A recombinant single-chain variable fragment (scFv) antibody was engineered to a tissue-specific carbohydrate epitope located on human sperm agglutination antigen-1 (SAGA-1), a sperm glycoform of CD52, demonstrating that active recombinant antibodies can be produced to a tissue-specific carbohydrate epitope on the human sperm surface, thereby creating opportunities for novel contraceptive agents.
The described materials and methods generated RASA, a recombinant scFv mini-antibody form of the murine anti-sperm mAb S19. Using recombinant DNA techniques, cDNAs encoding the variable regions of the S19 heavy and light chains were isolated from the parental MHS-8 hybridoma, linked to one another, ligated into an expression vector, and expressed in E. coli. Identification of the correct light chain cDNA was initially problematic due to a frame shift in the cDNA generated by the RPAS kit primers and was corrected through the generation of a unique primer. Use of the Van911 restriction enzyme to delete cDNAs encoding the aberrant endogenous light chain was a novel technique that expedited cloning of the authentic scFv. Both techniques may be of use to other researchers generating recombinant antibodies from murine hybridomas with homologous sequences to RASA. Over the past decade, the use of monoclonal antibodies has been expanded to a variety of clinical applications (Hollinger and Hoogenboom, 1998; Hudson, 1998, 1999); however, adverse effects, such as binding interference, caused by using whole murine antibodies in humans, and the expense of tissue culture have presented obstacles to their use. Recombinant antibody technology has presented a solution to both of these impediments, because the generation of recombinant antibody fragments, such as scFvs, not only removes the majority of the immunogenic murine sequence, but may also be more economical. 6 figures and 45 references
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