title: Effect of adjuvants on antibody titer of synthetic recombinant light chain of botulinum neurotoxin type B and its diagnostic potential for botulism.
abstract: Botulism is a neuroparalytic disease caused by Clostridium botulinum, which produces seven (A-G) antigenically diverse neurotoxins (BoNTs). BoNTs are the most poisonous substances known to humans, with a median lethal dose (LD₅₀) of approximately 1 ng/kg of body weight. Owing to their extreme potency and lethality, they have the potential to be used as a bioterrorism agent. The mouse bioassay is the gold standard for the detection of botulinum neurotoxins; however, it requires at least 3-4 days for completion. Attempts have been made to develop an ELISA-based detection system, which is potentially an easier and more rapid method of botulinum neurotoxin detection. The present study was designed using a synthetic gene approach. The synthetic gene encoding the catalytic domain of BoNT serotype B from amino acids 1-450 was constructed with PCR overlapping primers (BoNT/B LC), cloned in a pQE30 UA vector, and expressed in an E. coli M15 host system. Recombinant protein production was optimized at 0.5 mM IPTG final concentration, 4 h post induction, resulting in a maximum yield of recombinant proteins. The immunogenic nature of the recombinant BoNT/B LC protein was evaluated by ELISA. Antibodies were raised in BALB/c mice using various adjuvants. A significant rise in antibody titer (p<0.05) was observed in the Alum group, followed by the Titermax Classic group, Freund's adjuvant, and the Titermax Gold group. These developed high-titer antibodies may prove useful for the detection of botulinum neurotoxins in food and clinical samples.
authors.affiliation: Biotechnology Division, Defence Research and Development Establishment, Jhansi Road, Gwalior 474 002, Madhya Pradesh, India.
authors.forename: Swati
authors.initials: S
authors.lastname: Jain
authors.forename: S
authors.initials: S
authors.lastname: Ponmariappan
authors.forename: Om
authors.initials: O
authors.lastname: Kumar
authors.forename: Lokendra
authors.initials: L
authors.lastname: Singh
mesh_terms: D000276:Adjuvants, Immunologic / Q000008:administration & dosage*; D000818:Animals; D000992:Antitoxins / Q000097:blood*; D001905:Botulinum Toxins / Q000008:administration & dosage / Q000032:analysis* / Q000276:immunology; D019274:Botulinum Toxins, Type A; D001906:Botulism / Q000175:diagnosis; D019411:Clinical Laboratory Techniques / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005504:Food Analysis / Q000379:methods; D051379:Mice; D008807:Mice, Inbred BALB C; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000276:immunology
chemical_list: D000276:Adjuvants, Immunologic; D000992:Antitoxins; D011994:Recombinant Proteins; C096323:rimabotulinumtoxinB; D001905:Botulinum Toxins; D019274:Botulinum Toxins, Type A