Sp2/0 cell immortality improvement based hybridoma technique

Abstract

An sp2 / 0 cell immortality improvement based hybridoma technique for the field of biology is characterized in that hTERT with an immortality characteristic is adopted for transfection of rat sp2 / 0 tumor cells through a pcDNA3.1-hTERT recombinant retroviral vector to obtain immortality characteristic improved recombinant vector packaged rat sp2 / 0 tumor cells, then hTERT is guided into hybridoma cells through cell screening and fusion, and accordingly the immortality characteristic of the hybridoma cells is enhanced, the cell fusion success rate and fusion rate are increased, the cell survival rate in a cloning screening process is increased, the cell mortality rate in positive clone cell subculturing is decreased, confluence in cell adherence growth is increased, and a foundation is laid forindustrial amplification of fusion cells and further amplification of pathogenic genes in the fusion cells.

Description

technical field [0001] The invention relates to a hybridoma technology based on sp2 / 0 cell immortalization improvement in the field of biology, which is mainly used for cell hybridization and whole genome amplification and storage in the fields of scientific research, pharmacy and diagnosis. Background technique [0002] Hybridoma technology is lymphocyte hybridoma technology, also known as monoclonal antibody technology. [0003] Kohler and Milstein (1975) proved that myeloma cells were hybridized with spleen cells of immunized animals to form monoclonal antibodies that can secrete homogeneous and highly specific antibodies against the antigen. The technology is commonly known as hybridoma technology. Myeloma cells can be continuously passaged in vitro, while spleen cells are terminal cells that cannot be propagated in vitro. For example, if the myeloma cells of mice are hybridized with lymphocytes secreting certain antibodies or factors, the hybrid cells not only have th...

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Problems solved by technology

However, if it is transferred to the preparation of hybridoma cells hybridized between human gene mutant lymphocytes and mouse sp2 / 0 tumor cells, we found that although they can hybridize, there are the following three problems: (1) Human gene mutant hybridoma cells are attached Floating death while growing on the wall: There are always about 1 / 2 to 1 / 3 of the cells floating in the culture medium due to death, and the confluence of living cells attached to the wall is generally 30-50%, and usually does not reach 80-90% The above ideal confluence; (2) Rare disease hybridoma cells adhere to the wall while growing and dying: usually about 1 / 2 to 1 / 3 of the cells will die by attaching to the wall, and will not reach more than 80-90% viability Ideal confluence for adherent growth

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