High Level Expression Of A Functional Humanized Single-chain Variable Fragment (scFv) Antibody Against CD22 Marker In Pichia Pastoris | 49596
Journal of Biotechnology & Biomaterials
Like us on:
Our Group organises 3000+ Global Conferenceseries Events every year across USA, Europe & Asia with support from 1000 more scientific Societies and Publishes 700+ Open Access Journals which contains over 50000 eminent personalities, reputed scientists as editorial board members.
ScFvs have recently emerged as attractive candidates in targeted immunotherapy of various malignancies. The anti-CD22
scFv is able to target CD22, on B-cell surface and is being considered as a promising molecule in targeted immunotherapy
of B-cell malignancies. The recombinant anti-CD22 scFv has been successfully expressed in Escherichia coli; however, the
insufficient production yield has been a major bottleneck for its therapeutic application. Pichia pastoris has become a highly
popular expression host for the production of antibody fragments. In this study we used the Pichia expression system to
express a model scFv against CD22. The full-length humanized scFv gene was introduced into the expression vector pPICZαA
under the control of the AOX1 promoter and expressed in GS115 strain. The maximum production level of the scFv were
achieved at methanol concentration, 1%; pH, 6.0; inoculums density, OD600=3 and the induction time of 72 hours. Using
affinity chromatography, 72% of the recombinant protein was recovered with a purity and final yield of about 93% and 25
mg/L, respectively. The correlation between scFv gene dosage and expression level was also investigated by real time PCR and
the results confirmed the presence of such correlation up to 5 gene copies. Flowcytometry and biacore analysis demonstrated
binding to CD22 on the surface of human lymphoid cell line Raji and recombinant CD22, respectively. Taken together, the
presented data suggest that the Pichia pastoris can be considered as an efficient host for the large-scale production of anti-CD22
scFv as a possible promising carrier for targeted drug delivery.
Najmeh Zarei has completed her PharmD degree and PhD in the field of Pharmaceutical Biotechnology from Pasteur Institute of Iran. She is currently working as an Assistant Professor of the Pasteur Institute of Iran and has published more than 10 papers in reputed journals.