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ALT (atenolol) hapten, artificial antigen, antibody, preparation methods of ALT hapten and artificial antigen and applications of hapten and antibody

An artificial antigen and hapten technology, applied in chemical instruments and methods, preparation of carboxylic acid amides, preparation of organic compounds, etc., can solve the problems of cumbersome processing, long detection time, expensive instruments, etc., and achieve low cross-reaction rate and affinity. Good results

Active Publication Date: 2019-01-04
BEIJING KWINBON BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The instrumental detection method has the disadvantages of cumbersome sample pretreatment, long detection time, and expensive instruments, so it cannot be widely used in my country, and it does not meet the requirements of on-site detection "accurate detection and screening of a large number of samples in a short period of time and at low cost".

Method used

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  • ALT (atenolol) hapten, artificial antigen, antibody, preparation methods of ALT hapten and artificial antigen and applications of hapten and antibody
  • ALT (atenolol) hapten, artificial antigen, antibody, preparation methods of ALT hapten and artificial antigen and applications of hapten and antibody
  • ALT (atenolol) hapten, artificial antigen, antibody, preparation methods of ALT hapten and artificial antigen and applications of hapten and antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Example 1: Hapten synthesis

[0025] A kind of atenolol hapten, the molecular structure is:

[0026]

[0027] It is used as a raw material for the antigen system of animal immunity.

[0028] The preparation method of the above-mentioned atenolol hapten is as follows (see the attached synthetic route figure 1 ):

[0029] Take 0.5 g of 4-(epoxyethyl methoxy) phenylacetamide, add 60 mL of ethanol to dissolve, add 0.5 mL of 1 mol / L hydrochloric acid, stir and mix, add 0.29 g of L-β-homoalanine hydrochloride, Heat and stir in an oil bath, react at 80°C for 4 hours; stop the reaction, rotary steam, remove ethanol, add 80 mL of water, add 100 mL of chloroform, extract three times, combine the organic phases, evaporate to dryness, and purify by silica gel column chromatography. The volume ratio is The atenolol hapten can be obtained by eluting with 10:1 dichloromethane-methanol.

Embodiment 2

[0030] Example 2: Artificial antigen synthesis and identification

[0031] An artificial antigen of Atenolol made from the hapten of Atenolol described in Example 1. The molecular structure is:

[0032]

[0033] 1. Synthesis of immunogen

[0034] The synthesis method of immunogen is as follows:

[0035] Take 11mg of atenolol hapten, add 2mL N,N-dimethylformamide (DMF) to dissolve, add 8mg carbodiimide (EDC), 7mg N-hydroxysuccinimide (NHS), stir at room temperature for 2h , Get hapten activation solution A; take 50mg bovine serum albumin (BSA), add 3mL of 0.1mol / L phosphate buffer to dissolve to get B solution; add drop A to B solution, stir 4h, 0.02mol / L PB buffer solution was purified by dialysis for 3 days, and the medium was changed 3 times a day to obtain atenolol immunogen, aliquoted, and stored at -20°C.

[0036] Identification of artificial antigens:

[0037] According to the ratio of hapten, carrier protein and coupling product used in the synthesis of atenolol immunogen react...

Embodiment 3

[0042] Example 3: Monoclonal antibody preparation

[0043] A specific antibody of atenolol, which is obtained by immunizing white mice with the atenolol immunogen described in Example 2 and can react specifically with atenolol for monoclonal immunoglobulin G for Detection of atenolol residues.

[0044] The preparation method of the above-mentioned atenolol monoclonal antibody is as follows:

[0045] 1) Animal immunization: The immunogen was injected into Balb / c mice, and the immunization dose was 150 μg / mouse to produce antiserum.

[0046] 2) Cell fusion and cloning: The spleen cells of Balb / c mice that produce specific antibodies are fused with myeloma cells SP20, and the cell supernatant is determined by indirect competitive enzyme-linked immunoassay to screen positive wells. The positive wells were cloned by the limiting dilution method to obtain and establish a hybridoma cell line producing monoclonal antibodies.

[0047] 3) Cell cryopreservation and resuscitation: The hybridoma c...

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Abstract

The invention provides an ALT (atenolol) hapten and an artificial antigen. The molecular structural formula of the hapten is shown in the description. The invention further discloses preparation methods of the hapten, the artificial antigen and the antibody prepared from the antigen as well as applications of the hapten and the antibody. The final prepared antibody has high detection sensitivity and strong specificity.

Description

Technical field [0001] The present invention relates to selecting a compound with -COOH and most likely to contain the original structure of atenolol as the atenolol hapten, artificial antigens and specific antibodies made from the hapten; and such hapten Use of antigens, artificial antigens, and specific antibodies, and preparation methods of haptens, artificial antigens, and antibodies. Background technique [0002] In recent years, as new environmental pollutants, pharmaceuticals and personal care products (PPCPs) have received great attention due to their potential harm to humans and aquatic organisms. These drugs cannot be completely removed in wastewater treatment plants, so the effluent from wastewater treatment plants is considered to be the main source of PPCPs entering the water environment. Due to the characteristics of drug design, many drugs have physiological effects on humans and animals and are difficult to be biodegraded. Therefore, many drugs are frequently det...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07C231/12C07C235/34C07K14/765C07K16/44G01N33/577
CPCG01N33/577C07C231/12C07C235/34C07K14/765C07K16/44C07K19/00
Inventor 吴小胜崔廷婷万宇平崔海峰王兆芹张瑜张颖赵正苗曹东山肖杉木
Owner BEIJING KWINBON BIOTECH
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