Characterization of a Human Antibody Fragment Fab and Its Calcium Phosphate Nanoparticles that Inhibit Rabies Virus Infection with Vaccine

Abstract

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 6.7×108, 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).

A Human Anti-c-Met Fab Fragment Conjugated with Doxorubicin as Targeted Chemotherapy for Hepatocellular Carcinoma

“…Human peripheral blood mononuclear cells were isolated by Ficoll-Pacque gradient centrifugation, and total RNA was prepared by using an RNA Purification kit (QIAGEN, Valencia, CA, USA). For the amplification of Fab gene segments, a unique three-step PCR was used [54]. The resultant Fab was digested with Sfi I (Roche Molecular Biochemicals, Mannheim, Germany), and ligated into the phagemid pComb3XSS (provided by the Barbas III Laboratory).…”

Monoclonal antibodies for prophylactic and therapeutic use against viral infections

“…The antibody variable domains can be engineered into small fragments ( Table 2), including scFvs and F[ab] molecules [76][77][78] which do not require production in costly eukaryotic expression systems. Several of these small antibody fragments have been investigated regarding their antiviral activities [79,80], including camelid VHH domains. The serum of camels, dromedaries and llamas contains a unique type of antibodies devoid of antibody light chains [21].…”

Phage Displayed Peptides to Avian H5N1 Virus Distinguished the Virus from Other Viruses

“…There are reports regarding the phage-displayed peptides selected from combinatorial libraries that interacting with hepatitis B virus, adenovirus type 2, Andes virus, Sin Nombre virus and Hantaan virus and coronavirus [16]. At present, the phage display technology has become an increasingly attractive molecular tool to researchers in biotechnology related fields [17][18][19]. In our study, we used the H5N1 virion as an immobilized target and performed a biopanning using a 12-mer phage display peptide library.…”