Method for preparing monoclonal antibody
View Patent ↗A significantly increased amount of a monoclonal antibody is obtained from the culture medium of recombinant hybridoma prepared by introducing genes encoding a protein identical to the immunoglobulin heavy chain polypeptide of the specific monoclonal antibody into an immortalized B cell (hybridoma) producing the monoclonal antibody.
1. A method of producing a monoclonal antibody, comprising the steps of:
fusing a B cell of a transgenic mouse which expresses human antibodies with a myeloma cell line incapable of producing antibodies to obtain a hybridoma, wherein the hybridoma comprises a rearranged immunoglobulin heavy chain nucleotide sequence which expresses a heavy chain polypeptide, and a rearranged immunoglobulin light chain nucleotide sequence which expresses a light chain polypeptide;
introducing into the hybridoma an exogenous nucleotide sequence which encodes a heavy chain polypeptide identical to the heavy chain polypeptide expressed by the rearranged immunoglobulin heavy chain nucleotide sequence to thereby obtain a transformant hybridoma transformed with the exogenous nucleotide sequence wherein the exogenous nucleotide sequence does not encode the light chain polypeptide;
culturing the transformant in a cell culture medium; and
obtaining a monoclonal antibody produced by the transformant.
2. The method of claim 1 , wherein the myeloma cell line is a recombinant myeloma cell line.
3. The method of claim 1 , wherein the exogenous sequence further comprises a gene-amplification gene.
4. The method of claim 3 , wherein the gene-amplification gene is dihydrofolate reductase (DHFR) gene.
5. A transformant produced by a method comprising the steps of:
fusing a B cell of a transgenic mouse which expresses human antibodies with a myeloma cell line incapable of producing antibodies to obtain a hybridoma, wherein the hybridoma comprises a rearranged immunoglobulin heavy chain nucleotide sequence which expresses a heavy chain polypeptide, and a rearranged immunoglobulin light chain nucleotide sequence which expresses a light chain polypeptide;
introducing into the hybridoma an exogenous nucleotide sequence which encodes a heavy chain polypeptide identical to the heavy chain polypeptide expressed by the endogenous immunoglobulin heavy chain to thereby obtain a transformant hybridoma transformed with the exogenous nucleotide sequence wherein the exogenous nucleotide sequence does not encode the light chain polypeptide;
culturing the transformant in a cell culture medium; and
obtaining a transformant.