Recombinant human antibodies: linkage of an Fab fragment from a combinatorial library to an Fc fragment for expression in mammalian cell culture

Bender, Eckhard; Woof, Jennifer M.; Atkin, Julie D.; Barker, M. D.; Bebbington, Christopher; Burton, Dennis R.

doi:10.3233/hab-1993-4205

Cited by 48 publications

(12 citation statements)

References 25 publications

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“…Phage display of antibody libraries has provided a powerful tool for the isolation of human MAbs to important viral pathogens (4,7,12,16,24,51). Large repertoires of antibodies can be displayed on the surface of filamentous phage particles (e.g., M13), and antibodies with desired specificity can be isolated by panning on the antigens of interest (6,13,61).…”

mentioning

confidence: 99%

Identification by Phage Display and Characterization of Two Neutralizing Chimpanzee Monoclonal Antibodies to the Hepatitis E Virus Capsid Protein

Schofield

Glamann

Emerson³

et al. 2000

J Virol

155

109

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Two monoclonal antibodies (MAbs) against the ORF2 protein of the SAR-55 strain of hepatitis E virus (HEV) were isolated by phage display from a cDNA library of chimpanzee (Pan troglodytes) ␥1/ antibody genes. Both MAbs, HEV#4 and HEV#31, bound to reduced, denatured open reading frame 2 (ORF2) protein in a Western blot, suggesting that they recognize linear epitopes. The affinities (equilibrium dissociation constants, K d ) for the SAR-55 ORF2 protein were 1.7 nM for HEV#4 and 5.4 nM for HEV#31. The two MAbs also reacted in an enzyme-linked immunosorbent assay with recombinant ORF2 protein from a heterologous HEV, the Meng strain. Each MAb blocked the subsequent binding of the other MAb to homologous ORF2 protein in indirect competition assays, suggesting that they recognize the same or overlapping epitopes. Radioimmunoprecipitation assays suggested that at least part of the linear epitope(s) recognized by the two MAbs is located between amino acids 578 and 607. MAbs were mixed with homologous HEV in vitro and then inoculated into rhesus monkeys (Macaca mulatta) to determine their neutralizing ability. Whereas all control animals developed hepatitis (elevated liver enzyme levels in serum) and seroconverted to HEV, those receiving an inoculum incubated with either HEV#4 or HEV#31 were not infected. Therefore, each MAb neutralized the SAR-55 strain of HEV in vitro.Hepatitis E is an acute disease endemic in many countries throughout developing parts of the world, in particular on the continents of Africa and Asia, where epidemics have also occurred. The causative agent, hepatitis E virus (HEV), is transmitted via the fecal-oral route, predominantly through contaminated water (45). HEV is an RNA virus with a positive-sense genome approximately 7.5 kb in length. The genome contains three open reading frames (ORFs); ORF2 encodes the putative capsid protein (45). Hepatitis E has a low mortality rate in young adults (38). However, mortality rates can reach 20% in women in the third trimester of pregnancy (32, 58). Surprisingly, in industrialized countries such as the United States, where hepatitis E is not endemic, a significant proportion of healthy individuals within the general population are seropositive (over 20% in some areas; 39, 50). However, clinical hepatitis E is rare in these countries and usually is seen in individuals who acquired the infection during travel to a region in which HEV is endemic or epidemic.It has been suggested that animals serve as reservoirs for HEV and that human infections may, in part, be zoonoses. There have been several reports of HEV-specific (anti-HEV) antibody in animals (1, 9, 28, 29). Furthermore, an HEV-like virus was recently isolated from naturally infected swine in the United States (40). Although four genotypes of HEV have been identified, to date, only one serotype of HEV has been recognized. Therefore, it may be possible to produce a broadly protective vaccine. Passively transferred anti-HEV serum significantly reduced virus shedding in feces and abrogated disease in nonhu...

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mentioning

confidence: 99%

Identification by Phage Display and Characterization of Two Neutralizing Chimpanzee Monoclonal Antibodies to the Hepatitis E Virus Capsid Protein

Schofield

Glamann

Emerson³

et al. 2000

J Virol

155

109

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“…We also demonstrated that rFab fragment preincubated with anti-F(ab′) 2 showed the cell growth-inhibitory activity. This preliminary experiment suggests that the use of HBJ127 rFab genetically fused with the human IgG Fc casette 35) for cancer immunotherapy might be feasible. For clinical application of mAb, human antibodies against tumor antigens would be the best, though it is very difficult to isolate them because of the low immunogenicity of tumor antigens in humans.…”

Section: Discussionmentioning

confidence: 92%

Phage Display Cloning and Characterization of Monoclonal Antibody Genes and Recombinant Fab Fragment against the CD98 Oncoprotein

Itoh

Inoue

Hirooka

et al. 2001

Japanese Journal of Cancer Research

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The Fab gene of anti-CD98 heavy chain (h.c.) monoclonal antibody (mAb) HBJ127 was cloned and expressed as a recombinant Fab (rFab) fragment by means of a phage display system. The variable heavy and light chain genes of HBJ127 were found to be derived from VOx-1 and IgVk8-30 germline, respectively. Extensive somatic mutation was found in the heavy chain complementarity determining region 2. rFab fragment was purified homogeneously from crude bacterial lysates by Nichelate chromatography in a yield of 71.4 µ µ µ µg from 100 ml of culture. , respectively. Three-fold lower affinity of rFab fragment may be due to the difference of valency of the antibody preparation. Cell growth inhibition in vitro by rFab fragment preincubated with anti-Fab suggests that the rFab fragment produced by cloned gene-bearing Escherichia coli was identical to the Fab part of HBJ127 mAb. These results show that a small fragment with antigen binding activity similar to that of the parent mAb can easily be prepared by using a phage display system. To our knowledge, this is a first report of the production of anti-CD98 h.c. rFab fragment.

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“…PEGylation (Chapman, 2002), conjugation and fusion to an Fc fragment of an antibody (Smith et al, 2001) successfully increase the serum half-life of nanobodies. Fusion of nanobodies with proteins such as albumin can provide multifunctional proteins with several binding sites (Bender et al, 1993;Harmsen et al, 2005). The peculiar nature of nanobodies endows them more ability and they can be shared of immune-constructs.…”

Section: Therapeutic Applications Of Nanobodymentioning

confidence: 99%

Nanobody, New Agent for Combating Against Breast Cancer Cells

Rahbarizadeh¹,

Jamnani²,

Iri-Sofla³

2011

Breast Cancer - Current and Alternative Therapeutic Modalities

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Recombinant human antibodies: linkage of an Fab fragment from a combinatorial library to an Fc fragment for expression in mammalian cell culture

Cited by 48 publications

References 25 publications

Identification by Phage Display and Characterization of Two Neutralizing Chimpanzee Monoclonal Antibodies to the Hepatitis E Virus Capsid Protein

Identification by Phage Display and Characterization of Two Neutralizing Chimpanzee Monoclonal Antibodies to the Hepatitis E Virus Capsid Protein

Phage Display Cloning and Characterization of Monoclonal Antibody Genes and Recombinant Fab Fragment against the CD98 Oncoprotein

Nanobody, New Agent for Combating Against Breast Cancer Cells

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