Characterization of a human antibody fragment Fab and its calcium phosphate nanoparticles that inhibit rabies virus infection with vaccine

• Published in: PloS one 2011-05, Vol.6 (5), p.e19848
• Main Authors: Liu, Xinjian, Lin, Hong, Tang, Qi, Li, Chen, Yang, Songtao, Wang, Zhongcan, Wang, Changjun, He, Qing, Cao, Brian, Feng, Zhenqing, Guan, Xiaohong, Zhu, Jin
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Analysis; Animals; Antibodies; Antibodies, Viral - chemistry; Antibodies, Viral - immunology; Antibody libraries; Antigens - immunology; Biology; Blotting, Western; Calcium; Calcium phosphate; Calcium phosphates; Calcium Phosphates - immunology; Cell Line; Deoxyribonucleic acid; Display devices; DNA; DNA sequencing; Drug development; Drugs; Electrophoresis, Polyacrylamide Gel; Enzyme-Linked Immunosorbent Assay; Fab; Fluorescence; Fluorescent Antibody Technique; Gene sequencing; Glycoproteins; Health aspects; Humans; Immobilized Proteins - immunology; Immune system; Immunization; Immunofluorescence; Immunoglobulin Fab Fragments - chemistry; Immunoglobulin Fab Fragments - immunology; Immunoglobulins; Immunology; Immunoprecipitation; Immunosuppressive agents; In vivo methods and tests; Infectious diseases; Kaplan-Meier Estimate; Laboratories; Libraries; Lymphocytes; Lyssavirus; Mass Spectrometry; Mass spectroscopy; Materials Science; Medicine; Mice; Molecular Sequence Data; Nanoparticles; Nanoparticles - therapeutic use; Neutralization; Neutralization Tests; Nucleotide sequence; Panning; Peptide Library; Peripheral blood; Permeability; Phage display; Phages; Phosphates; Prophylaxis; Proteins; Rabies; Rabies - immunology; Rabies - prevention & control; Rabies - virology; Rabies Vaccines - immunology; Rabies virus - immunology; Vaccination; Vaccines; Virions; Viruses
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0019848

Recombinant antibody phage display technology has been used to mimic many aspects of the processes that govern the generation and selection of high-affinity natural human antibodies in the human immune system, especially for infectious disease prophylaxis. An anti-rabies virus immunized phage-display Fab library was constructed from peripheral blood lymphocytes from vaccinated volunteers. The immunized antibody library, with a diversity of 6.7×10(8), was used to select and produce antibodies that bound to rabies virus glycoprotein. After five rounds of immobilized fixed rabies virion panning, four unique DNA sequences were found in the higher binding clones, and only one, Fab094, showed neutralization activity. Fab094 components were analyzed by ELISA, immunoprecipitation and immunofluorescent staining. ELISA and immunofluorescence showed that Fab094 bound specifically to rabies virions. Immunoprecipitation and mass spectrometry showed that Fab094 reacted with rabies virus glycoprotein. To improve the penetration power of Fab094 antibodies, we developed Fab094 calcium phosphate nanoparticles (Fab094-CPNPs) and tested their efficacy. The rapid fluorescent focus inhibition test indicated that the neutralizing antibody titers of Fab094 and Fab094-CPNPs were reached at 200.17 IU/Kg and 246.12 IU/Kg, respectively. These findings were confirmed in vivo in a Kunming mouse challenge model. Our results demonstrate that human Fab094 and Fab094-CPNPs are efficacious candidate drugs to replace rabies immunoglobulin in post-exposure prophylaxis (PEP).