The sponsor was not involved in the study’s conductance. The authors would like to thank Staffan Paulie for the critical reading of the manuscript. “
“Monoclonal antibodies are a significant and growing class of therapeutics for a wide range of indications including cancer, metabolic, and inflammatory diseases. Phage display antibody libraries are an important tool for the discovery of human monoclonal antibodies, providing two marketed products, one under review by the FDA, and many more at various stages of clinical trials (Nelson et al., 2010). Specificity and affinity are key components
for the successful transition Akt cancer of an antibody from the lab to the clinic. Library size and diversity are extremely important in this endeavor as the larger and more diverse a library, the greater the chance of finding high affinity antibodies with diverse paratopes that bind diverse epitopes (Perelson and Oster, 1979, Perelson, 1989, Griffiths et al., 1994 and Vaughan OSI-906 price et al., 1996). The first fully human phage displayed antibody fragment library had 107 members and antibody fragments to four proteins were isolated with affinities as low as 86 nM (Marks et al., 1991). Other groups went on to construct larger human libraries: two Fab (6.5 × 1010 and 3.7 × 1010) (Griffiths et al., 1994 and de Haard et al., 1999) and one scFv (1.4 × 1010) (Vaughan et al., 1996). From
each library, antibody fragments with single-digit nanomolar affinities were isolated, and from the scFv library, two fragments were isolated with affinities
less than 1 nM. However, Fabs with only moderate affinities (> 800 nM) were recovered when selecting from a small portion of the Griffiths library (107 clones), supporting the claim that the larger the library, the greater the probability of isolating high affinity antibodies (Griffiths et al., 1994). To this end, we constructed two phagemid libraries, XFab1 and XscFv2, which display Fab and scFv fragments, respectively, each with more than 2.5 × 1011 members maximizing the potential for isolating high affinity antibodies against any target of interest. Antibody diversity Methane monooxygenase is influenced by the number of donors, donor tissues used, the types of variable regions from which antibody sequences are amplified and the choice in the utilization of V-gene frameworks. For each of XFab1 and XscFv2, variable regions were amplified from thirty racially-diverse healthy donors using a variety of tissues including bone marrow, PBMCs, spleen and lymph node. The amplification strategy encompasses variable domains derived from IgM, IgG, IgA, IgE and IgD. While other commercial phage display antibody libraries have restricted antibody frameworks to enhance stability or expression of the displayed fragments (Söderlind et al., 2000, Hoet et al., 2005 and Rothe et al., 2008), in the XFab1 and XscFv2 libraries, all prominent V-gene families encompassing the human repertoire were utilized to allow increased structural diversity.