A subtractive fluorescence-activated cell-sorting strategy to identify mimotopes of HBV–preS protein from bacterially displayed peptide library

• Published in: Journal of immunological methods Vol. 293; no. 1; pp. 13 - 21
• Main Authors: Xin, Zhong-Tao, Liu, Chuan, Dong, Bo, Gao, Ya-Ping, Shao, Ning-Sheng, Liu, Wei, Zhang, Jie, Dong, Jie, Ling, Shi-Gan, Xue, Yan-Ning
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.10.2004; Elsevier
• Subjects: Animals; Bacteria display; Biological and medical sciences; Blotting, Western; Cell Separation - methods; Epitope screening; FACS; Female; Flow Cytometry - methods; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Hepatitis B virus; Hepatitis B virus - immunology; Mice; Microbiology; Molecular immunology; Peptide Library; Protein Precursors - immunology; Rabbits; Techniques; Techniques used in virology; Viral Envelope Proteins - immunology; Virology; MFI; Peptide library; HRP; aFGF; Epitope screening; FACS; HBV; Bacteria display; Antigenic determinant; Peptides; Orthohepadnavirus; Identification; Protein; Immunological method; Virus; Hepadnaviridae; Bacteria; Hepatitis B virus
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/j.jim.2004.06.006

A novel subtractive fluorescence-activated cell-sorting (FACS) strategy using a model system is described here to identify disease-specific (DS) epitopes from a bacterially displayed random peptide library. In this process, preimmune serum was used as “Driver ” to block any common binding sites on the bacterial surface and the labeled anti-preS IgG polyclonal antibodies from immunized serum were used as “Tester” to enrich preS-specific mimotopes. Bacterial clones were identified out of this pool through an “antigen-independent” procedure only using both different sera samples. After four rounds of sub-FACS screening, 41 out of 50 bacterial clones were identified as reacting with the immunized serum but not reacting with the pre-immune one. Two motif sequences HQLD and DPAF were obtained from 13 clones. Immunization of mice with two representative bacterial clones elicited a strong specific response against native preS antigen in comparison with the control. This technique may provide a useful technology platform for high-throughput screening of disease-related epitope which is of importance to develop vaccine against some infectious diseases whose pathogen or immunodominant antigen is still unknown.