Neutralizing human Fab fragments against measles virus recovered by phage display.
Five human recombinant Fab fragments (Fabs) specific for measles virus (MV) proteins were isolated from three antibody phage display libraries generated from RNAs derived from bone marrow or splenic lymphocytes from three MV-immune individuals. All Fabs reacted in an enzyme-linked immunosorbent assay with MV antigens. In radioimmunoprecipitation assays two of the Fabs, MV12 and MT14, precipitated an approximately equal 80-kDa protein band corresponding to the hemagglutinin (H) protein from MV-infected Vero cell cultures, while two other Fabs, MT64 and GL29, precipitated an approximately equal 60-kDa protein corresponding the nucleocapsid (N) protein. In competition studies with MV fusion, H- and N protein-specific monoclonal antibodies (MAbs), the H-specific Fabs predominantly blocked the binding of H-specific MAbs, while the N-specific Fabs blocked MAbs to N. In addition, N-specific Fabs bound to denatured MV N protein in Western blotting. The specificity of the fifth Fab, MV4, could not be determined. By plaque reduction assays, three of the five Fabs, MV4, MV12, and MT14, exhibited neutralizing activity (80% cutoff) against MV (LEC-KI strain) at concentrations ranging between approximately 2 and 7 microg x ml(-1). Neutralization capacity against MV strains Edmonston and Schwarz was also detected, albeit at somewhat higher Fab concentrations. In conclusion, three neutralizing Fabs were isolated, two of them reactive against the H glycoprotein of MV and another reactive against an undefined epitope. This is the first study in which MV-neutralizing human recombinant Fab antibodies have been isolated from phage display libraries.
de Carvalho Nicacio
Cristina
C
Microbiology and Tumor Biology Center, Karolinska Institutet, S-171 77 Stockholm, Sweden.
Williamson
R Anthony
RA
Parren
Paul W H I
PW
Lundkvist
Ake
A
Burton
Dennis R
DR
Björling
Ewa
E
D000595:Amino Acid Sequence; D000914:Antibodies, Viral / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D000918:Antibody Specificity; D001435:Bacteriophages / Q000235:genetics*; D003001:Cloning, Molecular; D015723:Gene Library; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D007147:Immunoglobulin Light Chains / Q000235:genetics; D008214:Lymphocytes / Q000276:immunology; D008457:Measles / Q000276:immunology; D008459:Measles virus / Q000276:immunology*; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D011994:Recombinant Proteins / Q000096:biosynthesis
D000914:Antibodies, Viral; D007140:Immunoglobulin Fab Fragments; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D011994:Recombinant Proteins