A型肉毒毒素中和抗体S25全抗体的表达

目的:用HEK293细胞表达A型肉毒毒素(BoNT/A)中和抗体S25。方法:按哺乳动物偏好密码子设计合成S25可变区编码序列,κ链可变区序列构建到带有轻链恒定区的载体L293,重链可变区构建到带有重链恒定区的载体H293,轻、重链载体共转染至HEK293细胞,进行瞬时表达;用重链C端(HCC)及S25中和表位突变的HCC与细胞培养上清ELISA检测S25表达。结果:测序结果表明正确设计合成了S25可变区编码序列;ELISA结果显示分泌表达上清可以与抗原结合,而不能与S25中和表位突变的HCC结合。结论:按哺乳动物偏好密码子设计的S25编码序列可以用HEK293细胞进行表达,并且具有活性。

Expression of Neutralizing Antibody S25 for Botulinum Neurotoxin Type A

Objective：To express neutralizing antibody S25 of botulinum neurotoxin type A（BoNT/A） in HEK293 cells.Methods：Coding sequence for variable region of S25 was designed according to mammalian cell codon us-age preference.Vκ was constructed into vector L293 with constant region of light chain,while VH was constructed into vector H293 with constant region of heavy chain.Expression vectors for light chain and heavy chain were co-transfected into HEK293 cells,and antibody was transient expressed.S25 in expressing supernatant was detected by ELISA with HCC and its mutant lack of S25 neutralizing epitope.Results：DNA sequencing demonstrated that V region coding sequence was rightly synthesized.ELISA showed that HEK293 supernatant bound with HCC but not with the mutant.Conclusion：S25 with artificial synthesized V region can be secretively expressed by HEK293 asactive form.