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Hybridoma cell line YY capable of secreting anti-sildenafil monoclonal antibody and its application

A hybridoma cell line and monoclonal antibody technology, which can be applied to analytical materials, microorganism-based methods, biochemical equipment and methods, etc., can solve problems such as method limitation, high cost, and restrict the popularization of detection methods, and achieve simplified synthesis steps, The effect of good detection sensitivity and affinity

Active Publication Date: 2017-09-22
JIANGNAN UNIV +1
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Problems solved by technology

[0003] At present, physical and chemical methods such as ultraviolet spectroscopy, thin-layer chromatography and high-performance liquid chromatography are mostly used in the detection of sildenafil. Ultraviolet spectroscopy and thin-layer chromatography are difficult to exclude due to interference, and the methods are limited.
Instrumental detection methods can perform quantitative analysis and have a low detection limit, but usually require expensive instruments and complicated operations, and the pretreatment and detection time is long, which seriously restricts the promotion of these detection methods

Method used

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  • Hybridoma cell line YY capable of secreting anti-sildenafil monoclonal antibody and its application
  • Hybridoma cell line YY capable of secreting anti-sildenafil monoclonal antibody and its application
  • Hybridoma cell line YY capable of secreting anti-sildenafil monoclonal antibody and its application

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Embodiment 1

[0019] Embodiment 1: the preparation of the hybridoma cell line YY of anti-sildenafil monoclonal antibody

[0020] 1. Synthesis of sildenafil hapten: 500 mg sildenafil and 90 mg glycine were dissolved in 20 mL of methanol, and 236.4 mg of triethylamine was added while stirring. The mixture was reacted at 70°C for 4 hours. After the reaction was finished, 50ml of distilled water was added into the reactant to terminate the reaction. Then extract three times with ethyl acetate, combine organic layer, wash three times with saturated NaCl solution, NaCl 2 SO 4 Remove the water in the organic phase, pass through a silica gel column, and dry in vacuo to obtain a yellow solid powder, which is the hapten of sildenafil.

[0021] 2. Synthesis of complete antigen: take 3mg sildenafil hapten, add 4.5mg EDC (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) and 3.6mg NHS ( N-hydroxysuccinimide), dissolved in DMF (N,N-dimethylformamide), stirred at room temperature, activated...

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Abstract

The invention discloses a hybridoma cell line YY capable of secreting an anti-sildenafil monoclonal antibody and its application and belongs to the technical field of food safety immunoassay. The hybridoma cell line YY is preserved in the China general microbiological culture collection center (CGMCC) and has an accession number CGMCC No. 13100. The hybridoma cell line YY can secrete a monoclonal antibody having good affinity and high sensitivity to sildenafil and 50% inhibition concentration IC50 to sildenafil of 0.48 micrograms per milliliter. The hybridoma cell line YY can be used for preparation of a sildenafil immunoassay kit and a colloidal gold test strip and provides a good detection method for sildenafil in Chinese patent medicines and health products.

Description

technical field [0001] The invention relates to a hybridoma cell line YY secreting anti-sildenafil monoclonal antibody and application thereof, which belongs to the field of food safety immune detection. Background technique [0002] Sildenafil (Sild), full name Sildenafil citrate, is the main active pharmaceutical ingredient of Viagra (also known as "Viagra"). Sildenafil is a cyclic guanosine monophosphate (cGMP)-specific phosphodiesterase type 5 (PDE5) selective inhibitor, which can be used to treat erectile dysfunction (ED). In recent years, incidents of illegal addition of chemicals in Chinese patent medicines and health foods have been repeatedly banned. Chinese patent medicines for nourishing kidney and strengthening yang and health care products have become the hardest hit areas for illegal addition due to their huge number of users and huge market sales. Therefore, how to establish Cost-saving, targeted and efficient rapid inspection methods, cracking down on counte...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/20C07K16/44G01N33/577C12R1/91
Inventor 胥传来王忠兴匡华徐丽广刘丽强马伟吴晓玲宋珊珊胡拥明
Owner JIANGNAN UNIV
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