Sildenafil monoclonal antibody and colloidal gold chromatography test strip used for detecting sildenafil

A monoclonal antibody, sildenafil technology, applied in the direction of microorganism-based methods, analytical materials, measuring devices, etc., can solve the problems of no sildenafil monoclonal antibody report, long reaction time, high cost of experiments, etc. Achieve the effect of being convenient for human handling and quality control, easy to operate, and single biological activity

Active Publication Date: 2011-09-07
NANTONG EGENS BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, traditional methods are used to detect whether Viagra is contained in Chinese patent medicines or health care products. The extracted samples must be brought back to the laboratory for analysis, or large-scale test vehicles are used. This is very inconvenient and the reaction Long time, expensive experiment
However, there is no relevant report about sildenafil monoclonal antibody

Method used

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  • Sildenafil monoclonal antibody and colloidal gold chromatography test strip used for detecting sildenafil
  • Sildenafil monoclonal antibody and colloidal gold chromatography test strip used for detecting sildenafil
  • Sildenafil monoclonal antibody and colloidal gold chromatography test strip used for detecting sildenafil

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] A synthesizes N-acetate sildenafil (II-i)

[0051] (1) Alkylation: Add 20ml of dry dimethylformamide (DMF) and 1.708g, 3.6×10 -3 mol of sildenafil (IV-i), and through N 2 protection, stirred at room temperature, and after dimethylformamide (DMF) was completely dissolved, quickly weighed 0.173g, 4.32×10 -3 mol, concentration is 60% sodium hydride (NaH), and it is added in the described reaction bottle, stirs 60 minutes, until the solution produces without bubble, draws 1.202g, 7.2 * 10 with 1ml syringe then -3 mol of ethyl bromoacetate was slowly injected into the reaction flask, and after the injection was completed, it was stirred at room temperature for 12 hours, and then the reaction solution was poured into 60ml of water to disperse, extracted with 20ml×3 chloroform, and the extracts were combined to obtain Anhydrous Na 2 SO 4 Dry, then filter, and then concentrate under reduced pressure to obtain a solid, add 6ml of anhydrous ether to wash the solid, then filte...

Embodiment 2

[0065] The establishment of embodiment 2 hybridoma cell lines

[0066] Step 1 Animal Immunization

[0067] Artificially synthesized antigen N-sildenafil acetate-BSA was emulsified with an equal amount of complete Freund's adjuvant, and 0.2ml (1mg / ml) was subcutaneously injected into mice at multiple points. After 3 weeks, N-sildenafil acetate-BSA plus the same amount of Freund's incomplete adjuvant was fully mixed, and then subcutaneously injected 0.2ml (1mg / ml) into mice at multiple points. After 10 days, the tail was docked to collect blood One drop was used to measure the antibody titer, and mice with high titers were selected for fusion test. One month later, 0.1 ml of non-adjuvant antigen N-sildenafil acetate-BSA can be administered intravenously, and after 3 days, the mice are killed and spleens are taken for fusion.

[0068] A mouse myeloma line was induced from the Balb / c mouse line. Balb / c mice must be of pure line, both male and female, preferably 8-12 weeks old. ...

Embodiment 3

[0106] Example 3 In vivo induced ascites method to prepare monoclonal antibody

[0107] Usually, the method of producing monoclonal antibody in animals is adopted. Since the vast majority of animal hybridomas are obtained from the fusion of myeloma cells of Balb / c mice and spleen cells of the same strain, the animal used is of course the first choice of Balb / c mice. c mice. In this method, the hybridoma cells are inoculated into the peritoneal cavity of the mouse, the hybridoma is grown in the peritoneal cavity of the mouse, and ascites is produced, so a large amount of monoclonal antibody in the ascitic fluid can be obtained with a high antibody concentration. It can be seen that this method is easy to operate and economical, but ascites is often mixed with various miscellaneous proteins of mice, so in many cases it must be purified before use, and there is also the risk of contamination of animal viruses, so it is best to use SPF mice . The operation method is as follows: ...

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Abstract

The invention belongs to the field of medicament inspection and discloses a sildenafil monoclonal antibody and a colloidal gold chromatography test strip used for detecting sildenafil. The sildenafil monoclonal antibody is generated by a hybridoma cell line with the collection number of CCTCC No. C2010123. The sildenafil monoclonal antibody has highly uniform physical and chemical properties, single bioactivity and high specificity of combining the antigen sildenafil, is convenient to perform artificial treatment and quality control, and can be used for preparing the colloidal gold chromatography test strip for detecting the sildenafil. In the colloidal gold chromatography test strip for detecting sildenafil, the gold-labelled antibody coated on a gold-labelled combination pad is the colloidal gold marker of the sildenafil monoclonal antibody. By using the colloidal gold chromatography test strip, no any reagent and instrument are required, and site operation can be performed; and after a sample is mixed with an extraction solution, the supernatant is taken for detection, the detection result can be obtained within 5-10 minutes, and the method is simple, convenient and rapid and has high timeliness.

Description

technical field [0001] The invention belongs to the field of drug inspection, and relates to a sildenafil monoclonal antibody and a colloidal gold chromatography test strip for detecting sildenafil. Background technique [0002] Sildenafil (Sildenafil), also translated as sildenafil, is a drug for the treatment of male erectile dysfunction accidentally invented when developing drugs for the treatment of cardiovascular diseases. Hong Kong registered name Wei Ergang) is widely known. However, compared with the product name Sildenafil, the common name "Viagra" in China is more widely used and has a greater impact. At present, sildenafil is illegally added to many domestic Chinese patent medicines or health products to achieve clinical efficacy. As a patient, taking Chinese patent medicines or health products containing sildenafil without knowing for a long time will cause serious side effects . At present, traditional methods are used to detect whether Viagra is contained in...

Claims

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Application Information

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IPC IPC(8): C12N5/20C07K16/44G01N33/577C12R1/91
Inventor 欧卫军吴迪宏蒋澄王蕾
Owner NANTONG EGENS BIOTECH
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