Hybridoma technology generally refers to B lymphocyte hybridoma technology. It is an experimental technique to hybridize immune spleen cells with homologous animal myeloma cells by cell fusion technique and obtain the hybrid cells secreting predetermined specific antibodies through screening. Redbert (Beijing) Biotechnology Co., Ltd. will introduce hybridoma technology to you.

In 1975, two scholars, Kohler and Milstein, developed a technique based on cell fusion. The hybridoma cell line secreting SRBC monoclonal antibody was established by fusing mouse myeloma cells in vitro with mouse spleen cells immunized with sheep red blood cell (SRBC). The results showed that these cells could be continuously cultured in vitro. It opened a new era in antibody preparation and application.

Hybridoma technique is a long cycle and continuous experimental technique system. The experiments mainly include animal immunity, preparation of myeloma cells and immune spleen cells, cell fusion, screening of positive hybridoma, clone culture, preparation, purification and identification of monoclonal antibody.

According to Burnet antibody selection theory, a B lymphocyte can produce a kind of specific antigenic determinant antibody against its recognition, from a small plant cloning is the nature of the same uniform antibodies, called monoclonal antibodies, has exactly the same molecular structure and biological characteristics, but the B lymphocyte secretion has important effect on long-term survival in vitro.

Cell fusion technique was used to fuse B lymphocytes with myeloma cells with long-term proliferation in vitro. Hybridoma cells obtained by selective culture are called hybridoma cells. This work not only has the function of immune B lymphocyte secreting specific mAb, but also has the infinite proliferation ability of myeloma cells, using hybridoma cells to produce monoclonal antibody.

There are two pathways to DNA biosynthesis in tumor cells. The first is the main pathway of biosynthesis, the synthesis of nucleotides from amino acids and other small molecular compounds to DNA. Folic acid derivatives play an important role in nucleotide synthesis during nucleic acid synthesis. Aminopterin is a folic acid antagonist that blocks the main pathway of DNA biosynthesis in cells.

At this point, if the medium contains the intermediate products hypoxanthine and thymine of nucleotide synthesis, DNA synthesis of the cell can be carried out by another pathway, i.e. nucleotide synthesis by exogenous hypoxanthine and thymine catalyzed by HGPRT and TK. In the absence of any of these enzymes, the pathway is blocked.

After the main pathway of DNA synthesis was blocked by A, the non-fused myeloma cells and their homonuclear fused bodies in HAT medium could not utilize H in culture medium due to lack of HGPRT. Although TK can use T, it cannot complete the DNA synthesis process. The unfused splenic cells and their homonuclear fused bodies could not grow in the medium for a long time and died about 7 days later.

Heteronuclear fusion of spleen cells with myeloma cells (i.e. hybridoma cells) not only obtains HGPRT and TK from spleen cells, synthesizes DNA using H and T in the culture medium, but also obtains unlimited proliferation ability from myeloma cells, enabling them to survive in HAT culture medium.

Monoclonal antibody technology has become an important routine technology in immunology, and has been widely used in cell biology, genetics, microbiology, biochemistry, oncology and other life sciences. The diagnostic revolution caused by monoclonal antibodies is no longer limited to scientific research. It will have an increasingly important impact in clinical therapeutics and other areas of the economy. At present, the Beacon instrument has a single-cell light guide system. By integrating photoelectric positioning technology and microfluidic technology, it realizes the high-throughput cell biology research based on a single cell in the nano-upgrade cell of the chip, which is of great significance to the single-cell cloning technology.

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