The core principle is to combine the variable region (V region) genes (including light chain VL and heavy chain VH) of mouse monoclonal antibodies with the constant region (C region) genes (CL and CH) of human antibodies through genetic engineering recombination. The V region contains the complementary determining region (CDR), which directly mediates the high specificity of antigen recognition. The human C region determines the immune effector function of the antibody (such as complement activation, ADCC), and because it is derived from humans, it significantly reduces the rejection of the human immune system to the constant region of heterologous antibodies (HAMA reaction). 

In the implementation of the technology, the V region gene was amplified from mouse hybridoma cells, cloned by PCR and ligated into the expression vector carrying the human C region. The recombinant gene was introduced into host cells (such as CHO or HEK293), and antibody folding and glycosylation modification were completed using the eukaryotic system. The final expressed chimeric antibody not only retains the high affinity to the target antigen, but also has the long-term and low immunogenicity of the human antibody.

(1) Fast development cycle : Based on the verified variable region (V region) of mouse monoclonal antibody (mAb), the human constant region (C region) was directly cloned and recombined, without the need to screen human antibody genes from scratch, which greatly shortened the development time. 

(2) Retain high affinity : By directly utilizing the high specific antigen binding ability of mouse V region (especially CDR region), the affinity loss of humanized antibody caused by CDR transplantation can be avoided.

(1) Reduce HAMA response : The proportion of human constant region is more than 70 %, which significantly reduces the rejection of heterologous antibodies (human anti-mouse antibody response, HAMA) and prolongs the half-life of antibodies. For example, the tolerance of rituximab is significantly better than the first generation of murine monoclonal antibodies. 

(2) Avoiding the Challenge of Full Humanization : Compared with the time-consuming screening and potential functional loss of fully humanized antibodies, chimeric antibodies can balance immunogenicity and functionality through simple V-C recombination.

(1) Flexible selection of constant source area : According to the treatment needs, different IgG subtypes of C region can be selected (such as IgG1 enhances ADCC / CDC effect, IgG4 reduces inflammatory response). 

(2) Clinically verified functional retention : For example, infliximab (an anti-TNF-α chimeric antibody) activates complements and immune cells through the human IgG1 Fc segment, enhancing the inhibition of inflammatory responses.

(1) Mature expression system : Mammalian cells (such as CHO, HEK293) can efficiently express complete IgG to ensure the correctness of antibody folding and glycosylation modification. 

(2) Cost-effectiveness : Compared with the complex screening process of fully humanized antibodies, the production process of chimeric antibodies is more standardized and stable, and it is easier to achieve large-scale preparation.

The mRNA in mouse B cells was transcribed into cDNA by reverse transcriptase. The variable region genes (VH and VL) of the mouse antibody were amplified by PCR and then cloned into the framework of the human antibody. The heavy chain (VH) and light chain (VL) genes of the chimeric antibody were precisely inserted to ensure that they could be correctly expressed.

The constructed antibody gene was transfected into suitable host cells (such as CHO cells). Promote the growth of host cells through appropriate culture conditions and enable them to produce recombinant antibodies in large quantities. Commonly used host cells are CHO cells (Chinese hamster ovary cells) and HEK293 cells (human embryonic kidney cells), which have efficient protein expression capabilities.

Chimeric antibodies secreted from host cells were purified using affinity chromatography (such as Protein A affinity chromatography). The purified antibody will be further detected by SDS-PAGE and Western Blot to ensure the purity and quality of the antibody. 

Animal experiments such as mice were carried out in the laboratory to evaluate the immunogenicity, safety and efficacy of chimeric antibodies. If the experiment is successful, further clinical trials can be carried out and eventually become antibody drugs for clinical application.