Monoclonal antibody of human insulin-like growth factor binding protein-1, relative host cell, diagnostic reagent and diagnostic reagent kit

A technology of monoclonal antibodies and diagnostic kits, applied in the fields of genetic engineering and immunology, can solve the problems of undetectable and achieve the effect of sensitive response and low cost

Inactive Publication Date: 2010-08-18
无锡博慧斯生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0011] 4) Amnioscopic examination: For suspicious medical records, the fetal membranes can be directly observed under the amniotic microscope, but certai...

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1 Antigen preparation (amino acid sequence shown in SEQ ID NO: 1)

[0036] Put the peptide synthesis column in the Pioneer TM In a peptide synthesizer, the peptides were synthesized under nitrogen according to the amino acid sequence of the small peptides.

[0037] The resin was washed with THF for about 5 minutes.

[0038]Filter and centrifuge the filtrate in cold diethyl ether, pour off the supernatant and repeat the centrifugation with cold diethyl ether until the peptide is completely precipitated, and the obtained crude peptide is purified by preparative C18 silica gel column chromatography, eluted with acetonitrile gradient, and the fraction containing the target peptide is collected. Fractions were eluted and lyophilized to yield the target peptide.

Embodiment 2

[0039] Embodiment 2 animal immunization

[0040] The antigen obtained in Example 1 was coupled to KLH, 4 mg of the conjugate was dissolved in 5 ml of normal saline, mixed with complete Freund's adjuvant 1:1 (volume ratio), and 8-week-old magnetic Balb / c mice were taken, Animal immunizations were performed according to the following protocol:

[0041] (1) Initial immunization (small peptide 50 μg, plus Freund's complete adjuvant subcutaneous multi-point injection, 1ml 0.2ml / point);

[0042] (2) Second immunization three weeks later (same dose as above, subcutaneous injection with Freund's incomplete adjuvant, 0.5ml)

[0043] (3) The third immunization after three weeks (dose as above, subcutaneous injection without adjuvant, 7 days later blood collection to measure its titer and immune effect);

[0044] (4) Booster immunization three weeks later (dose 500 μg, subcutaneous injection without adjuvant);

[0045] (5) Take the spleen for fusion.

[0046] (6) At the end of the fo...

Embodiment 3

[0047] Example 3 Cell Fusion and Screening

[0048] Sacrifice the mice, prepare a single splenocyte suspension, mix in a 50ml centrifuge tube for 10 8 splenocytes and 10 7 Mouse myeloma cells (NS-1 cells) growing in the logarithmic phase, and add 30ml simple medium, centrifuge at 800 rpm at room temperature for 8 minutes, remove the supernatant, put the centrifuge tube in a water bath at 37°C, and slowly Add 0.8ml of 50% (percentage by weight) polyethylene glycol-4000, complete the addition in 2 minutes, shake the centrifuge tube while adding, then slowly add the simple medium, add 4ml within 3 minutes, centrifuge at room temperature for 8 minutes at 800 rpm, Remove the supernatant, add 45ml of 20% (volume percentage) HAT selective medium of calf serum, inoculate in three 96-well plates after blowing, in 5% CO 2 After 7 days, the culture solution in the culture plate was encrypted, observed after 10 days, and the antibody titer of the hybridoma cell culture solution was dete...

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PUM

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Abstract

The invention provides a monoclonal antibody of human insulin-like growth factor binding protein-1, a monoclonal antibody host cell, a diagnostic reagent with the monoclonal antibody of the human insulin-like growth factor binding protein-1and a diagnostic reagent kit, wherein antigen of the monoclonal antibody is antigenic determinant of the human insulin-like growth factor binding protein-1; preferably, the antigenic determinant comprises an amino acid sequence represented by SEQIDNO:1; and the monoclonal antibody host cell contains the monoclonal antibody of the human insulin-like growth factor binding protein-1. The monoclonal antibody of the human insulin-like growth factor binding protein-1 can perform specific detection on the human insulin-like growth factor binding protein-1, and the prepared diagnostic reagent has high specificity of premature rupture of membrane diagnosis, rapid reaction, low cost and suitability for large-scale popularization and application.

Description

technical field [0001] The present invention relates to the technical field of genetic engineering and immunology, more specifically to the technical field of monoclonal antibody, in particular to a monoclonal antibody of human insulin-like growth factor binding protein-1, related host cells, diagnostic reagents and diagnostic Reagent test kit. Background technique [0002] Premature rupture of membranes (PROM) is called premature rupture of membranes before full-term delivery; the incidence of premature rupture of membranes is about 10%, and about 30% of pregnant women are clinically tested for PROM; At present, the number of pregnant women nationwide is about 10-14 million. [0003] Therefore, premature rupture of membranes is one of the most common causes of premature birth and neonatal complications, and it is also one of the most difficult treatment difficulties encountered in current obstetrics clinics. Diagnosis after rupture of membranes 24 hours increases fetal mo...

Claims

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Application Information

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IPC IPC(8): C07K16/18C12N5/20G01N33/577
Inventor 韦彦余刘丽雷震张丽新李雅丽
Owner 无锡博慧斯生物医药科技有限公司
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