title: Development of protective agent against Hottentotta saulcyi venom using camelid single-domain antibody.
abstract: Hottentotta saulcyi, medically important scorpion species, causes some of harmful toxic exposure in Iran. Administrated, conventional antivenom-based immunotherapy is still limited and hardly meet ideal characteristic of effective treatment for scorpion envenomation. In this study we aimed to develop a neutralizing agent directed against scorpion venom based on VHH, variable domain of the Camelidae heavy chain antibody or Nanobody. This promising biomolecule is well-established as an advantageous tool for therapeutic purposes due to its small size, stability, monomeric performance and less immunogenicity. In this study, a large Nb library was constructed and phage displayed after successful camel immunization using H. saulcyi scorpion crude venom. After a series of biopanning rounds on Sephadex G50 purified venom fraction and screening by monoclonal phage ELISA, the best reactive Nb was retrieved and designated Nb12. The selected Nb was then expressed as soluble protein in Escherichia coli, purified and confirmed by SDS-PAGE analysis and western blotting. The lead candidate Nb12 bound scorpion venom with Kaff value of 5×10(7)M(-1). Nb12 was shown to be capable of neutralizing 2 LD50 of whole venom of scorpion toxin when injected in the ratio of the Nb/toxin of 1.4:1 into C57BL/6 mice. In challenge experiment, Nb succeeded to rescue all i.p. lethal dose injected mice even when administrated i.v., 20min after envenoming. These results with ease of production and superior neutralizing activity make Nb a suitable anti-toxin candidate for treatment of scorpion envenoming.
authors.affiliation: Biotechnology Research Center, Venom & Biotherapeutics Molecules Lab, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Maryam
authors.initials: M
authors.lastname: Darvish
authors.affiliation: Biotechnology Research Center, Venom & Biotherapeutics Molecules Lab, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Mahdi
authors.initials: M
authors.lastname: Behdani
authors.affiliation: National Cell Bank of Iran, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Mohammad Ali
authors.initials: MA
authors.lastname: Shokrgozar
authors.affiliation: Biotechnology Research Center, Venom & Biotherapeutics Molecules Lab, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Kamran
authors.initials: K
authors.lastname: Pooshang-Bagheri
authors.affiliation: Biotechnology Research Center, Venom & Biotherapeutics Molecules Lab, Pasteur Institute of Iran, Tehran, Iran. Electronic address: shahbazzadeh@yahoo.com.
authors.forename: Delavar
authors.initials: D
authors.lastname: Shahbazzadeh
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology*; D000915:Antibody Affinity; D000997:Antivenins / Q000276:immunology* / Q000494:pharmacology; D002162:Camelus / Q000276:immunology*; D007114:Immunization; D007143:Immunoglobulin Heavy Chains / Q000276:immunology / Q000494:pharmacology; D007167:Immunotherapy / Q000379:methods; D007492:Iran; D051379:Mice; D008810:Mice, Inbred C57BL; D011485:Protein Binding / Q000276:immunology; D065008:Scorpion Stings / Q000188:drug therapy* / Q000473:pathology; D012604:Scorpion Venoms / Q000037:antagonists & inhibitors* / Q000276:immunology; D012605:Scorpions / Q000378:metabolism; D061905:Single-Domain Antibodies / Q000276:immunology / Q000494:pharmacology*
chemical_list: D057134:Antibodies, Neutralizing; D000997:Antivenins; D007143:Immunoglobulin Heavy Chains; D012604:Scorpion Venoms; D061905:Single-Domain Antibodies
keywords: Antivenom; Heavy chain antibody (HC-Ab); Hottentotta saulcyi; Nanobody; Phage display