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
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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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