Recombinant botulinum neurotoxin serotype A binding domain [BoNT/A(Hc)], expressed in @entity10056 , was developed as a vaccine candidate for preventing botulinum neurotoxin type A (BoNT/A) intoxication. After fermentation and cell disruption, BoNT/A(Hc) was purified by using a three-step chromatographic process consisting of expanded-bed chromatography, Mono S cation-exchange chromatography, and hydrophobic interaction chromatography. Two pools of immunogenic product were separated on the Mono S column and processed individually. Both products were more than 95% pure and indistinguishable by @entity3208 - @entity2975 gel electrophoresis, Western blot analysis, and enzyme-linked immunosorbent assay (ELISA). Each protein was assayed for potency in @entity19 at immunogen doses ranging from 2.4 ng to 10 microg, followed by challenge with 1,000 @entity19 intraperitoneal 50% lethal doses (i.p. LD50) of BoNT/A. The calculated 50% effective dose for both peaks was approximately 0.1 microg/ @entity19 . @entity20690 was evaluated further in a @entity19 efficacy assay. @entity19 were injected either once, twice, or three times at five different doses and subsequently challenged with 100,000 @entity19 i.p. LD50 of BoNT/A. In general, multiple injections protected better than one, with complete or nearly complete protection realized at doses of >/=0.5 microg/mouse. Serum neutralization and ELISA titers were also determined. Tellingly, 82 of 83 mice with antibody titers of >/=1, 600, as measured by ELISA, survived, but only 6 of 42 @entity19 with titers of </=100 survived. This work shows that the purified BoNT/A(Hc) produced was a highly effective immunogen, able to protect against a high challenge dose of neurotoxin.

Purification, potency, and efficacy of the botulinum neurotoxin type A binding domain from XXXX as a recombinant vaccine candidate.

@entity3208 :: ('MESH:D012967', 'Chemical') :: ['sodium dodecyl sulfate'] @entity10056 :: ('4922', 'Species') :: ['Pichia pastoris'] @entity19 :: ('10090', 'Species') :: ['mice', 'mouse', 'Mice'] @entity20690 :: ('244895(Tax:10090)', 'Gene') :: ['Peak 1'] @entity2975 :: ('CHEBI:51135', 'Chemical') :: ['polyacrylamide']
@entity10056 :: (4922,Species) :: ['Pichia pastoris']
