Preparation method of human antinuclear antibody

Abstract

The invention discloses a preparation method of a human antinuclear antibody, comprising the following steps: collecting single nuclear cells of peripheral blood of a patient with the autoimmune disease, extracting the total RNA (Ribonucleic Acid), and carrying out reverse transcription to obtain a cDNA (Complementary Deoxyribonucleic Acid) library; amplifying a light chain k,1 gene and an immunoglobulin molecule heavy chain Fd gene by taking the acquired cDNA library as a template through a PCR (Polymerase Chain Reaction) method; cloning the light chain k,1 gene into a pComb3Hss carrier to construct a light chain library; loading the heavy chain Fd gene into a carrier to construct and finish a combinatorial library; transforming the obtained recombinant plasmid into escherichia coli XL1-Blue, infecting by using a helper phage M12KO7, and expressing the random combinatorial library on the surface of a filamentous phage to finish phage surface display; and purifying the Fab segment of the human antinuclear antibody by utilizing affinity chromatography. By using the preparation method, the defects that the random combinatorial antibody library has strong randomness, the screening workload is large and a specific antibody is not easy to obtain are overcome, and the specific antibody can be directly screened from a phage antibody library; and in addition, the method is simple and feasible, and the experimental period is shortened.

Description

technical field [0001] The invention belongs to the field of biochemistry, in particular to a preparation method of human antinuclear antibody. Background technique [0002] Tumor is the biggest threat to human health and life at present. The incidence rate is increasing, and the mortality rate has leapt to the top of all diseases. Surgery, radiotherapy, chemotherapy and other methods are mainly used for treatment, but satisfactory curative effect is still not obtained. Tumor necrosis therapy is a new therapy emerging recently, which belongs to the category of radioimmunotherapy. Its basic principle is to use the discontinuity of microvascular endothelial cells of tumor tissue, the incompleteness of basement membrane and the high permeability of tumor cell membrane. , the application of targeting molecules that can combine with the nuclear antigen components of cells to carry tumor-killing substances, stay or locate in tumor tissues, and cause tumor-specific killing. The ta...

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Problems solved by technology

However, the application of this method to prepare monoclonal antibodies for radioimmunotherapy has the following defects: first, as a mouse / human chimeric monoclonal antibody, it still has relatively serious host antibody reactions, and it is not suitable for multiple applications; second, the preparation technology It is relatively complicated, difficult and demanding, and the success rate is low; third, the monoclonal antibody obtained so far has a large molecular weight and poor tissue penetration, which is not conducive to bringing tumor-killing substances into the tumor; fourth, because it targets Tumor necrosis tissue, the antigen composition is more complex, so its specificity is poor

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