Gene amplification methods

A method of producing a transformed cell that co-expresses a combination of VH and VL regions to form a multivalent composition from a subject's B-cell lymphoma cells, comprising:

• a) providing:
• i) malignant cells isolated from a subject having a B-cell lymphoma;
• ii) an amplification vector comprising a recombinant oligonucleotide having a sequence encoding a first inhibitable enzyme operably linked to a heterologous promoter;
• iii) a T lymphoid parent cell line;
• b) isolating nucleic acid from said malignant cells, said nucleic acid comprising nucleotide sequences selected from the group consisting of nucleotide sequences encoding at least one VH region and at least two VL regions, nucleotide sequences encoding at least two VH regions and at least one VL region, and nucleotide sequences encoding at least two VH regions and at least two VL regions, wherein said at least two VL regions differ by at least one idiotope, wherein said at least two VH regions differ by at least one idiotope, and wherein said nucleic acid comprising nucleic acid sequences encoding said VH and VL regions is derived from nucleic acid encoding immunoglobulin molecules expressed by said malignant cells;
• c) inserting said nucleic acid comprising nucleotide sequences encoding said VH regions into a plurality of first expression vectors, and inserting said nucleic acid comprising nucleotide sequences encoding said VL regions into a plurality of second expression vectors;
• d) introducing said pluralities of said first and second expression vectors and said amplification vector into said parent cell line to generate transformed cells, wherein a ratio ranging from 2:20 to 2:50 of said amplification vector to said first or second expression vector is employed;
• e) introducing said transformed cells into a first aqueous solution, said first aqueous solution comprising an inhibitor capable of inhibiting said first inhibitable enzyme, wherein the concentration of said inhibitor present in said first aqueous solution is sufficient to prevent growth of said parent cell line;
• and
• f) identifying a transformed cell capable of growth in said first aqueous solution, wherein said transformed cell capable of growth expresses a combination of VH and VL regions selected from the group consisting of at least one VH region and at least two VL regions, at least two VH regions and at least one VL region, and at least two VH regions and at least two VL regions, wherein said at least two VL regions differ by at least one idiotope, wherein said at least two VH regions differ by at least one idiotope, and wherein said VH and VL regions comprise a protein molecule,
• wherein said multivalent composition comprises said expressed combination of VH and VL regions.