Phage displayed antibodies to heat stable alkaline phosphatase: framework region as a determinant of specificity.
Human antibodies against specific targets of tumor cells are the most desirable molecules for possible immunotherapy. They could be developed by using the combinatorial antibody library displayed on a phage. We selected four human antibody fragments (scFv) binding to the oncoplacental antigen Heat Stable Alkaline Phosphatase (HSAP, the placental isozyme of alkaline phosphatase) from a synthetic human antibody library. Characterization of these scFvs showed they bound HSAP with moderate affinity but did not have isozyme specificity, as determined by binding to cell lines exhibiting differential expression of isozymes of alkaline phosphatase. The V(H) sequences of two of these scFvs were similar and although both bound to HSAP only one was cross-reactive with albumin. The sequences revealed a difference in the framework region (FR1) of these antibodies, indicating a role for this region in the determination of specificity. This is also significant considering that the heavy chains generated the diversity of the synthetic library used in this study, and only a single light chain showing binding to BSA was used for the entire library.
D000469:Alkaline Phosphatase / Q000276:immunology*; D000595:Amino Acid Sequence; D000918:Antibody Specificity; D019921:Biotinylation; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006358:Hot Temperature; D006801:Humans; D007128:Immunoglobulin Fragments / Q000276:immunology*; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology*; D007527:Isoenzymes / Q000276:immunology; D008969:Molecular Sequence Data; D019151:Peptide Library; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D012710:Serum Albumin, Bovine / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D007527:Isoenzymes; D019151:Peptide Library; D011994:Recombinant Proteins; D012710:Serum Albumin, Bovine; D000469:Alkaline Phosphatase