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A fluorescent detection method for bacterial drug resistance and its diagnostic kit

A technology of fluorescence detection and detection method, which is applied in the fields of medicine and biology, can solve the problems of non-specific detection of broad-spectrum β-lactamases, etc., and achieve the effect of facilitating early clinical treatment, gaining treatment time, and improving the success rate of treatment

Active Publication Date: 2017-04-12
SHANGHAI BAIKE BIO TECH CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

GeneBLAze reagent is also based on fluorescence detection, however, this reagent can only be used to detect the presence of all β-lactamases indiscriminately, but cannot specifically detect the broad-spectrum β-lactamases that can degrade the third and fourth generations of cephalosporins

Method used

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  • A fluorescent detection method for bacterial drug resistance and its diagnostic kit
  • A fluorescent detection method for bacterial drug resistance and its diagnostic kit
  • A fluorescent detection method for bacterial drug resistance and its diagnostic kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Embodiment 1, the preparation of fluorescent substrate C-1

[0060] synthetic route

[0061]

[0062] Cefotaxime (cefotaxime, 455.5 mg, 1.00 mmol) and toluenesulfonic acid (172.2 mg, 1.00 mmol) were dissolved in 5 mL of acetonitrile and 8 mL of aqueous solution; Diphenyldiazomethane (199.2 mg, 1.00 mmol , 1 equivalent); mixed and stirred at -20°C for 4 to 5 minutes until the characteristic purple color disappeared; after adding sodium hydroxide solution to the mixture to adjust the pH value to 9, it was extracted with ethyl acetate (50 ml); organic The layers were washed twice with water (15 mL), then dried over anhydrous calcium chloride; the solvent was removed using a rotary evaporator. The enrichment was separated and purified on a 50 g silica gel column with 30% n-hexane-ethyl acetate; the collected fractions were removed by rotary evaporation to obtain 500 mg of compound 1 as a white solid (yield 80%).

[0063] Proton NMR spectrum (deuterated chloroform, 600M...

Embodiment 2

[0073] Example 2 Preparation of Bacterial Drug Resistance Fluorescent Detection Kit and Detection of Bacterial Drug Resistance Experiment

[0074] 1) The solutions required to configure the detection kit are as follows:

[0075] Fluorescence substrate 1mL, concentration 1 micromol / L, prepared with double distilled water before use.

[0076] Non-denaturing cell buffer. 1 mL includes: 20 mmol / L sodium phosphate buffer at pH 7.5, 150 mmol / L sodium chloride, 1% polyethylene glycol octylphenyl ether (Triton X-100 )

[0077] 2) The use and detection method of the fluorescent detection kit for bacterial resistance

[0078] The method for using the fluorescent detection kit for bacterial resistance, that is, the detection operation procedure or detection method of the fluorescent detection kit for bacterial resistance.

[0079] The detection method of the present invention is based on the fluorescence resonance energy transfer phenomenon, so the detection of drug-resistant bacteria...

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Abstract

The invention provides a fluorescence detection method for detecting bacterial drug resistance in an individual and a diagnostic kit / diagnostic reagent thereof. According to the fluorescence detection method, absorbance difference under the condition of different wavelengths is detected by using a fluorogenic substrate, which only can be degraded by broad spectrum beta lactamase, through double fluorescence signals and further the bacterial drug resistance of the individual or the biological sample of the individual can be detected. The detection method and the obtained detection kit based on the method can detect the 3rd and 4th generations of drug-resistance bacteria of cephalosporin, which cannot be detected by products available in the markets and can reach a detection value of 10<5>-10<8> drug-resistance bacteria, so that a market blank is filled in, more effective monitoring method and use guide are provided for the use of antibiotics in clinic, and extremely great practical values are achieved.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a fluorescent detection of bacterial drug resistance and its diagnostic kit, more specifically to a fluorescent detection of bacterial drug resistance treated with β-lactam antibiotics and its diagnosis The test kit belongs to the technical field of medicine. Background technique [0002] Since humans first discovered β-lactam antibiotics represented by penicillin in the 1920s, they have become the first-line drugs for the treatment of bacterial infections and saved countless lives. Compared with other classes of antibiotics, β-lactam antibiotics have the characteristics of high curative effect and low toxicity. However, bacteria are also constantly evolving various resistance mechanisms. Currently, drug-resistant bacterial infections have become a global problem. Especially in China, India, and some other countries, due to the phenomenon of overuse of antibiotics, the emergence o...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/64
Inventor 王郁生
Owner SHANGHAI BAIKE BIO TECH CO LTD
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