The average length of rabbit CDR-H3 is 12 amino acids, which is similar to its human and mouse counterparts, while the CDR-元 length is much longer than its human and mouse counterparts. In addition, the somatic mutations in VH and VL are higher than its human and mouse counterparts, with VH regions accumulating two-third more mutations than humans and mice and VL regions accumulating more mutations in fragment region (FR)-1 and FR-3. In this study, IGHV1S40 and IGHV1S45 were found to dominate the VH repertoire of naïve rabbit, while IGHV1S69 contributed significantly to immunized rabbit with a 16-mer peptide ( 22). Recently, the comprehensive rabbit antibody repertoire was analyzed with next-generation sequencing (NGS) technology ( 22). In addition, the rabbit VL repertoire displays a larger complementarity-determining region (CDR)-元 loop length than its human and mouse counterparts ( 22). Most rabbit antibodies are derived from the IGHV1 gene ( 21). The diversity of rabbit VH repertoire is more limited, and the V light (VL) repertoire is more diverse than that in mice and humans ( 17– 20). Also, more than 50 IG Kappa V and 17 IG Lambda V functional genes were identified with genomic sequencing (IMGT database). The rabbit immunoglobulin heavy chain (IGH) locus contains over 200 IGH variable germline genes, with over 50% have been found to be “non-functional” ( 16, 17). These technology platforms have proven to be efficient and robust for generation of large panels of antigen-specific recombinant antibodies from immunized rabbits within 1 week, which promoted the production of rabbit mAbs. Recently, some new platforms have been developed for generating rabbit mAbs from antigen-specific memory B cells or plasma cells by FACS or manual micromanipulation, and expression of recombinant mAbs in prokaryotic, eukaryotic, or cell-free expression systems ( 9, 12– 15). Owing to these favorable features, more than 9,519 rabbit mAbs were generated by Abcam, which were identified for major signaling pathways including apoptosis, cell cycle, epidermal growth factor receptor signaling, and transforming growth factor-β signaling. Compared with traditional mouse mAbs, the rabbit mAbs have advantages in diagnostics with high affinity and specificity toward antigens, more diverse epitope recognition, and greatly improved response to small-size epitopes and mouse antigens ( 11). The rabbit immune system generates antibody diversity and optimizes affinity by mechanisms different from those of mice and other rodents ( 10). The methods for single-cell isolation are currently reliant on fluorescence-activated cell sorting (FACS) and manual micromanipulation, and the antibody genes were transferred to mammalian cells for mAb expression and further characterization ( 8, 9). Antigen-specific memory B cells expressing surface IgG and IgG-secreting plasma cells have been exploited extensively as a source of mAbs. Moreover, this technique exploits the natural process of affinity, specificity, and stability profiles of mAbs. This technology avoids the inefficient hybridoma fusion step and retains the natural heavy and light chain pairing, thereby allowing a more thorough interrogation of the B cell population. More recently, a single B cell-based method has been developed that allows direct sampling of the immune repertoire from a single B-cell or the clonally expanded progeny of single cell ( 7). In addition, the natural cognate pairing information of antibodies is lost in display methods, which reduced the specific diversity of antibodies. Although these antibody generation technologies were widely adopted for mAbs screening, these methods were inefficient and required time-consuming operations. In the past few decades, several display techniques such as phage display ( 3), yeast surface display, ribosome display ( 4, 5), and mRNA display technologies ( 6) have been used for producing mAbs. The mouse hybridoma method described by Kohler and Milstein in 1975 was the first and most widely used approach for obtaining mouse mAbs ( 2). Many methods exist for the generation and identification of mAbs for both research and therapeutic purposes. mAb therapeutics has revolutionized the approach to many serious human diseases with an increasing speed, and over 230 mAbs were evaluated in phase clinical studies in early 2017 ( 1). Monoclonal antibodies (mAbs) are essential tools in biochemistry, molecular biology, and medicine research.
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