CN117110607B - Composition for detecting vibrio parahaemolyticus and application thereof - Google Patents
Composition for detecting vibrio parahaemolyticus and application thereof Download PDFInfo
- Publication number
- CN117110607B CN117110607B CN202311195101.4A CN202311195101A CN117110607B CN 117110607 B CN117110607 B CN 117110607B CN 202311195101 A CN202311195101 A CN 202311195101A CN 117110607 B CN117110607 B CN 117110607B
- Authority
- CN
- China
- Prior art keywords
- vibrio parahaemolyticus
- reaction
- composition
- concentration
- solution
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 241000607272 Vibrio parahaemolyticus Species 0.000 title claims abstract description 62
- 239000000203 mixture Substances 0.000 title claims abstract description 28
- 239000000523 sample Substances 0.000 claims abstract description 32
- 238000001514 detection method Methods 0.000 claims abstract description 22
- 238000006243 chemical reaction Methods 0.000 claims abstract description 20
- GEYOCULIXLDCMW-UHFFFAOYSA-N 1,2-phenylenediamine Chemical compound NC1=CC=CC=C1N GEYOCULIXLDCMW-UHFFFAOYSA-N 0.000 claims abstract description 15
- 239000000243 solution Substances 0.000 claims description 40
- 239000011572 manganese Substances 0.000 claims description 32
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 30
- 238000001035 drying Methods 0.000 claims description 21
- 108090000790 Enzymes Proteins 0.000 claims description 20
- 102000004190 Enzymes Human genes 0.000 claims description 20
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 claims description 18
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims description 17
- 229940098773 bovine serum albumin Drugs 0.000 claims description 17
- 238000002156 mixing Methods 0.000 claims description 16
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 15
- 239000002122 magnetic nanoparticle Substances 0.000 claims description 14
- 238000003756 stirring Methods 0.000 claims description 12
- LMDZBCPBFSXMTL-UHFFFAOYSA-N 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide Chemical compound CCN=C=NCCCN(C)C LMDZBCPBFSXMTL-UHFFFAOYSA-N 0.000 claims description 10
- 238000002835 absorbance Methods 0.000 claims description 10
- 238000000034 method Methods 0.000 claims description 10
- NQTADLQHYWFPDB-UHFFFAOYSA-N N-Hydroxysuccinimide Chemical compound ON1C(=O)CCC1=O NQTADLQHYWFPDB-UHFFFAOYSA-N 0.000 claims description 9
- 230000004913 activation Effects 0.000 claims description 8
- 229910021380 Manganese Chloride Inorganic materials 0.000 claims description 7
- GLFNIEUTAYBVOC-UHFFFAOYSA-L Manganese chloride Chemical compound Cl[Mn]Cl GLFNIEUTAYBVOC-UHFFFAOYSA-L 0.000 claims description 7
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 claims description 7
- ZOIORXHNWRGPMV-UHFFFAOYSA-N acetic acid;zinc Chemical compound [Zn].CC(O)=O.CC(O)=O ZOIORXHNWRGPMV-UHFFFAOYSA-N 0.000 claims description 7
- RBTARNINKXHZNM-UHFFFAOYSA-K iron trichloride Chemical compound Cl[Fe](Cl)Cl RBTARNINKXHZNM-UHFFFAOYSA-K 0.000 claims description 7
- 239000011565 manganese chloride Substances 0.000 claims description 7
- 229940099607 manganese chloride Drugs 0.000 claims description 7
- 235000002867 manganese chloride Nutrition 0.000 claims description 7
- 229940093429 polyethylene glycol 6000 Drugs 0.000 claims description 7
- 230000035484 reaction time Effects 0.000 claims description 7
- 239000001632 sodium acetate Substances 0.000 claims description 7
- 235000017281 sodium acetate Nutrition 0.000 claims description 7
- 239000001509 sodium citrate Substances 0.000 claims description 7
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 claims description 7
- 239000004246 zinc acetate Substances 0.000 claims description 7
- 108091023037 Aptamer Proteins 0.000 claims description 6
- 239000007795 chemical reaction product Substances 0.000 claims description 6
- 239000011259 mixed solution Substances 0.000 claims description 6
- 238000010517 secondary reaction Methods 0.000 claims description 6
- 238000012417 linear regression Methods 0.000 claims description 5
- 238000002360 preparation method Methods 0.000 claims description 5
- 238000001179 sorption measurement Methods 0.000 claims description 5
- 238000005119 centrifugation Methods 0.000 claims description 4
- 239000002244 precipitate Substances 0.000 claims description 4
- 230000003213 activating effect Effects 0.000 claims description 3
- WGCNASOHLSPBMP-UHFFFAOYSA-N hydroxyacetaldehyde Natural products OCC=O WGCNASOHLSPBMP-UHFFFAOYSA-N 0.000 claims description 3
- -1 aminopropyl Chemical group 0.000 claims 1
- 230000000007 visual effect Effects 0.000 abstract description 8
- 238000004737 colorimetric analysis Methods 0.000 abstract description 5
- 238000011084 recovery Methods 0.000 abstract description 5
- 230000035945 sensitivity Effects 0.000 abstract description 4
- 239000000758 substrate Substances 0.000 abstract description 4
- 230000009977 dual effect Effects 0.000 abstract description 3
- 244000005700 microbiome Species 0.000 abstract description 2
- 241000894006 Bacteria Species 0.000 description 14
- 235000013305 food Nutrition 0.000 description 8
- 230000015572 biosynthetic process Effects 0.000 description 4
- 239000003086 colorant Substances 0.000 description 4
- 238000003786 synthesis reaction Methods 0.000 description 4
- 208000019331 Foodborne disease Diseases 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 238000012512 characterization method Methods 0.000 description 3
- 238000000227 grinding Methods 0.000 description 3
- 238000007789 sealing Methods 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- 206010016952 Food poisoning Diseases 0.000 description 2
- 241000186779 Listeria monocytogenes Species 0.000 description 2
- 241000293869 Salmonella enterica subsp. enterica serovar Typhimurium Species 0.000 description 2
- 241000191967 Staphylococcus aureus Species 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 230000002452 interceptive effect Effects 0.000 description 2
- 238000007885 magnetic separation Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 238000000870 ultraviolet spectroscopy Methods 0.000 description 2
- 241001474374 Blennius Species 0.000 description 1
- 241001646719 Escherichia coli O157:H7 Species 0.000 description 1
- 208000005577 Gastroenteritis Diseases 0.000 description 1
- 241000269821 Scombridae Species 0.000 description 1
- 206010040047 Sepsis Diseases 0.000 description 1
- 208000012873 acute gastroenteritis Diseases 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 244000052616 bacterial pathogen Species 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000004186 food analysis Methods 0.000 description 1
- 244000078673 foodborn pathogen Species 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 235000020640 mackerel Nutrition 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000002114 nanocomposite Substances 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 230000005180 public health Effects 0.000 description 1
- 235000014102 seafood Nutrition 0.000 description 1
- 208000013223 septicemia Diseases 0.000 description 1
- 238000004088 simulation Methods 0.000 description 1
- 238000011895 specific detection Methods 0.000 description 1
- 229910001220 stainless steel Inorganic materials 0.000 description 1
- 239000010935 stainless steel Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 238000009210 therapy by ultrasound Methods 0.000 description 1
- 238000002371 ultraviolet--visible spectrum Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56911—Bacteria
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
- G01N33/532—Production of labelled immunochemicals
- G01N33/535—Production of labelled immunochemicals with enzyme label or co-enzymes, co-factors, enzyme inhibitors or enzyme substrates
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
- G01N33/54326—Magnetic particles
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
- G01N33/54346—Nanoparticles
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/195—Assays involving biological materials from specific organisms or of a specific nature from bacteria
- G01N2333/28—Assays involving biological materials from specific organisms or of a specific nature from bacteria from Vibrionaceae (F)
-
- 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
Abstract
The invention relates to the technical field of microorganism detection. The invention provides a composition for detecting vibrio parahaemolyticus and application thereof, wherein the composition comprises Fe 3 O 4 -Zn-Mn-Apt probe, 3' 5-tetramethylbenzidine and o-phenylenediamine. The composition of the invention can be used for detecting vibrio parahaemolyticus, has higher sensitivity, the detection limit is 1.12cfu/mL, and the linear range is 0-1 multiplied by 10 4 cfu/mL，R 2 = 0.9934. As the concentration of Vibrio parahaemolyticus increases, the reaction solution changes from blue-green to green and then to yellow, so that semi-quantitative detection is carried out by visual observation, and the visual detection limit is 10cfu/mL. In addition, the developed dual substrate colorimetric method has good selectivity and recovery rate for the Vibrio parahaemolyticus for the determination of Vibrio parahaemolyticus.
Description
Technical Field
The invention relates to the technical field of microorganism detection, in particular to a composition for detecting vibrio parahaemolyticus and application thereof.
Background
In recent years, food safety problems caused by food-borne pathogens have become one of the public health problems of global public concern. Vibrio parahaemolyticus is a halophilic bacterium. The vibrio parahaemolyticus food poisoning is caused by eating food containing the strain, mainly comes from seafood, and in life practice, the occurrence rate of food-borne disease events caused by vibrio parahaemolyticus pollution is relatively high due to the effect of various factors. The diseases are food poisoning, acute gastroenteritis, septicemia, etc. Seriously threatens the health and life safety of people.
Therefore, it is important to develop a product capable of rapidly and specifically detecting Vibrio parahaemolyticus.
Disclosure of Invention
The invention aims to provide a composition for detecting vibrio parahaemolyticus and application thereof, and the composition can realize rapid and specific detection of the vibrio parahaemolyticus.
In order to achieve the above object, the present invention provides the following technical solutions:
the invention provides a composition for detecting vibrio parahaemolyticus, which is characterized in that the composition comprises Fe 3 O 4 -Zn-Mn-Apt probe, 3' 5-tetramethylbenzidine and o-phenylenediamine.
Preferably, the Fe 3 O 4 The preparation method of the Zn-Mn-Apt probe comprises the following steps:
(1) Mixing ferric trichloride, glycol, sodium acetate, sodium citrate and polyethylene glycol 6000, reacting, and drying to obtain Fe 3 O 4 Magnetic nanoparticles;
(2) Mixing manganese chloride, water and bovine serum albumin solution, stirring, adding sodium hydroxide solution for reaction, adding zinc acetate solution for secondary reaction, centrifuging, and drying the precipitate to obtain Zn-Mn nano enzyme;
(3) Zn-Mn nano enzyme and Fe 3 O 4 Mixing magnetic nano particles with water, stirring for reaction, collecting reaction products, and drying to obtain Fe 3 O 4 -Zn-Mn nanoenzyme;
(4) Fe with 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide and N-hydroxysuccinimide 3 O 4 -Zn-Mn nano enzyme activation site, fe after activating site 3 O 4 And mixing the Zn-Mn nano enzyme with the vibrio parahaemolyticus aptamer to obtain the Fe3O4-Zn-Mn-Apt probe.
Preferably, in the step (1), the dosage ratio of the ferric trichloride, the ethylene glycol, the sodium acetate, the sodium citrate and the polyethylene glycol 6000 is as follows: 18-22 mM: 18-22 mL: 70-80 mM: 30-40 mM: 0.18-0.22 g, wherein the temperature of the reaction in the step (1) is 190-210 ℃, the reaction time is 17-19 h, and the drying temperature is 55-65 ℃.
Preferably, the ratio of the manganese chloride, water and bovine serum albumin solution in the step (2) is 95-105 mM: 1.8-2.2 mL: 9-11 mL, wherein the concentration of the bovine serum albumin solution is 4-6 mg/mL, the stirring time in the step (2) is 25-35 min, the amount of the sodium hydroxide solution is the same as that of the bovine serum albumin solution, the concentration of the sodium hydroxide solution is 60-80 mM, the reaction time in the step (2) is 4-6 min, the amount of the zinc acetate solution is 1.8-2.2 times that of the bovine serum albumin solution, the secondary reaction time is 10-14 h, the centrifugation condition is 11000-13000 rpm, 13-17 min, and the drying temperature in the step (2) is 45-55 ℃.
Preferably, the Zn-Mn nanoenzyme and Fe in the step (3) 3 O 4 The dosage ratio of the magnetic nano particles to the water is 18-22 mg:0.18 to 0.22g: 45-55 mL, wherein the temperature of the stirring reaction is 55-65 ℃ and the time is 1.8-2.2 h, the reaction products are collected by magnetic adsorption, and the drying temperature in the step (3) is 55-65 ℃.
Preferably, the concentration of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in step (4) is 8-12 mg +.mL, the concentration of the N-hydroxysuccinimide is 8-12 mg/mL, the 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and Fe 3 O 4 The dosage ratio of the Zn-Mn nano enzyme is 1.8-2.2 mL: 1.8-2.2 mL:1mg, wherein the activation time is 25-35 min.
The invention also provides application of the composition in preparation of a vibrio parahaemolyticus detection kit.
The invention also provides a kit for detecting vibrio parahaemolyticus, which comprises the Fe as claimed in claim 1 3 O 4 -Zn-Mn-Apt probe, 3' 5-tetramethylbenzidine and o-phenylenediamine.
The invention also provides a using method of the kit, which comprises the following steps: sample to be measured, fe 3 O 4 Mixing a Zn-Mn-Apt probe, 3', 5-tetramethylbenzidine and o-phenylenediamine to obtain a mixed solution, and detecting the absorbance of the mixed solution at 652 nm;
the linear regression equation is adopted as Y= -0.2215X+0.9405, R 2 = 0.9934 to calculate the concentration of vibrio parahaemolyticus in the sample to be tested, where Y is absorbance, X is LogC, and C is the concentration of vibrio parahaemolyticus in cfu/mL.
The invention provides a composition for detecting vibrio parahaemolyticus and application thereof, wherein the composition comprises Fe 3 O 4 -Zn-Mn-Apt probe, 3' 5-tetramethylbenzidine and o-phenylenediamine. The composition of the invention can be used for detecting vibrio parahaemolyticus, has higher sensitivity, the detection limit is 1.12cfu/mL, and the linear range is 0-1 multiplied by 10 4 cfu/mL，R 2 = 0.9934. As the concentration of Vibrio parahaemolyticus increases, the reaction solution changes from blue-green to green and then to yellow, so that semi-quantitative detection is carried out by visual observation, and the visual detection limit is 10cfu/mL. In addition, the developed dual substrate colorimetric method has good selectivity and recovery rate for the Vibrio parahaemolyticus for the determination of Vibrio parahaemolyticus.
Drawings
FIG. 1 is a synthetic Fe 3 O 4 -a characterization map of Zn-Mn nanoenzyme;
FIG. 2 is a sensitivity graph and a linear regression graph of Vibrio parahaemolyticus corresponding to different concentrations with different colors;
FIG. 3 shows that under the optimal condition, the interference bacteria and the Vibrio parahaemolyticus are respectively added into a detection system, and are detected by an ultraviolet-visible spectrophotometer.
Detailed Description
The invention provides a kit for detecting vibrio parahaemolyticus, which comprises the Fe as claimed in claim 1 3 O 4 -Zn-Mn-Apt probe, 3' 5-tetramethylbenzidine and o-phenylenediamine.
In the present invention, the Fe 3 O 4 The preparation method of the Zn-Mn-Apt probe comprises the following steps:
(1) Mixing ferric trichloride, glycol, sodium acetate, sodium citrate and polyethylene glycol 6000, reacting, and drying to obtain Fe 3 O 4 Magnetic nanoparticles;
(2) Mixing manganese chloride, water and bovine serum albumin solution, stirring, adding sodium hydroxide solution for reaction, adding zinc acetate solution for secondary reaction, centrifuging, and drying the precipitate to obtain Zn-Mn nano enzyme;
(3) Zn-Mn nano enzyme and Fe 3 O 4 Mixing magnetic nano particles with water, stirring for reaction, collecting reaction products, and drying to obtain Fe 3 O 4 -Zn-Mn nanoenzyme;
(4) Fe with 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide and N-hydroxysuccinimide 3 O 4 -Zn-Mn nano enzyme activation site, fe after activating site 3 O 4 Mixing Zn-Mn nano enzyme with vibrio parahaemolyticus aptamer to obtain Fe 3 O 4 -Zn-Mn-Apt probe.
In the present invention, the dosage ratio of the ferric trichloride, the ethylene glycol, the sodium acetate, the sodium citrate and the polyethylene glycol 6000 in the step (1) is preferably: 18-22 mM: 18-22 mL: 70-80 mM: 30-40 mM:0.18 to 0.22g, more preferably 20mM:20mL:75mM:35mM: the temperature of the reaction in step (1) is preferably 190 to 210 ℃, more preferably 200 ℃, the time of the reaction is preferably 17 to 19 hours, more preferably 18 hours, and the temperature of the drying is preferably 55 to 65 ℃, more preferably 60 ℃.
In the present invention, the ratio of the manganese chloride, water and bovine serum albumin solution in the step (2) is preferably 95 to 105mM: 1.8-2.2 mL:9 to 11mL, more preferably 100mM:2.0mL:10mL, the concentration of the bovine serum albumin solution is preferably 4 to 6mg/mL, more preferably 5mg/mL, the stirring time in the step (2) is preferably 25 to 35min, more preferably 30min, the amount of the sodium hydroxide solution is the same as that of the bovine serum albumin solution, the concentration of the sodium hydroxide solution is preferably 60 to 80mM, more preferably 70mM, the reaction time in the step (2) is preferably 4 to 6min, more preferably 5min, the amount of the zinc acetate solution is preferably 1.8 to 2.2 times, more preferably 2 times that of the bovine serum albumin solution, the time of the secondary reaction is preferably 10 to 14h, more preferably 12h, the centrifugation condition is preferably 11000 to 13000rpm, 13 to 17min, more preferably 12000rpm, 15min, and the drying temperature in the step (2) is preferably 45 to 55 ℃, more preferably 50 ℃.
In the present invention, the Zn-Mn nanoenzyme, fe in the step (3) 3 O 4 The dosage ratio of the magnetic nano particles to the water is preferably 18-22 mg:0.18 to 0.22g:45 to 55mL, more preferably 20mg:0.20g:50mL, the temperature of the stirring reaction is preferably 55-65 ℃, more preferably 60 ℃, and the time is preferably 1.8-2.2 h, more preferably 2h, the reaction product is preferably collected by magnetic adsorption, and the drying temperature in the step (3) is preferably 55-65 ℃, more preferably 60 ℃.
In the present invention, the concentration of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide in step (4) is preferably 8 to 12mg/mL, more preferably 10mg/mL, the concentration of N-hydroxysuccinimide is preferably 8 to 12mg/mL, more preferably 10mg/mL, the concentration of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide, N-hydroxysuccinimide and Fe 3 O 4 The dosage ratio of the Zn-Mn nano enzyme is preferably 1.8-2.2 mL: 1.8-2.2 mL:1mg, more preferably 2mL:2mL:1mg, wherein the activation time is 25-35 min.
In the present invention, fe 3 O 4 The enzymatic activity of the-Zn-Mn-Apt probe induces oxidation of TMB to TMB oxide (TMB) + Bluish green) and o-phenylenediamine (OPD) to form OPD oxide (yellow), multiple colors are generated due to the complementary nature of bluish green and yellow. With TMB + The absorbance at 652nm was measured for Vibrio parahaemolyticus.
The invention also provides a using method of the kit, which comprises the following steps: sample to be measured, fe 3 O 4 Mixing a Zn-Mn-Apt probe, 3', 5-tetramethylbenzidine and o-phenylenediamine to obtain a mixed solution, and detecting the absorbance of the mixed solution at 652 nm;
the linear regression equation is adopted as Y= -0.2215X+0.9405, R 2 = 0.9934 to calculate the concentration of vibrio parahaemolyticus in the sample to be tested, where Y is absorbance, X is LogC, and C is the concentration of vibrio parahaemolyticus in cfu/mL.
In the present invention, based on a two-substrate colorimetric method, in Fe 3 O 4 The presence of Zn-Mn gives rise to various colours. Catalysis produces polychromatic light. By utilizing the phenomenon, a convenient multicolor comparison method is established, and the vibrio parahaemolyticus is quantitatively and semi-quantitatively detected. Fe when Vibrio parahaemolyticus is present 3 O 4 Certain sites on the surface of the Zn-Mn aptamer are occupied, resulting in TMB with increasing Vibrio parahaemolyticus concentration + Gradually decreasing in the generation of (a). Thus, the solution shows a multi-colored light signal. These colors correspond to different concentrations of Vibrio parahaemolyticus and can be visually distinguished. With the increase of the concentration of the vibrio parahaemolyticus, the reaction solution changes from blue-green to green and then to yellow, so that the visual observation semi-quantitative detection is carried out, and the visual detection limit is 10cfumL -1 。
The technical solutions provided by the present invention are described in detail below with reference to examples, but they should not be construed as limiting the scope of the present invention.
EXAMPLE 1Fe 3 O 4 -Zn-Mn-Apt probe synthesis
(1) First, fe is synthesized 3 O 4 Magnetic nanoparticles:
the mixture was obtained by ultrasonic treatment of 20mM ferric trichloride in 20mL of ethylene glycol, 75mM sodium acetate, 35mM sodium citrate and 0.2g of polyethylene glycol 6000 until the mixture was dissolved uniformly. The mixture was placed in a stainless steel autoclave and reacted at 200℃for 18 hours. Naturally cooling to room temperature, and washing with distilled water and absolute ethyl alcohol respectively. Then placing the mixture into a 60 ℃ oven, drying, grinding, collecting, sealing and preserving.
(2) Zn-Mn nano enzyme synthesis:
100mM manganese chloride was dissolved in 2mL of water, 10mL of a bovine serum albumin solution (5 mg/mL) was added, and the mixture was vigorously stirred for 30min. 10mL of sodium hydroxide (70 mM) was then added and the reaction was continued for 5min. 20mL of zinc acetate (10 mM) was added dropwise, and the reaction was continued for 12 hours. Centrifuge at 12000rpm for 15min. The precipitate was collected and washed 3 times with absolute ethanol. Drying at 50deg.C overnight, grinding, collecting, and sealing.
③Fe 3 O 4 Zn-Mn nano enzyme synthesis
Taking 20mg of Zn-Mn nano enzyme and Fe 3 O 4 0.2g of magnetic nanoparticles was dissolved in 50mL of water. The reaction was stirred vigorously at 60℃for 2h. Magnetic adsorption and collection, and drying at 60 ℃. Grinding, collecting, sealing and storing.
④Fe 3 O 4 Synthesis of-Zn-Mn-Apt nanoprobe
Fe was reacted with 1- (3-dimethylaminopropyl) -3-Ethylcarbodiimide (EDC) (10 mg/mL) and N-hydroxysuccinimide (NHS) (10 mg/mL) 3 O 4 The Zn-Mn magnetic nano enzyme activation site is subjected to 30min, so that the surface group of the Zn-Mn magnetic nano enzyme activation site is easy to couple with an aptamer, supernatant is removed by magnetic adsorption, and 100 mu L of 100 mu M of vibrio parahaemolyticus aptamer (Apt) (the nucleotide sequence is NH) is added 2 TCTAAAAAT GGG CAAAGAAACAGT GAC TCG TTGAGATAC T) suspension overnight coupling, followed by 3 washes with sterile PBS to give Fe 3 O 4 -Zn-Mn-Apt probe.
For the prepared Fe 3 O 4 Characterization of Zn-Mn nanoenzyme is carried out, the result is shown in figure 1, A in figure 1 is Fe 3 O 4 TEM characterization of magnetic nanoparticles shows that the prepared magnetic nanoparticles are round and have uniform particle size. B in FIG. 1 is the productThe Zn-Mn nanoenzyme was prepared, and TEM results showed a cloud form, which was based on bovine serum albumin. C in FIG. 1 is Fe 3 O 4 TEM results of Zn-Mn nanoenzyme, as shown in the figure, the resulting nanocomposite has a rough surface and a size greater than Fe 3 O 4 Magnetic nanoparticles. D in FIG. 1 is the application of Fe 3 O 4 Capturing vibrio parahaemolyticus by using-Zn-Mn-Apt nano probe, and displaying Fe 3 O 4 The Zn-Mn-Apt nanoprobes aggregate on the bacterial surface. Has excellent identification function to target bacteria.
EXAMPLE 2 construction of Vibrio parahaemolyticus detection System
First, 0.5mg of Fe was added to a sample bottle 3 O 4 Zn-Mn-Apt probe, 1mL Vibrio parahaemolyticus solutions (10-10) of different concentrations 4 cfu/mL), mixer incubation for 30min, magnetic separation and supernatant removal. Next, 500. Mu.L of OPD solution (0.15 mM) and 500. Mu.L of LTMB solution (8 mM) were left at room temperature for 5min. Magnetic separation, taking the supernatant into a 1.5mL centrifuge tube, and recording color change and ultraviolet-visible spectrum in the range of 500-800 nm. A standard curve was constructed using the concentration and absorbance values (652 nm) of Vibrio parahaemolyticus. The results are shown in the interpolated diagram of fig. 2. With the sequential increase of the concentration of the target bacteria, the color of the experimental tube is gradually changed from blue-green to green, then changed into yellow, and the corresponding ultraviolet absorption peak at 652nm is gradually reduced. Constructing a linear regression curve by taking an ultraviolet absorption peak of A652nm as an ordinate and a Log value of the concentration of the target bacteria as an abscissa, wherein R is 2 = 0.9934, the results are shown to have good consistency.
Example 3 interference experiments
Because the food matrix is complex and various, other bacteria are often present for interference detection, and for testing the anti-interference performance of a detection system, common 4 food-borne pathogenic bacteria are selected as representatives, including Staphylococcus Aureus (SA), listeria Monocytogenes (LM), salmonella Typhimurium (ST) and escherichia coli O157: h7 (the concentrations are 10 each 3 cfu/mL) as potential co-existing bacteria. PBS samples served as reagent blanks. Under the optimal condition, the interference bacteria and the vibrio parahaemolyticus are respectively added into a detection system, and are detected by an ultraviolet-visible spectrophotometer. Blank group, 4The common bacteria and the mixed bacteria are blue-green, which indicates Fe 3 O 4 The surface sites of the Zn-Mn-aptamer were not captured. In the case of Vibrio parahaemolyticus alone or in combination with other bacteria, the colour of the tube is light and there is a significant change in absorbance in the sample containing interfering bacteria. In contrast, when vibrio parahaemolyticus is present, the absorbance change in the sample tube is small, indicating that the presence of these interfering bacteria has little effect on the detection result.
Example 4 food simulation test
In order to further study the potential application prospect of the developed multicolor method in vibrio parahaemolyticus food analysis, common food samples (mackerel and seaweed) polluted by vibrio parahaemolyticus are detected by adopting a colorimetric method so as to determine the practicability of the method. Food samples were processed according to literature and tested according to the same procedure. In addition, 7 and 70cfu/mL levels of Vibrio parahaemolyticus were repeatedly detected in three samples. The results summarized are shown in table 1, with recovery ranging from 89.14 to 115.86% and RSD ranging from 3.4 to 13.89% (n=3). These results show that the method has potential application prospect in detecting vibrio parahaemolyticus in real food samples. In conclusion, the multicolor method has wide application prospect for analyzing the vibrio parahaemolyticus. The method demonstrates its utility and potential for use in the real world by successfully detecting and quantifying Vibrio parahaemolyticus in a normal food sample.
TABLE 1 detection of recovery and RSD values of bacteria in the added samples [ (]n＝3).
From the above embodiments, the present invention provides a detectionComposition of vibrio parahaemolyticus and application thereof, wherein the composition comprises Fe 3 O 4 -Zn-Mn-Apt probe, 3' 5-tetramethylbenzidine and o-phenylenediamine. The composition of the invention can be used for detecting vibrio parahaemolyticus, has higher sensitivity, the detection limit is 1.12cfu/mL, and the linear range is 0-1 multiplied by 10 4 cfu/mL，R 2 = 0.9934. As the concentration of Vibrio parahaemolyticus increases, the reaction solution changes from blue-green to green and then to yellow, so that semi-quantitative detection is carried out by visual observation, and the visual detection limit is 10cfu/mL. In addition, the developed dual substrate colorimetric method has good selectivity and recovery rate for the Vibrio parahaemolyticus for the determination of Vibrio parahaemolyticus.
The foregoing is merely a preferred embodiment of the present invention and it should be noted that modifications and adaptations to those skilled in the art may be made without departing from the principles of the present invention, which are intended to be comprehended within the scope of the present invention.
Claims (8)
1. A composition for detecting Vibrio parahaemolyticus, characterized in that the composition comprises Fe 3 O 4 -Zn-Mn-Apt probe, 3' 5-tetramethylbenzidine and o-phenylenediamine;
the Fe is 3 O 4 The preparation method of the Zn-Mn-Apt probe comprises the following steps:
(1) Mixing ferric trichloride, glycol, sodium acetate, sodium citrate and polyethylene glycol 6000, reacting, and drying to obtain Fe 3 O 4 Magnetic nanoparticles;
(2) Mixing manganese chloride, water and bovine serum albumin solution, stirring, adding sodium hydroxide solution for reaction, adding zinc acetate solution for secondary reaction, centrifuging, and drying the precipitate to obtain Zn-Mn nano enzyme;
(3) Zn-Mn nano enzyme and Fe 3 O 4 Mixing magnetic nano particles with water, stirring for reaction, collecting reaction products, and drying to obtain Fe 3 O 4 -Zn-Mn nanoenzyme;
(4) With 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide andn-hydroxysuccinimide reacting Fe 3 O 4 -Zn-Mn nano enzyme activation site, fe after activating site 3 O 4 Mixing the Zn-Mn nano enzyme with the Vibrio parahaemolyticus aptamer Apt to obtain Fe 3 O 4 -Zn-Mn-Apt probe.
2. The composition of claim 1, wherein the ratio of the amounts of ferric trichloride, ethylene glycol, sodium acetate, sodium citrate, and polyethylene glycol 6000 in step (1) is: 18-22 mM: 18-22 mL: 70-80 mM: 30-40 mM: 0.18-0.22 g, wherein the temperature of the reaction in the step (1) is 190-210 ℃, the reaction time is 17-19 h, and the drying temperature is 55-65 ℃.
3. The composition of claim 2, wherein the manganese chloride, water and bovine serum albumin solution in step (2) are used in an amount ratio of 95 to 105mM: 1.8-2.2 mL: 9-11 mL, wherein the concentration of the bovine serum albumin solution is 4-6 mg/mL, the stirring time in the step (2) is 25-35 min, the amount of the sodium hydroxide solution is the same as that of the bovine serum albumin solution, the concentration of the sodium hydroxide solution is 60-80 mM, the reaction time in the step (2) is 4-6 min, the amount of the zinc acetate solution is 1.8-2.2 times that of the bovine serum albumin solution, the secondary reaction time is 10-14 h, the centrifugation condition is 11000-13000 rpm, the centrifugation time is 13-17 min, and the drying temperature in the step (2) is 45-55 ℃.
4. A composition according to claim 3, wherein in step (3) the Zn-Mn nanoenzyme, fe 3 O 4 The dosage ratio of the magnetic nano particles to the water is 18-22 mg:0.18 to 0.22g: 45-55 mL, wherein the temperature of the stirring reaction is 55-65 ℃ and the time is 1.8-2.2 h, the reaction products are collected by magnetic adsorption, and the drying temperature in the step (3) is 55-65 ℃.
5. The composition of claim 4, wherein in step (4) the 1- (3-dimethyl) isThe concentration of the aminopropyl) -3-ethyl carbodiimide is 8-12 mg/mL, the concentration of the N-hydroxysuccinimide is 8-12 mg/mL, and the 1- (3-dimethylaminopropyl) -3-ethyl carbodiimide, the N-hydroxysuccinimide and Fe 3 O 4 The dosage ratio of the Zn-Mn nano enzyme is 1.8-2.2 mL: 1.8-2.2 mL:1mg, wherein the activation time is 25-35 min.
6. Use of the composition according to any one of claims 1 to 5 for the preparation of a vibrio parahaemolyticus detection kit.
7. A kit for detecting Vibrio parahaemolyticus, comprising the Fe of claim 1 3 O 4 -Zn-Mn-Apt probe, 3' 5-tetramethylbenzidine and o-phenylenediamine.
8. The method of using the kit of claim 7, comprising the steps of: mixing the sample to be tested with Fe 3 O 4 After the Zn-Mn-Apt probe is mixed and incubated, adding 3' 5-tetramethyl benzidine and o-phenylenediamine for mixing to obtain a mixed solution, and detecting the absorbance of the mixed solution at 652 nm;
the linear regression equation is adopted as Y= -0.2215X+0.9405, R 2 = 0.9934 to calculate the concentration of vibrio parahaemolyticus in the sample to be tested, where Y is absorbance, X is LogC, and C is the concentration of vibrio parahaemolyticus in cfu/mL.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311195101.4A CN117110607B (en) | 2023-09-16 | 2023-09-16 | Composition for detecting vibrio parahaemolyticus and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311195101.4A CN117110607B (en) | 2023-09-16 | 2023-09-16 | Composition for detecting vibrio parahaemolyticus and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN117110607A CN117110607A (en) | 2023-11-24 |
| CN117110607B true CN117110607B (en) | 2024-01-26 |
Family
ID=88805559
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202311195101.4A Active CN117110607B (en) | 2023-09-16 | 2023-09-16 | Composition for detecting vibrio parahaemolyticus and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117110607B (en) |
Citations (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB2078372A (en) * | 1979-12-26 | 1982-01-06 | Matsushita Electric Ind Co Ltd | Atmosphere sensing element |
| CN102253193A (en) * | 2010-05-20 | 2011-11-23 | 上海医脉赛科技有限公司 | Magnetic fluorescent kit for rapidly detecting microbes as well as preparation method and use method thereof |
| CN103065751A (en) * | 2011-10-19 | 2013-04-24 | 中国科学院宁波材料技术与工程研究所 | Magnetic MFe2O4/C/M' compound and preparation method thereof using aniline small organic molecules as a carbon source |
| CN105102979A (en) * | 2013-01-29 | 2015-11-25 | 生物辐射海法有限公司 | Hand maneuverable laser welding gun |
| WO2016112195A1 (en) * | 2015-01-09 | 2016-07-14 | Etubics Corporation | Methods and compositions for combination immunotherapy |
| CN107045061A (en) * | 2017-06-05 | 2017-08-15 | 吉林大学 | Based on Magnetic Isolation and quantum dot-labeled vibrio parahemolyticus detection kit |
| CN107202884A (en) * | 2017-06-16 | 2017-09-26 | 吉林大学 | Based on immunomagnetic beads and MnO2Nano-particle detects vibrio parahemolyticus kit |
| WO2018152422A1 (en) * | 2017-02-16 | 2018-08-23 | Essenlix Corporation | Assay with textured surface |
| CN113281507A (en) * | 2021-05-23 | 2021-08-20 | 吉林大学 | Rapid detection method and kit for staphylococcus aureus |
| CN114371287A (en) * | 2021-12-02 | 2022-04-19 | 鲁东大学 | Staphylococcus aureus detection kit based on immunomagnetic separation and click chemical reaction and detection method thereof |
| CN114891755A (en) * | 2022-05-26 | 2022-08-12 | 华中农业大学 | Vibrio parahaemolyticus phage VPP1 of Changtail family and application thereof |
| CN115672374A (en) * | 2022-10-31 | 2023-02-03 | 中国科学技术大学 | Metal monoatomic/metal nanoparticle nanocomposite with cascaded catalytic activity and preparation and application thereof |
| CN116203235A (en) * | 2022-09-13 | 2023-06-02 | 鲁东大学 | Rapid detection kit for staphylococcus aureus and detection method thereof |
| CN116713002A (en) * | 2023-04-25 | 2023-09-08 | 西南交通大学 | Nano enzyme with peroxidase-like activity, and preparation method and application thereof |
-
2023
- 2023-09-16 CN CN202311195101.4A patent/CN117110607B/en active Active
Patent Citations (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB2078372A (en) * | 1979-12-26 | 1982-01-06 | Matsushita Electric Ind Co Ltd | Atmosphere sensing element |
| CN102253193A (en) * | 2010-05-20 | 2011-11-23 | 上海医脉赛科技有限公司 | Magnetic fluorescent kit for rapidly detecting microbes as well as preparation method and use method thereof |
| CN103065751A (en) * | 2011-10-19 | 2013-04-24 | 中国科学院宁波材料技术与工程研究所 | Magnetic MFe2O4/C/M' compound and preparation method thereof using aniline small organic molecules as a carbon source |
| CN105102979A (en) * | 2013-01-29 | 2015-11-25 | 生物辐射海法有限公司 | Hand maneuverable laser welding gun |
| WO2016112195A1 (en) * | 2015-01-09 | 2016-07-14 | Etubics Corporation | Methods and compositions for combination immunotherapy |
| WO2018152422A1 (en) * | 2017-02-16 | 2018-08-23 | Essenlix Corporation | Assay with textured surface |
| CN107045061A (en) * | 2017-06-05 | 2017-08-15 | 吉林大学 | Based on Magnetic Isolation and quantum dot-labeled vibrio parahemolyticus detection kit |
| CN107202884A (en) * | 2017-06-16 | 2017-09-26 | 吉林大学 | Based on immunomagnetic beads and MnO2Nano-particle detects vibrio parahemolyticus kit |
| CN113281507A (en) * | 2021-05-23 | 2021-08-20 | 吉林大学 | Rapid detection method and kit for staphylococcus aureus |
| CN114371287A (en) * | 2021-12-02 | 2022-04-19 | 鲁东大学 | Staphylococcus aureus detection kit based on immunomagnetic separation and click chemical reaction and detection method thereof |
| CN114891755A (en) * | 2022-05-26 | 2022-08-12 | 华中农业大学 | Vibrio parahaemolyticus phage VPP1 of Changtail family and application thereof |
| CN116203235A (en) * | 2022-09-13 | 2023-06-02 | 鲁东大学 | Rapid detection kit for staphylococcus aureus and detection method thereof |
| CN115672374A (en) * | 2022-10-31 | 2023-02-03 | 中国科学技术大学 | Metal monoatomic/metal nanoparticle nanocomposite with cascaded catalytic activity and preparation and application thereof |
| CN116713002A (en) * | 2023-04-25 | 2023-09-08 | 西南交通大学 | Nano enzyme with peroxidase-like activity, and preparation method and application thereof |
Non-Patent Citations (4)
| Title |
|---|
| Effect of physiochemical property of Fe3O4 particle on magnetic lateral flow immunochromatographic assay;Yan Jun 等;SENSORS AND ACTUATORS B-CHEMICAL;第197卷;129-136 * |
| Mesoporous MnFe2O4 magnetic nanoparticles as a peroxidase mimic for the colorimetric detection of urine glucose;Ke Liu 等;RSC Adv.;第11卷;28375-28380 * |
| 双抗体夹心ELISA法检测食品中出血性大肠杆菌O15;黄海燕;陈亮;陈萍;;食品工业科技(第11期);185-194 * |
| 基于纳米材料的光学适配体传感器在农药残留检测中的应用研究进展;付瑞杰 等;食品科学;第42卷(第03期);330-338 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN117110607A (en) | 2023-11-24 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN112904007B (en) | Method for detecting vomitoxin based on blue magnetic porous double-nanoenzyme/quantum dot double-signal amplification immunochromatographic test strip | |
| CN110118759B (en) | Oxytetracycline fluorescence detection method based on surface passivation and covalent coupling | |
| Dudak et al. | Development of an immunosensor based on surface plasmon resonance for enumeration of Escherichia coli in water samples | |
| CN111024938B (en) | Rapid detection method for pathogenic microorganisms | |
| Meng et al. | Novel chitosan-based fluorescent materials for the selective detection and adsorption of Fe3+ in water and consequent bio-imaging applications | |
| CN108358814B (en) | AIE fluorescent probe based on salicylhydrazone derivative and preparation method and application thereof | |
| Mo et al. | A simple paper-based ratiometric luminescent sensor for tetracyclines using copper nanocluster-europium hybrid nanoprobes | |
| CN105527427B (en) | A kind of method of quick detection Listeria monocytogenes | |
| CN117110607B (en) | Composition for detecting vibrio parahaemolyticus and application thereof | |
| Chen et al. | N-doped MoS2-nanoflowers as peroxidase-like nanozymes for total antioxidant capacity assay | |
| Celik et al. | Preparation of natural indicator incorporated media and its logical use as a colorimetric biosensor for rapid and sensitive detection of Methicillin-resistant Staphylococcus aureus | |
| Liu et al. | A colorimetric sensor for Staphylococcus aureus detection based on controlled click chemical-induced aggregation of gold nanoparticles and immunomagnetic separation | |
| Bai et al. | Portable dual-mode biosensor based on smartphone and glucometer for on-site sensitive detection of Listeria monocytogenes | |
| CN113281507B (en) | Rapid detection method and kit for staphylococcus aureus | |
| Yildirim et al. | Synthesis of new fluorescent metal-organic framework (4-sulfo-1, 8-naphthalimide-UiO66) for detection of Cr (III)/Cr (VI) ions in aqueous media and its antimicrobial studies | |
| CN116735580B (en) | Meat freshness detection sensor based on bimodal monoatomic nano enzyme and preparation method thereof | |
| CN1828303A (en) | Method for quick detection of microbe | |
| CN116203235A (en) | Rapid detection kit for staphylococcus aureus and detection method thereof | |
| CN110501494A (en) | A kind of microorganism detection method based on manganese dioxide nano flower and fluorescent material | |
| CN111366727B (en) | Kit for detecting salmonella typhimurium and preparation method thereof | |
| CN115404074A (en) | Fluorescence detection nano probe, preparation method and application | |
| CN115246823A (en) | Difunctional near-infrared emission fluorescent probe for detecting bisulfite and indicating freshness of fish meat and synthetic method and application thereof | |
| CN114371287A (en) | Staphylococcus aureus detection kit based on immunomagnetic separation and click chemical reaction and detection method thereof | |
| Miyamoto et al. | Rapid detection and counting of viable bacteria in vegetables and environmental water using a photon-counting TV camera | |
| CN111323596A (en) | Staphylococcus aureus detection kit and preparation method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |