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Preparation of anti cytoplasmic thymidine kinase-IgY and tumor diagnosis composition

A technology of thymidine kinase and cytoplasm, which is applied in the field of tumor diagnostic compositions, and achieves the effects of high sensitivity, extended service life, and reduced cost

Inactive Publication Date: 2003-04-30
SHENZHEN HUARUI TONGKANG BIOTECHNOLOGICAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Enhanced chemiluminescent (ECL) dot blot method is used for early serological diagnosis of malignant tumors, and the establishment of cancer detection indicators is the first at home and abroad.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1: Obtaining the amino acid sequence of the active site of cytoplasmic thymidine kinase.

[0048] TK1 in HeLa cells is a tetrameric molecule with a molecular weight of 96kD, and its monomer is composed of 234 amino acids. The C-terminal 40 amino acids of TK1 (amino sequence from 194 to 234) are the functions of TK1 to regulate cell cycle activity. key sequence. If these 40 amino acids are eliminated, TK1 will lose the activity of regulating the cell cycle. If the terminal 10 amino acids were eliminated, TK1 still showed cell cycle regulation activity. Through activity screening experiments, we selected 15 peptides with amino acid sequences from 211 to 225 and 31 peptides with amino acid sequences from 194 to 225. This polypeptide is artificially synthesized and then linked with a coupling molecule (BSA) to form an antibody preparation technology. It has been confirmed that the antibody prepared with TK1-31 peptide antigen (amino acid sequence from 194 to 225) ...

Embodiment 2

[0049] Example 2: Cell culture.

[0050] Using conventional cell culture methods, the CEM TK used in the experiment + , CEM TK - Cells were cultured (using 1640 culture medium containing 10% calf serum), when the number of cells in the culture flask grew to 40×10 6 , the harvested cells were processed.

Embodiment 3

[0051] Example 3: Preparation and purification of pT7 hTK1 engineering strain.

[0052] by literature [6] Prepared by the provided method. Purified hTK1 (15 amino acids at the N-terminal cut off) was identified as a band by natural gel electrophoresis and SDS electrophoresis, with a molecular weight of 22Kd. Further analysis by ECL Western Blot showed that hTK1 had an immune reaction with the purified anti-TK1-gY antibody, and a clear imprinted band was seen at the 22Kd position.

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Abstract

An anti-cytoplasm thymidine kinase-IgY antibody prepared from the C-terminal 31 peptide of human cervical cancer cell (HeLa)TK1, the application of the reagent kit prepared from said antibody in diagnosing tumor, the process for preparing said antibody and reagent kit, and its test method are disclosed.

Description

Technical field: [0001] The invention relates to the use of the C-terminal 31 peptide of human cervical cancer cell (HeLa) TK1 as an antigen to prepare an anti-cytoplasmic thymidine kinase-IgY antibody and a tumor diagnosis composition with the IgY antibody as an active ingredient. Background of the invention: [0002] Cancer is called "incurable disease" by contemporary people, and has become one of the major diseases that threaten human health and life. The domestic cure rate is only about 10%. The main reason for the low cure rate is that there is no ideal early diagnosis method at present. Most patients are already at an advanced stage when they see a doctor, so the key to improving the cure rate lies in the "three early", that is, early detection, early diagnosis and early treatment. There are dozens of tumor markers in China, such as AFP (alpha-fetoprotein), CEA (carcinoembryonic antigen), HCG (chorionic gonadotropin), PSA (prostate specific antigen), VCA (laryngocarci...

Claims

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Application Information

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IPC IPC(8): C07K14/435C07K16/18C12P21/02G01N33/574
Inventor Q何吴传京
Owner SHENZHEN HUARUI TONGKANG BIOTECHNOLOGICAL
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