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Although nowadays many different libraries for the selection of antibodies are available, hybridoma cells and immunized animals are still an important source for the production and selection of specific binders. Therefore, this protocol describes the isolation of the coding mRNA, PCR amplification of VL and VH domains and the final assembly of these domains into the scFv format, fused to an M13 phage...
Today, recombinant protein expression in E. coli is usually considered a routine technology. Nonetheless, problems with soluble expression, rooted in protein aggregation competing with folding, are still frequently encountered. Many antibody fragments from natural sources are aggregation-prone during folding. Antibodies and their derivatives need to have disulfide bonds for functional expression,...
Recombinant antibodies have become a standard component of research, diagnostics, and therapy. In the development of recombinant antibodies – irrespective of the final format – a monovalent construct is virtually always the first protein to be tested. This is due to the fact that essentially all selection systems use such formats, and that the periplasmic production of scFv and Fab fragments has now...
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