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Single chain antibody of human anti-placenta growth factor

A technology for placental growth factor and single-chain antibody, which is applied to the preparation method of anti-growth factor immunoglobulin, anti-animal/human immunoglobulin, peptide, etc. Single-chain antibodies, etc.

Inactive Publication Date: 2010-02-10
SHANGHAI TENTH PEOPLES HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] There is no report on the successful preparation of human-derived anti-placental growth factor single-chain antibody

Method used

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  • Single chain antibody of human anti-placenta growth factor
  • Single chain antibody of human anti-placenta growth factor
  • Single chain antibody of human anti-placenta growth factor

Examples

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Embodiment 1

[0033] Extract the complete set of genetic information of antibodies from peripheral blood lymphocytes of patients with ovarian cancer, construct a single-chain antibody cDNA library, screen out the target antibody by immunomagnetic bead method, and enrich the target antibody gene by in situ PCR to obtain a high-concentration human Source anti-placental growth factor (placental growth factor, PIGF) single-chain antibody, the specific implementation method is as follows:

[0034] (1) Dilute 3mL of peripheral blood of ovarian cancer patient volunteers with an equal amount of PBS, add to 6mL of human lymphocyte separation medium, centrifuge at 2500r / min for 20min, and collect the lymphocyte layer;

[0035] (2) Use TRIZOL reagent (Cat.#SK1312, purchased from Shanghai Yuanxiang Biological Co., Ltd.) and extract total RNA according to its instructions;

[0036] (3) Using total RNA as a template, Oligo dT as a primer, and dNTP as a raw material, synthesize the first strand of cDNA un...

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Abstract

The invention relates to a single chain antibody of human anti-placenta growth factor, which is encoded by a heavy-chain gene with an SEQ NO.1 sequence and light-chain gene with an SEQ NO.2 sequence.The invention also provides a preparation method thereof, comprising the following steps in sequence: amplifying human total RNA heavy-chain variable region, light-chain variable region and constant region gene of the whole set by a PCR technique, splicing and constructing an overall-length single-chain antibody cDNA library; expressing the cDNA library by an externally-coupled transcription / translation system to obtain an antibody-ribosome-mRNA compound; performing affiliation selection and clution on the compound by a magnetic bead coated by specific antigen; amplifying selected destinationantibody gene by the PCR technique; and expressing the amplified antibody gene using the externally-coupled transcription / translation system repeatedly to obtain the single-chain antibody. The antibody is adapted to prepare medicament for treating ovarian cancer.

Description

technical field [0001] The present invention relates to human-derived anti-placental growth factor single-chain antibody. Background technique [0002] Placental growth factor (placental growth factor, PIGF) was first isolated and purified from the human placenta cDNA library by Maglione et al. in 1991. It is a subtype of VEGF (vascular endothelial growth factor), which is independent of VEGF in the main expression VEGFR-1 endothelial cells, macrophages, bone marrow stem progenitor cells and tumor cells. Studies have shown that serum and tumor PIGF levels are related to tumor grade, vascular distribution, recurrence, metastasis and growth status. Levels of αVEGF (anti-VEGF antibody) were not decreased in cancer patients treated with αVEGF, nor in patients treated with radiation, suggesting that PIGF plays a key role in the angiogenic rescue program. [0003] The latest research shows that anti-placental growth factor antibodies can inhibit tumor growth. Fischer et al. rep...

Claims

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Application Information

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IPC IPC(8): C07K16/22C12P21/08C07K1/22
Inventor 李芳苏平平朱怀仕
Owner SHANGHAI TENTH PEOPLES HOSPITAL
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