Abstract
The immunoglobulin heavy- and light-chain variable region (VH and VK) genes were isolated from 8E5 hybridoma cells, which secreted monoclonal antibody against human fibrin by RT-PCR. An expression vector pOPE51-8E5 was constructed for the recombinant VH-VK scFv expression. The primary sequence of the variable regions was determined. Expression product was found in the periplasmic space and inclusion bodies by SDS-PAGE and immunoblotting. It was a 30 KDa single chain fragment (scFv) with the antigen-binding specificity of the parental monoclonal antibody. A light chain shuffling with an unspecific VL did not result in a loss of fibrin binding specificity.
MeSH terms
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Amino Acid Sequence
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Animals
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Antibodies, Monoclonal / genetics*
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Antibodies, Monoclonal / immunology
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Base Sequence
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Cell Line
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Cloning, Molecular
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Fibrin / immunology*
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Fibrin / metabolism
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Humans
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Immunoglobulin Fragments / genetics
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Immunoglobulin Heavy Chains / genetics*
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Immunoglobulin Heavy Chains / immunology
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Immunoglobulin Light Chains / genetics*
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Immunoglobulin Light Chains / immunology
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Immunoglobulin Variable Region / genetics*
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Immunoglobulin Variable Region / immunology
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Mice
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Molecular Sequence Data
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Recombinant Proteins / genetics
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Recombinant Proteins / metabolism
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Restriction Mapping
Substances
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Antibodies, Monoclonal
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Immunoglobulin Fragments
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Immunoglobulin Heavy Chains
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Immunoglobulin Light Chains
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Immunoglobulin Variable Region
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Recombinant Proteins
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immunoglobulin Fv
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Fibrin
Associated data
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GENBANK/N11725
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GENBANK/Y12778