title: A quantitative bifunctional in vitro potency assay for botulinum neurotoxin serotype A.
abstract: BACKGROUND
  Botulinum neurotoxin type A (BoNTA) is one of seven serotypes produced by Clostridium botulinum (types A thru G) and is the serotype most widely used to treat both cosmetic and medical conditions. Potency for botulinum toxin preparations is expressed in mouse LD50 units. There is a need to develop a non-animal based replacement for this potency assay.
  
  
  METHODS
  An in vitro potency assay measuring BoNTA activity has been developed that addresses both BoNTA heavy chain binding to its cell receptor SV2C and BoNTA light chain enzymatic activity in cleaving SNAP-25, an intracellular protein essential in neurotransmitter release. This bifunctional assay utilizes a 96 well microtiter format and well defined reagents. Assay characterization determined that the relative standard deviation for intermediate precision was less than 10%.
  
  
  RESULTS
  The assay standard curve covers the range of BoNTA concentrations from 0.0624 to 32 ng/mL. Specificity was demonstrated with purified BoNTA heavy chain which inhibited the activity in a dose dependent manner. A correlation between this bifunctional assay and the mouse LD50 potency assay was demonstrated.
authors: Revance Therapeutics, 7444 Gateway Blvd., Newark, CA 94560, United States.
authors: Richard W
authors: RW
authors: Gregory
authors: Revance Therapeutics, 7444 Gateway Blvd., Newark, CA 94560, United States. Electronic address: bwerner@revance.com.
authors: William E
authors: WE
authors: Werner
authors: Revance Therapeutics, 7444 Gateway Blvd., Newark, CA 94560, United States.
authors: Curtis
authors: C
authors: Ruegg
mesh_terms: D000818:Animals; D019274:Botulinum Toxins, Type A / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism* / Q000633:toxicity; D057075:Enzyme Assays / Q000379:methods*; D007928:Lethal Dose 50; D051379:Mice; D013379:Substrate Specificity
chemical_list: D019274:Botulinum Toxins, Type A
keywords: Botulinum neurotoxin type A (BoNTA) in vitro potency assay; LD(50); Methods; SV2C