against SNAP25197, we generated human and murine recombinant versions (rMAb) using specific backbone immunoglobulins. In this study, we validated the specificity of our anti-SNAP25197 rMAbs in several different assays and performed side-by-side comparisons to commercially-available and in-house antibodies against SNAP25. Our rMAbs were highly specific for SNAP25197 in all assays and on several different BoNT/A-treated tissues, showing no cross-reactivity with full-length SNAP25. This was not the case with other reportedly SNAP25197-selective antibodies, which were selective in some, but not all assays. The rMAbs described herein represent effective new tools for detecting BoNT/A activity within cells.