CN115856297B - Preparation method of kit for detecting salmonella typhimurium and kit - Google Patents
Preparation method of kit for detecting salmonella typhimurium and kit Download PDFInfo
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Classifications
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention provides a preparation method of a kit for detecting salmonella typhimurium and the kit, and belongs to the technical field of kits. According to the invention, magnetic beads with superparamagnetism are prepared firstly, the magnetic beads are used as magnetic cores, organic covalent frameworks (COFs) are coated to form core-shell structures, namely MCOF nanometer magnetic particles, then gold nanometer particles are loaded on MCOF nanometer magnetic particles by an in-situ reduction method, and finally the gold nanometer particles are coupled with an aptamer of salmonella typhimurium to synthesize a nanometer probe apt-MCOF-AuNPs with high fluorescence emission. The salmonella typhimurium and the nano-probe are incubated together to form a probe-thallus complex, a quencher Fe 3+ is added to quench the fluorescence of the probe, and the fluorescence of the probe is not quenched by Fe 3+ due to the protective effect of the thallus on the nano-probe and the competitive combination of the thallus and Fe 3+, so that the solution is bright green fluorescence under 365nm ultraviolet light irradiation. Thereby realizing the rapid and specific detection of salmonella typhimurium.
Description
Technical Field
The invention relates to the technical field of kits, in particular to a preparation method of a kit for detecting salmonella typhimurium and a kit prepared by the preparation method.
Background
Salmonella typhimurium (Salmonella typhimurium, S.tyrmium) is a common food-borne pathogenic bacterium that can cause an outbreak of food-borne diseases. Salmonella typhimurium often contaminates food products such as milk, eggs, fruits and vegetables, meats, and the like by animal manure. Salmonella typhimurium infection can cause symptoms of typical food poisoning, such as abdominal pain, diarrhea, nausea, vomiting, fever, and other gastrointestinal symptoms. It is reported that the incidence of food poisoning caused by salmonella typhimurium is still on the rise. At present, the bacteria are necessary pathogenic bacteria for agricultural and sideline products such as eggs, meat, milk and the like.
Traditional salmonella typhimurium detection methods include plate culture, enzyme-linked immunosorbent assay (ELISA) and Polymerase Chain Reaction (PCR). Although these techniques have become gold standard or recommended detection methods, in order to meet the needs of salmonella typhimurium on-site detection, there is also a need to construct a detection method that is more convenient, easy to operate, and easy to read the results. Over the past few years, various new types of biosensors have been widely developed, such as colorimetric sensors, fluorescence sensors, surface Enhanced Raman Scattering (SERS), and electrochemical biosensors. They have the advantages of quick response, convenient use, low cost, good sensitivity, good selectivity, etc. Among them, the fluorescence sensor has advantages of detection sensitivity and less susceptibility to the influence of fluorescent signals, and detects an analyte by a fluorescent color change using a portable optical measurement device. In addition, in combination with the image acquisition and image analysis system of the smart phone, qualitative and semi-quantitative analysis services can be provided.
Disclosure of Invention
In view of the above, in order to meet the requirement of salmonella typhimurium on-site detection, on the one hand, the invention provides a preparation method of a kit for detecting salmonella typhimurium, which comprises the steps of firstly preparing magnetic beads with superparamagnetism, taking the magnetic beads as magnetic cores, coating organic covalent frameworks (COFs) to form core-shell structures, namely MCOF nanometer magnetic particles, then loading gold nanometer particles on MCOF nanometer magnetic particles by an in-situ reduction method, and finally coupling with an aptamer of salmonella typhimurium to synthesize a nanometer probe apt-MCOF-AuNPs with high fluorescence emission. The salmonella typhimurium and the nano-probe are incubated together to form a probe-thallus complex, a quencher Fe 3+ is added to quench the fluorescence of the probe, and the fluorescence of the probe is not quenched by Fe 3+ due to the protective effect of the thallus on the nano-probe and the competitive combination of the thallus and Fe 3+, so that the solution is bright green fluorescence under 365nm ultraviolet light irradiation. Thereby realizing the rapid and specific detection of salmonella typhimurium.
In order to achieve the above purpose, the present invention provides the following technical solutions:
a preparation method of a kit for detecting salmonella typhimurium comprises the following steps:
Step one: preparing Fe 3O4 nano particles with superparamagnetism;
step two: preparation of magnetic organic framework MCOF
Taking the Fe 3O4 nano-particles prepared in the first step as a magnetic core, coating an organic covalent framework (COF) to form a core-shell structure, and obtaining MCOF magnetic nano-particles;
step three: preparation of MCOF-AuNPs nanocomposite
Loading gold nanoparticles on MCOF magnetic nanoparticles prepared in the second step to obtain a MCOF-AuNPs nanocomposite;
Step four: preparation of apt-MCOF-AuNPs nanoprobe
Coupling MCOF-AuNPs nano composite material obtained in the step three with an aptamer of salmonella typhimurium to synthesize a nano probe apt-MCOF-AuNPs with high fluorescence emission;
Step five: preparing Fe 3+ quencher, sealing and keeping away from light for standby.
Preferably, the specific steps are as follows:
step one: preparation of Fe 3O4 nanoparticles
Magnetically stirring FeCl 3·6H2O、CH3COONa·3H2 O in ethylene glycol at room temperature until the FeCl 3·6H2O、CH3COONa·3H2 O is completely dissolved, transferring the obtained homogeneous solution into a reaction kettle, placing the reaction kettle into an oven for reaction, separating black brown solid matters from the reaction solution by using a permanent magnet, and cleaning to obtain Fe 3O4 nano particles with superparamagnetism;
step two: preparation of magnetic organic framework MCOF
Dissolving 1,3, 5-tri (4-aminophenyl) benzene and 2, 5-dimethoxybenzene-1, 4-dicarboxaldehyde in 1, 4-dioxane and n-butanol, fully dissolving, adding Fe 3O4 nano particles prepared in the step one, dispersing, adding acetic acid for reaction, separating brown solid substances from the reaction solution by using a permanent magnet, and cleaning to obtain MCOF magnetic nano particles;
step three: preparation of MCOF-AuNPs nanocomposite
Adding the chlorauric acid trihydrate solution into MCOF magnetic nano particles obtained in the second step, completely and uniformly mixing, adding a trisodium citrate solution for reaction, separating brown solid matters from the reaction solution by using a permanent magnet, and cleaning to obtain a MCOF-AuNPs nano composite material;
Step four: preparation of apt-MCOF-AuNPs nanoprobe
Adding a tris (2-carbonyl ethyl) phosphorus hydrochloride solution into a salmonella aptamer aqueous solution, carrying out mixed spinning at room temperature to obtain an activated aptamer solution, adding MCOF-AuNPs nano composite material obtained in the step three into the activated aptamer solution, and carrying out mixed spinning at room temperature overnight to obtain an apt-MCOF-AuNPs nano probe;
Step five: preparation of the quencher
Adding FeCl 3·6H2 O into double distilled water, uniformly mixing to obtain Fe 3+ quencher, and sealing and keeping in dark for later use.
Preferably, in the first step, the mass ratio of FeCl 3·6H2 O to CH 3COONa·3H2 O is 1.2-3:3.2-8.
Preferably, in the second step, the mass ratio of the 1,3, 5-tri (4-aminophenyl) benzene, the 2, 5-dimethoxybenzene-1, 4-dicarboxaldehyde and the Fe 3O4 nano particles is 102-106:86-88:115-120.
Preferably, in the second step, the volume ratio of the 1, 4-dioxane to the n-butanol is 0.8-1:1-1.2.
Preferably, in the third step, the concentration of the aqueous chloroauric acid solution is 1-1.2mM, and the concentration of the trisodium citrate solution is 100-120mM.
Preferably, in the third step, the volume mass ratio of the chloroauric acid trihydrate solution, the trisodium citrate solution and the MCOF nano particles is 35-40mL:1.5-1.6mL:30-32mg.
Preferably, in step four, the volume μl of salmonella aptamer aqueous solution: the mass mg of MCOF-AuNPs nano composite material is 9-10:0.18-0.2.
Preferably, in the fifth step, the volume of double distilled water mL: the mass mg of FeCl 3·6H2 O is 95-100:25-27.
On the other hand, the invention also provides a kit which is prepared by adopting the preparation method.
Compared with the prior art, the invention has the following beneficial effects:
According to the preparation method of the kit for detecting salmonella typhimurium, provided by the invention, magnetic beads with superparamagnetism are firstly prepared, the magnetic beads are used as magnetic cores, organic covalent frameworks (COFs) are coated to form nuclear shell structures, namely MCOF nanometer magnetic particles, then gold nanometer particles are loaded on MCOF nanometer magnetic particles by an in-situ reduction method, and finally the gold nanometer particles are coupled with an aptamer of salmonella typhimurium to synthesize the high-fluorescence-emission nanoprobe apt-MCOF-AuNPs. The salmonella typhimurium and the nano-probe are incubated together to form a probe-thallus complex, a quencher Fe 3+ is added to quench the fluorescence of the probe, and the fluorescence of the probe is not quenched by Fe 3+ due to the protective effect of the thallus on the nano-probe and the competitive combination of the thallus and Fe 3+, so that the solution is bright green fluorescence under 365nm ultraviolet light irradiation. Thereby realizing the rapid and specific detection of salmonella typhimurium.
The invention utilizes the nanocomposite apt-MCOF-AuNPs to detect the salmonella typhimurium, utilizes the quenching inhibition effect to realize fluorescence detection, shortens the detection time, has small variation coefficient during quantitative detection, has the lowest detection concentration of 4CFU/mL, has the recovery rate of a simulation sample of 97.38-104.56%, has high sensitivity and good stability, and has certain applicability. Image analysis is realized by combining a smart phone color analysis APP (e.g. ColorColl) to obtain semi-quantitative detection results.
Drawings
FIG. 1 is a graph of a salmonella detection standard;
FIG. 2 is a flow chart of the detection of Salmonella typhimurium by the apt-MCOF-AuNPs provided by the invention.
Detailed Description
The invention provides a preparation method of a kit for detecting salmonella typhimurium, which comprises the following steps:
Step one: the Fe 3O4 nano-particles with superparamagnetism are prepared by the following preparation method:
Preferably, the mass ratio is 1.35:3.6 FeCl 3·6H2 O and CH 3COONa·3H2 O are magnetically stirred in ethylene glycol at room temperature until complete dissolution, and the resulting homogeneous solution is transferred to a reaction kettle, preferably in an oven at 200 ℃ for 16h; separating black brown solid substances from the reaction solution by using a permanent magnet, and alternately cleaning for 3-5 times by using ultrapure water and absolute ethyl alcohol to obtain Fe 3O4 nano particles with superparamagnetism;
step two: preparation of magnetic organic framework MCOF
The Fe 3O4 nano-particles prepared in the first step are used as magnetic cores, and coated with organic covalent frameworks (COFs) to form core-shell structures, so that MCOF magnetic nano-particles are obtained, and the preferable preparation method is as follows:
Dissolving 1,3, 5-tri (4-aminophenyl) benzene and 2, 5-dimethoxybenzene-1, 4-dicarboxaldehyde in 1, 4-dioxane and n-butanol, adding Fe 3O4 nano particles prepared in the step one to disperse the nano particles, adding acetic acid, stirring the mixture for 48 hours preferably at 70 ℃, separating green solid matters from a reaction solution by using a permanent magnet, alternately washing MCOF magnetic nano particles for multiple times by using acetone and tetrahydrofuran to remove unreacted matters, finally drying the mixture to obtain Fe 3O4 @Ag magnetic nano particles, and keeping the Fe 3O4 @Ag magnetic nano particles away from light for standby at 4 ℃;
Wherein, the mass ratio of the 1,3, 5-tri (4-aminophenyl) benzene, the 2, 5-dimethoxybenzene-1, 4-dicarboxaldehyde and the Fe 3O4 nano particles is preferably 105.4:87.4:120;
wherein, the volume ratio of the 1, 4-dioxane to the n-butanol is preferably 1:1;
step three: preparation of MCOF-AuNPs nanocomposite
Gold nanoparticles are loaded on MCOF magnetic nanoparticles prepared in the second step to obtain a MCOF-AuNPs nanocomposite, and the preferred preparation method is as follows:
Adding the solution of chloroauric acid trihydrate to MCOF obtained in the second step, completely mixing, adding the solution of trisodium citrate, and stirring and reacting for 40min at 20 ℃ preferably. Separating brown solid substances from the reaction solution by using a permanent magnet, washing MCOF-AuNPs nano composite materials by using double distilled water for multiple times to remove unreacted substances, and finally drying to obtain MCOF-AuNPs nano composite materials, and storing the MCOF-AuNPs nano composite materials at a temperature of 4 ℃ in a dark place for later use;
The concentration of chloroauric acid trihydrate solution is preferably 1mM, and the concentration of trisodium citrate solution is preferably 100mM; the volume mass ratio of chloroauric acid trihydrate solution, trisodium citrate solution and MCOF nano particles is preferably 40mL:1.6mL:32mg;
Step four: preparation of apt-MCOF-AuNPs nanoprobe
Coupling MCOF-AuNPs nanocomposite obtained in the step three with an aptamer of Salmonella typhimurium to synthesize a nanoprobe apt-MCOF-AuNPs with high fluorescence emission, wherein the preparation method is preferably as follows:
Adding a tris (2-carbonyl ethyl) phosphorus hydrochloride solution into a salmonella aptamer aqueous solution, carrying out mixing and spinning at room temperature for preferably 1h, activating a modified sulfhydryl group on the aptamer to obtain an activated aptamer solution, taking MCOF-AuNPs nanocomposite, preferably washing with distilled water for 2 times, resuspending with distilled water, adding the activated aptamer, carrying out mixing and spinning at room temperature overnight, carrying out magnetic separation, removing supernatant, washing with distilled water for three times to obtain apt-MCOF-AuNPs, and storing at 4 ℃ in a dark place for later use;
Wherein, the concentration of the tris (2-carbonyl ethyl) phosphorus hydrochloride solution is preferably 1mM, the concentration of the salmonella aptamer aqueous solution is preferably 10 mu M, and the volume ratio of the tris (2-carbonyl ethyl) phosphorus hydrochloride solution to the salmonella aptamer aqueous solution is preferably 1: volume μl of salmonella aptamer aqueous solution: the mass mg of MCOF-AuNPs nanoparticles is preferably 10:0.2;
Wherein, the size of the apt-MCOF-AuNPs nano probe is preferably 300nm;
The salmonella-resistant aptamer provided by the invention has the sequence as follows: 5'-SH- (CH 2)6 -TAT GGC GGC GTC ACC CGA CGG GGA CTT GAC CTT GACATTATGACA G-3', synthesized by Shanghai Biotechnology;
Step five: the preparation method of the Fe 3+ quencher is to store in a sealed and light-proof way for standby, and the preferred preparation method is as follows:
Adding FeCl 3·6H2 O into double distilled water, uniformly mixing to obtain Fe 3+ quencher, and sealing and keeping in dark for later use.
Wherein, double distilled water volume mL: feCl 3·6H2 O mass mg is 100:27.
The technical scheme of the present invention will be explained in detail with reference to specific examples.
Example 1
Step one: preparation of Fe 3O4 nanoparticles with superparamagnetism
First, fe 3O4 was prepared by a hydrothermal method, and 1.35g of FeCl 3·6H2 O and 3.6g of CH 3COONa·3H2 O were magnetically stirred in 40mL of ethylene glycol at room temperature until they were completely dissolved. Then transferring the obtained homogeneous solution into a reaction kettle, and reacting for 16 hours in a baking oven at 200 ℃; separating black brown solid substances from the reaction solution by using a permanent magnet, and alternately cleaning for 3 times by using ultrapure water and absolute ethyl alcohol to obtain Fe 3O4 nano particles with superparamagnetism;
step two: preparation of magnetic organic framework MCOF
Preparation MCOF process: 105.4mg of 1,3, 5-tris (4-aminophenyl) benzene and 87.4mg of 2, 5-dimethoxybenzene-1, 4-dicarboxaldehyde are dissolved in a mixed solution containing 20mL of 1, 4-dioxane and 20mL of n-butanol, after the complete dissolution, 120mg of Fe 3O4 nano-particles prepared are added to disperse the nano-particles therein, 0.5mL of 12M acetic acid is added, preferably, the mixture is kept stand at 70 ℃ for 48 hours, green solid matters are separated from the reaction solution by using permanent magnets, MCOF magnetic nano-particles are alternately washed for multiple times by using acetone and tetrahydrofuran to remove unreacted matters, and finally, the mixture is dried;
step three: preparation of MCOF-AuNPs nanocomposite
MCOF-AuNPs were prepared by in situ reduction, 32mg of MCOF was added to 40mL of 1mM HAuCl 4 solution, dissolved completely, 1.6mL of 100mM trisodium citrate solution was added, and the reaction was stirred at 20℃for 40min. Separating brown solid substances from the reaction solution by using a permanent magnet, washing MCOF-AuNPs nano composite materials by using double distilled water for multiple times to remove unreacted substances, and finally drying to obtain MCOF-AuNPs nano composite materials, and storing the MCOF-AuNPs nano composite materials at a temperature of 4 ℃ in a dark place for later use;
Step four: preparation of apt-MCOF-AuNPs nanoprobe
1. Mu.L of 1mM tris (2-carboxyethyl) phosphorus hydrochloride solution was added to 10. Mu.L of 10. Mu.M Salmonella aptamer aqueous solution, and the mixture was spun at room temperature for 1 hour, the thiol group modified on the aptamer was activated, 0.2mg MCOF-AuNPs nanocomposite was taken, washed 2 times with distilled water, resuspended in 1mL distilled water, and the activated aptamer was added and spun at room temperature overnight. Magnetic separation, supernatant removal and washing with distilled water for three times to obtain the apt-MCOF-AuNPs nano probe, and preserving the nano probe at 4 ℃ in a dark place for later use, wherein the aptamer sequence of salmonella used in the method is as follows: 5'-SH- (CH 2)6 -TAT GGC GGC GTC ACC CGA CGG GGA CTT GAC CTT GACATTATGACA G-3') was synthesized by Shanghai Biochemical Co.
Step five: preparation of the quencher (Fe 3+ standard solution)
27Mg of FeCl 3·6H2 O is weighed and dissolved in 100mL of double distilled water to obtain the Fe 3+ quencher, and the quencher is stored in a sealed and light-proof manner for standby.
Examples 2-4 were prepared in the same manner as in example 1 with the following parameter configurations in Table 1
Table 1 parameter configurations of examples 2-4
The kits prepared in examples 1-4 above were tested as follows.
Preparation of fungus liquid standard substance
Storing strain at-80deg.C, activating the strain, streaking and purifying the activated strain on 3% sodium chloride tryptic vein agar plate, culturing at 37deg.C for 18-24 hr, selecting single colony, inoculating 3% sodium chloride tryptic liquid culture medium, and shake culturing at 37deg.C for 12-18 hr. Viable bacteria were counted by a dilution plate decanting method with a 10-fold ratio of 1mL of bacterial liquid. Inactivating the rest bacterial liquid with 1% formaldehyde at room temperature for 10min, centrifuging the inactivated bacterial liquid at 3000rpm for 3min, collecting bacterial cells, mixing with PBS solution, preparing bacterial suspension, regulating bacterial suspension concentration to 10 9CFU·mL-1 with PBS solution, and storing in a refrigerator at 4deg.C for use.
Method for detecting salmonella typhimurium based on apt-MCOF-AuNPs and quencher
The detection flow is shown in FIG. 2, 800. Mu.L of the sample to be detected and 200. Mu.L of 2mg/mL apt-MCOF-AuNPs of examples 1-4 are respectively taken for testing, and the mixture is spun for 30min at room temperature and in a dark place. 200. Mu.L of 1mM Fe 3+ was added and reacted for 3min. The method comprises the steps of irradiating 365nm ultraviolet light in a darkroom, photographing by a smart phone, performing HSV analysis by using a color analysis APP, outputting three channel values of Hue (H), saturation (S) and Value (brightness, V), and taking the Hue Value as an output quantitative signal of the concentration of salmonella typhimurium. Referring to the standard curve made in FIG. 1, the amount of Salmonella typhimurium in the sample was determined. The concentration range of the bacteria is 10-10 7 CFU/mL.
FIG. 1 is a graph of the above salmonella detection criteria, wherein the abscissa represents the concentration logarithmic value of Salmonella typhimurium and the ordinate represents the tone value. Detection concentration range: 10-10 7 CFU/mL.
The method has the advantages of stable detection, low detection limit of 4CFU/ml, short detection time, simple operation and good detection effect. The salmonella typhimurium, listeria monocytogenes, staphylococcus aureus, vibrio parahaemolyticus, escherichia coli O157:H27 and the like were tested, and only the salmonella test results in examples 1-4 were positive, the rest were negative, the method was strong in specificity, no false positive and false negative results were found, and the results are shown in tables 2-5.
TABLE 2 specific detection results of example 1 on Salmonella
TABLE 3 results of specific detection of Salmonella in example 2
TABLE 4 example 3 results of specific detection of Salmonella
TABLE 5 results of specific detection of Salmonella in example 4
Note that: the positive and negative of Salmonella typhimurium are shown.
Detection of simulated samples
0.5G of chicken was ground and mixed with 50mL of sterile PBS to form a homogenate. 1 ml of milk and egg liquid were diluted 100-fold with sterile PBS and the filtrate was suction filtered through a 0.22 μm filter. Then, each of the actual samples was inoculated with Salmonella typhimurium for culture, and after dilution, plate counting was performed and detection and comparison were performed by the method of the present invention.
Taking 800 mu L of food sample to be tested and 200 mu L of apt-MCOF-AuNPs in examples 1-4, respectively testing, and mixing and rotating for 30min at room temperature in a dark place. Fe 3+ was added and the reaction was carried out for 3min. Irradiation with 365nm uv light in darkroom, smart phone photographing, HSV analysis with APP developed independently, and Hue value (Hue value) as output quantitative signal of salmonella typhimurium concentration. The plate counting result is compared with the result of the method according to the invention, and referring to Table 6, the method according to the invention has stable detection and the standard adding recovery rate reaches 97.38% -104.56%.
TABLE 6 example 1 results of detection of Salmonella typhimurium in food samples
Table 7 example 2 results of detecting salmonella typhimurium in food samples
TABLE 8 example 3 results of detection of Salmonella typhimurium in food samples
Table 9 example 4 results of detecting salmonella typhimurium in food samples
* Log C: concentration (CFU/mL) log of Salmonella typhimurium.
Comparative example 1
Table 10 comparison of the kit with commercially available ELISA kits
| Detection time | Detection device | Sensitivity of | Coefficient of variation | |
| Example 1 | About 33 minutes | Smart phone | 4CFU/mL | Less than 6% |
| Example 2 | About 33 minutes | Smart phone | 9CFU/mL | Less than 6% |
| Example 3 | About 33 minutes | Smart phone | 12CFU/mL | Less than 9% |
| Example 4 | About 33 minutes | Smart phone | 6CFU/mL | Less than 10% |
| ELISA kit for market | For more than 2.5 hours | Precise enzyme label instrument | 10-104CFU/mL | Less than 10-15% |
The above description is only a preferred embodiment of the present invention, but the scope of the present invention is not limited thereto. Any person skilled in the art can make equivalent substitutions or changes according to the technical scheme of the present invention and the improved conception thereof within the technical scope of the present disclosure; are intended to be encompassed within the scope of the present invention.
Claims (10)
1. The preparation method of the kit for detecting salmonella typhimurium is characterized by comprising the following steps of:
Step one: preparing Fe 3O4 nano particles with superparamagnetism;
step two: preparation of magnetic organic framework MCOF
Taking the Fe 3O4 nano-particles prepared in the first step as a magnetic core, coating an organic covalent framework (COF) to form a core-shell structure, and obtaining MCOF magnetic nano-particles;
step three: preparation of MCOF-AuNPs nanocomposite
Loading gold nanoparticles on MCOF magnetic nanoparticles prepared in the second step to obtain a MCOF-AuNPs nanocomposite;
Step four: preparation of apt-MCOF-AuNPs nanoprobe
Coupling MCOF-AuNPs nano composite material obtained in the step three with an aptamer of salmonella typhimurium to synthesize a nano probe apt-MCOF-AuNPs with high fluorescence emission;
Step five: preparing Fe 3+ quencher, sealing and keeping away from light for standby.
2. The method for preparing the kit for detecting salmonella typhimurium according to claim 1, wherein the method comprises the following specific steps:
step one: preparation of Fe 3O4 nanoparticles
Magnetically stirring FeCl 3·6H2O、CH3COONa·3H2 O in ethylene glycol at room temperature until the FeCl 3·6H2O、CH3COONa·3H2 O is completely dissolved, transferring the obtained homogeneous solution into a reaction kettle, placing the reaction kettle into an oven for reaction, separating black brown solid matters from the reaction solution by using a permanent magnet, and cleaning to obtain Fe 3O4 nano particles with superparamagnetism;
step two: preparation of magnetic organic framework MCOF
Dissolving 1,3, 5-tri (4-aminophenyl) benzene and 2, 5-dimethoxybenzene-1, 4-dicarboxaldehyde in 1, 4-dioxane and n-butanol, fully dissolving, adding Fe 3O4 nano particles prepared in the step one, dispersing, adding acetic acid for reaction, separating brown solid substances from the reaction solution by using a permanent magnet, and cleaning to obtain MCOF magnetic nano particles;
step three: preparation of MCOF-AuNPs nanocomposite
Adding the chlorauric acid trihydrate solution into MCOF magnetic nano particles obtained in the second step, completely and uniformly mixing, adding a trisodium citrate solution for reaction, separating brown solid matters from the reaction solution by using a permanent magnet, and cleaning to obtain a MCOF-AuNPs nano composite material;
Step four: preparation of apt-MCOF-AuNPs nanoprobe
Adding a tris (2-carbonyl ethyl) phosphorus hydrochloride solution into a salmonella aptamer aqueous solution, carrying out mixed spinning at room temperature to obtain an activated aptamer solution, adding MCOF-AuNPs nano composite material obtained in the step three into the activated aptamer solution, and carrying out mixed spinning at room temperature overnight to obtain an apt-MCOF-AuNPs nano probe;
Step five: preparation of the quencher
Adding FeCl 3·6H2 O into double distilled water, uniformly mixing to obtain Fe 3+ quencher, and sealing and keeping in dark for later use.
3. The method for preparing a kit for detecting salmonella typhimurium according to claim 2, wherein in the first step, the mass ratio of FeCl 3·6H2 O to CH 3COONa·3H2 O is 1.2-3:3.2-8.
4. The method for preparing the kit for detecting salmonella typhimurium according to claim 2, wherein in the second step, the mass ratio of the 1,3, 5-tris (4-aminophenyl) benzene, 2, 5-dimethoxybenzene-1, 4-dicarboxaldehyde and Fe 3O4 nano particles is 102-106:86-88:115-120.
5. The method for preparing a kit for detecting salmonella typhimurium according to claim 2, wherein in the second step, the volume ratio of 1, 4-dioxane to n-butanol is 0.8-1:1-1.2.
6. The method for preparing a kit for detecting salmonella typhimurium according to claim 2, wherein in the third step, the concentration of the aqueous chloroauric acid solution is 1-1.2mM, and the concentration of the trisodium citrate solution is 100-120mM.
7. The method for preparing the kit for detecting salmonella typhimurium according to claim 2, wherein in the third step, the volume mass ratio of the chloroauric acid hydrate solution, the trisodium citrate solution and the MCOF magnetic nanoparticles is 35-40mL:1.5-1.6mL:30-32mg.
8. The method for preparing a kit for detecting salmonella typhimurium according to claim 2, wherein in the fourth step, the volume μl of the salmonella aptamer aqueous solution is: the mass mg of MCOF-AuNPs nano composite material is 9-10:0.18-0.2.
9. The method for preparing a kit for detecting salmonella typhimurium according to claim 2, wherein in the fifth step, the volume of double distilled water is mL: the mass mg of FeCl 3·6H2 O is 95-100:25-27.
10. A kit prepared by the method of any one of claims 1-9.
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Citations (17)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104857933A (en) * | 2015-05-18 | 2015-08-26 | 苏州汇通色谱分离纯化有限公司 | Preparation and application of core-shell type magnetic metal organic framework nano-particles |
| JP2016144445A (en) * | 2015-01-28 | 2016-08-12 | ユニバーサル・バイオ・リサーチ株式会社 | Automatic bacteria detecting apparatus and method thereof |
| CN105866079A (en) * | 2016-03-31 | 2016-08-17 | 同济大学 | A molecular logic gate method building based on interactions among a fluorochrome NMM, G-quadruplex DNA, crown ether and metal ions |
| CN105920620A (en) * | 2016-06-21 | 2016-09-07 | 东南大学 | Magnetic fluorescent multimodal nano biological probe as well as preparation method and application thereof |
| CN109239037A (en) * | 2018-09-28 | 2019-01-18 | 长沙理工大学 | Biosensor and its preparation method and application based on MOFs as energy acceptor |
| CN109298180A (en) * | 2018-11-19 | 2019-02-01 | 中南大学 | A method of detection salmonella typhimurium |
| CN111024943A (en) * | 2019-12-20 | 2020-04-17 | 吉林农业大学 | Switch-on/off type composite fluorescent nano probe for rapid detection of salmonella and preparation method thereof |
| CN111235229A (en) * | 2020-01-15 | 2020-06-05 | 徐州医科大学 | Ratiometric fluorescent probe for detecting PTK7 and preparation method thereof |
| CN111366727A (en) * | 2020-03-19 | 2020-07-03 | 吉林大学 | Kit for detecting salmonella typhimurium and preparation method thereof |
| WO2021104435A1 (en) * | 2019-11-30 | 2021-06-03 | 康码（上海）生物科技有限公司 | Biomagnetic microsphere and preparation method therefor and use thereof |
| WO2021145796A2 (en) * | 2020-01-14 | 2021-07-22 | Максим Петрович НИКИТИН | Method of determining analyte content in a sample with the aid of a nanoparticle-based agent and a recognition oligonucleotide |
| CN113514435A (en) * | 2021-04-25 | 2021-10-19 | 重庆医科大学 | Based on UCNPs and GO @ Fe3O4Fluorescent biosensor for immediately detecting escherichia coli |
| CN113567521A (en) * | 2021-07-14 | 2021-10-29 | 山西大学 | Magnetic COF surface molecularly imprinted electrochemical sensor and preparation method and application thereof |
| CN113624967A (en) * | 2021-08-11 | 2021-11-09 | 郑州轻工业大学 | Magnetic MOF @ aptamer and method for detecting food-borne pathogenic bacteria by using same |
| CN113663646A (en) * | 2021-07-19 | 2021-11-19 | 中国计量大学 | Preparation method and application of wide-spectrum specific antibody modified magnetic metal organic framework material |
| CN114381553A (en) * | 2022-01-27 | 2022-04-22 | 云南大学 | Biological material and kit for detecting African swine fever virus and detection method of African swine fever virus for non-diagnosis purpose |
| CN114720437A (en) * | 2022-03-01 | 2022-07-08 | 江西省科学院微生物研究所(江西省流域生态研究所) | Fe-PDAN-based efficient fluorescence quenching test strip and preparation method and detection method thereof |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20150024376A1 (en) * | 2013-07-19 | 2015-01-22 | Battelle Memorial Institute | Trap and flow system and process for capture of target analytes |
| US20180274013A1 (en) * | 2015-09-23 | 2018-09-27 | Nanyang Technological University | Metal-organic framework nanosheet |
-
2023
- 2023-01-04 CN CN202310008162.9A patent/CN115856297B/en active Active
Patent Citations (17)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2016144445A (en) * | 2015-01-28 | 2016-08-12 | ユニバーサル・バイオ・リサーチ株式会社 | Automatic bacteria detecting apparatus and method thereof |
| CN104857933A (en) * | 2015-05-18 | 2015-08-26 | 苏州汇通色谱分离纯化有限公司 | Preparation and application of core-shell type magnetic metal organic framework nano-particles |
| CN105866079A (en) * | 2016-03-31 | 2016-08-17 | 同济大学 | A molecular logic gate method building based on interactions among a fluorochrome NMM, G-quadruplex DNA, crown ether and metal ions |
| CN105920620A (en) * | 2016-06-21 | 2016-09-07 | 东南大学 | Magnetic fluorescent multimodal nano biological probe as well as preparation method and application thereof |
| CN109239037A (en) * | 2018-09-28 | 2019-01-18 | 长沙理工大学 | Biosensor and its preparation method and application based on MOFs as energy acceptor |
| CN109298180A (en) * | 2018-11-19 | 2019-02-01 | 中南大学 | A method of detection salmonella typhimurium |
| WO2021104435A1 (en) * | 2019-11-30 | 2021-06-03 | 康码（上海）生物科技有限公司 | Biomagnetic microsphere and preparation method therefor and use thereof |
| CN111024943A (en) * | 2019-12-20 | 2020-04-17 | 吉林农业大学 | Switch-on/off type composite fluorescent nano probe for rapid detection of salmonella and preparation method thereof |
| WO2021145796A2 (en) * | 2020-01-14 | 2021-07-22 | Максим Петрович НИКИТИН | Method of determining analyte content in a sample with the aid of a nanoparticle-based agent and a recognition oligonucleotide |
| CN111235229A (en) * | 2020-01-15 | 2020-06-05 | 徐州医科大学 | Ratiometric fluorescent probe for detecting PTK7 and preparation method thereof |
| CN111366727A (en) * | 2020-03-19 | 2020-07-03 | 吉林大学 | Kit for detecting salmonella typhimurium and preparation method thereof |
| CN113514435A (en) * | 2021-04-25 | 2021-10-19 | 重庆医科大学 | Based on UCNPs and GO @ Fe3O4Fluorescent biosensor for immediately detecting escherichia coli |
| CN113567521A (en) * | 2021-07-14 | 2021-10-29 | 山西大学 | Magnetic COF surface molecularly imprinted electrochemical sensor and preparation method and application thereof |
| CN113663646A (en) * | 2021-07-19 | 2021-11-19 | 中国计量大学 | Preparation method and application of wide-spectrum specific antibody modified magnetic metal organic framework material |
| CN113624967A (en) * | 2021-08-11 | 2021-11-09 | 郑州轻工业大学 | Magnetic MOF @ aptamer and method for detecting food-borne pathogenic bacteria by using same |
| CN114381553A (en) * | 2022-01-27 | 2022-04-22 | 云南大学 | Biological material and kit for detecting African swine fever virus and detection method of African swine fever virus for non-diagnosis purpose |
| CN114720437A (en) * | 2022-03-01 | 2022-07-08 | 江西省科学院微生物研究所(江西省流域生态研究所) | Fe-PDAN-based efficient fluorescence quenching test strip and preparation method and detection method thereof |
Non-Patent Citations (5)
| Title |
|---|
| A fluorescence biosensor for Salmonella typhimurium detection in food based on the nanoself- assembly of alendronic acid modified upconversion and gold nanoparticles;Min Chen;《The Royal Society of Chemistry》;20211231;全文 * |
| A Novel Fluorescence Aptasensor Based on Magnetic Beads/Gold Nanoparticles/DNA-Stabilized Silver Nanoclusters for Detection of Salmonella Typhimurium;Shiqian Fu;《foods》;20220218;全文 * |
| A portable smartphone-assisted highly emissive magnetic covalent organic framework-based fluorescence sensor for the detection of Salmonella typhimurium;Shengnan Weia;《Sensors and Actuators: B. Chemical》;20230602;全文 * |
| 基于纳米粒子荧光传感器的食源性病原菌检测技术研究;吴静;李耘;王天琪;任亚林;;农产品质量与安全;20171210(06);全文 * |
| 沙门氏菌检测方法研究综述;蓝兰;吕素玲;李秀桂;;广西畜牧兽医;20181115(06);全文 * |
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