Isolation and Characterization of A Panel of Anti-HER2 Scfv Molecules by Phage Display Technology

Document Type : Original Article

Authors

1 Ph.D. Student of Genetics, College of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran. Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran. Visiting Researcher at Department of Medical Oncology, Fox Chase Cancer Center, Philadelphia, Pennsylvania, USA.

2 Professor, Department of Genetics, College of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran. Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran.

3 Associate Professor, Department of Genetics,College of Agriculture, Ferdowsi University of Mashhad, Mashhad, Iran. Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran.

4 Associate Professor, Department of Molecular Cell Biology, College of Applied Sciences, Ferdowsi University of Mashhad, Mashhad, Iran. Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran.

5 Associate Professor, Department of Physiology, College of Veterinary Medicine, Ferdowsi University of Mashhad, Mashhad, Iran. Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran.

6 Associate Professor, Department of Medical Oncology,Fox Chase Cancer Center, Philadelphia, Pennsylvania, USA.

7 Professor, Department of Medical Oncology, Fox Chase Cancer Center, Philadelphia, Pennsylvania, USA.

Abstract

Introduction: The scFvs (single-chain fragment variable) are popular recombinant antibody formats that are produced by advanced molecular biology and genetic engineering techniques. Moreover, HER2-ECD recognized as a suitable target for a wide range of cancer cells. The current study was accomplished for isolation of specific scFv antibodies.
Methods: To isolate specific scFv fragments from phage library, phage display technology was carried out by coating HER2 extra cellular domain (HER2-ECD) protein onto an immuno tube. The resultant scFv antibodies were expressed in Escherichia coli TG1. The recombinant products were purified by metal affinity chromatography against His-tag. These scFvs were characterized for binding to HER2-ECD by western blot, biosensor surface resonance and flow cytometry assays.
Results: After three rounds of panning against HER2-ECD protein yielded three different scFv fragments that confirmed by subsequent DNA sequencing. The kinetic results indicated that all of isolated scFvs had binding tendency to HER2-ECD on micro molar to nano molar range. Western blot and flow cytometry results demonstrated that all of isolated scFvs have had binding tendency to HER2-ECD.
Conclusion: Phage display is a powerful tool for isolate promising scFv clones that bind specifically to the HER2 receptor, a cell surface of breast cancer antigen.

Keywords

References

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