CN113372439A - Manganese metal organic framework biological composite material and application thereof in detection of food-borne pathogenic bacteria - Google Patents
Manganese metal organic framework biological composite material and application thereof in detection of food-borne pathogenic bacteria Download PDFInfo
- Publication number
- CN113372439A CN113372439A CN202110642668.6A CN202110642668A CN113372439A CN 113372439 A CN113372439 A CN 113372439A CN 202110642668 A CN202110642668 A CN 202110642668A CN 113372439 A CN113372439 A CN 113372439A
- Authority
- CN
- China
- Prior art keywords
- organic framework
- mof
- composite material
- manganese metal
- metal organic
- 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.)
- Pending
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
- C08G83/00—Macromolecular compounds not provided for in groups C08G2/00 - C08G81/00
- C08G83/008—Supramolecular polymers
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
- C12N15/115—Aptamers, i.e. nucleic acids binding a target molecule specifically and with high affinity without hybridising therewith ; Nucleic acids binding to non-nucleic acids, e.g. aptamers
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/02—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating impedance
- G01N27/04—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating impedance by investigating resistance
- G01N27/06—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating impedance by investigating resistance of a liquid
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
-
- 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
- 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
- G01N33/56916—Enterobacteria, e.g. shigella, salmonella, klebsiella, serratia
-
- 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
- G01N33/56938—Staphylococcus
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2310/00—Structure or type of the nucleic acid
- C12N2310/10—Type of nucleic acid
- C12N2310/16—Aptamers
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Hematology (AREA)
- Physics & Mathematics (AREA)
- Urology & Nephrology (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Genetics & Genomics (AREA)
- Organic Chemistry (AREA)
- Medicinal Chemistry (AREA)
- Biotechnology (AREA)
- Analytical Chemistry (AREA)
- Microbiology (AREA)
- Virology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Food Science & Technology (AREA)
- Cell Biology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Electrochemistry (AREA)
- Biophysics (AREA)
- General Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Polymers & Plastics (AREA)
- Plant Pathology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Investigating Or Analyzing Materials By The Use Of Electric Means (AREA)
Abstract
The invention discloses a manganese metal organic framework biological composite material and application thereof in detection of food-borne pathogenic bacteria. The manganese metal organic framework biological composite material is formed by covalently coupling Mn-MOF-74 nano microspheres and a specific biological recognition material, and the particle size of the Mn-MOF-74 nano microspheres is 200-300 nm. The manganese metal organic framework biological composite material has the advantages of sensitive electrochemical impedance signal response, stable structure and specific identification of target pathogenic bacteria, namely the listeria monocytogenes impedance type electrochemical immunosensor, wherein the Mn-MOF-74 crystals have uniform size, good biological affinity and easy modification, the prepared manganese metal organic framework biological composite material has specific marking capability on target bacteria and is easy to be reduced by hydrogen peroxide to generate manganese ions with good conductivity, and the listeria monocytogenes electrochemical detection technology developed by using the biological composite material has the advantages of simple operation, low detection limit and high efficiency and rapidness in the detection process.
Description
Technical Field
The invention relates to a manganese metal organic framework biological composite material and application thereof in detection of food-borne pathogenic bacteria, belonging to the technical field of detection of food-borne pathogenic bacteria.
Background
Metal Organic Frameworks (MOFs), as an advanced porous material, have a variety of functional characteristics such as designability of structure, ordered spatial conformation, adjustable framework size, high porosity, large surface area, etc., and are currently used in selective adsorption/separation, novel catalysis, microelectronic materials, chemical/biological sensing, and in vivo drug delivery. Because the particle size of the MOF crystal is larger, the stability of acid/alkali solution is sensitive and the non-specific adsorption is limited, the MOF is less researched as an electrochemical detection probe. Existing MOF electrochemical sensor studies are mainly based on classical MOF structures, i.e. forming composite materials with other materials and then generating electrical signals by chemical reactions. The main detection mode is also based on the huge specific surface area of the MOF, and the MOF is used as an adsorption material. The application of MOF materials as electrochemical probes currently has two main difficulties: (1) most of MOFs have low conductivity, extra electric signal interference ions can be introduced by modification strategies of electrochemical signals of a material doped conductive substance (graphene, tin oxide, biomacromolecule enzyme and the like) lifter, and the acquisition of signals is interfered in the actual sample detection process, so that the detection sensitivity and accuracy are reduced. In addition, the MOF is used as a good adsorption material, but the MOF is lack of selective adsorption capacity, and a target object and impurities are easily adsorbed and captured together in the detection process, so that detection errors are caused.
Diseases caused by food-borne pathogens continue to be a global problem threatening public health. It is counted that more than 200 million people die each year from diarrheal diseases, many of which are infected by intake of water and bacteria-bearing food contaminated with pathogenic microorganisms (viruses, bacteria, harmful fungi, parasites). These threats are mostly due to contamination in water and food. Listeria monocytogenes (L.m) is a typical food-borne pathogen, a gram-positive Brevibacterium, facultative anaerobic, first isolated in 1924 in the blood of septic rabbits. Although the incidence of listeriosis is low, its fatality rate is still as high as 30%, making it the third most fatal disease of all food-borne diseases. The susceptible patient group of the disease is mainly the elderly, pregnant women, newborns and adults with low immunity. Once infected, it causes mononucleosis, fever, muscle pain, diarrhea and other gastrointestinal symptoms in the blood of the body, typical symptoms being septicemia and meningitis. Because the pathogen can contaminate drinking water and food and has extremely low residual quantity, a corresponding detection method is established, the improvement of detection precision and efficiency is an important means for ensuring the avoidance of outbreak of food-borne diseases, and the development of a novel functional biosensor becomes the trend of food safety detection. Therefore, it is expected to employ a metal organic framework biocomposite as a biosensor to time-limit high-sensitivity detection of food harmful substances.
Disclosure of Invention
The invention aims to provide a manganese metal organic framework biological composite material, namely a composite material of a manganese metal organic framework (Mn-MOF-74 nano microsphere) and a specific biological recognition material (anti-Listeria monoclonal antibody), which can be used as an impedance immunosensor, has the characteristics of easy generation of electrochemical signals and stable signal response, has good specific capture on target pathogenic bacteria such as Listeria monocytogenes, and can be used for detecting food-borne pathogenic bacteria.
The manganese metal organic framework biological composite material provided by the invention is formed by covalently coupling Mn-MOF-74 nano microspheres and a specific biological recognition material.
The particle size of the Mn-MOF-74 nano microspheres is 200-300 nm.
The specific biological recognition material can be a monoclonal antibody, a polyclonal antibody, a genetic engineering antibody or an aptamer, and specifically depends on the biological activity and stability of the recognition material and the specific recognition capability of the target to be detected, and the specific embodiment of the invention adopts a mouse monoclonal antibody for resisting listeria monocytogenes.
The invention further provides a preparation method of the manganese metal organic framework biological composite material, which comprises the following steps:
coupling the Mn-MOF-74 nano microspheres and the specific biological recognition material by adopting an active ester method;
the active ester method comprises the following steps:
1) dispersing the Mn-MOF-74 nano microspheres in a PBS buffer solution, adding EDC for reaction to obtain the Mn-MOF-74 nano microspheres activated by carboxyl;
2) dispersing the specific biological recognition material in PBS (phosphate buffer solution) of NHS (polyethylene glycol succinate), mixing with the system obtained in the step 1), reacting, and centrifuging to obtain the manganese metal organic framework biological composite material.
In the steps 1) and 2), the pH value of the PBS buffer solution is 7.1-7.4.
In the step 1), after the Mn-MOF-74 nano microspheres are added into a PBS (phosphate buffer solution), carrying out ultrasonic treatment (for example, for 30min) so as to uniformly disperse the Mn-MOF-74 nano microspheres in the PBS buffer solution;
in the PBS buffer solution, the concentration of the Mn-MOF-74 nano microspheres can be 5.0-10 mg/mL.
In step 1), the reaction is carried out under vortex conditions;
and after the reaction is finished, centrifuging, washing the EDC which does not participate in the reaction by using the PBS buffer solution, and re-suspending by using the PBS buffer solution.
In the step 2), the obtained PBS buffer solution resuspension can be mixed with the PBS buffer solution dispersed with NHS;
the molar ratio of the NHS to the EDC may be 1: 2-6, preferably 1: 2;
vortex reaction can be carried out for 2h at 37 ℃;
the resulting composite material can be resuspended in bovine serum albumin blocking solution (0.1% BSA-PBS), and then stored at 4 ℃ under a low temperature and sealed condition.
In the preparation method, the Mn-MOF-74 nano microspheres are prepared from manganese chloride tetrahydrate and 2, 5-dihydroxy terephthalic acid through a solvothermal reaction method;
the molar ratio of the manganese chloride tetrahydrate to the 2, 5-dihydroxy terephthalic acid is 2.0-4.0: 1, preferably 3.3: 1.
the solvent thermal reaction method comprises the following steps:
dissolving the manganese chloride tetrahydrate and the 2, 5-dihydroxy terephthalic acid in a mixed solvent of DMF, methanol and water to obtain a mixed solution; and transferring the mixed solution into a sleeve of a reaction kettle, heating at 130-140 ℃ for 20-24 h for crystallization, and centrifuging to obtain the Mn-MOF-74 nano microspheres.
In the mixed solvent, the volume ratio of the DMF, the methanol and the water is preferably 1:1: 15.
the specific biological identification material modified on the surface of the manganese metal organic framework biological composite material can perform specific identification on various pathogenic bacteria such as escherichia coli, staphylococcus aureus, salmonella, listeria monocytogenes and the like.
The manganese metal organic framework biological composite material can be reacted by various reducing substances (hydrogen peroxide, vitamin C and glutathione) and release manganese (II) ions with good conductive signals when being used as an impedance type electrochemical signal probe.
The manganese metal organic framework biological composite material can be used for detecting food-borne pathogenic bacteria such as salmonella, staphylococcus aureus, escherichia coli and listeria monocytogenes by changing biological identification materials aiming at different targets;
the method can be used for detecting food-borne pathogenic bacteria in the following samples:
1) ambient water;
2) domestic water and drinking water;
3) dairy products, including milk, yogurt, fruit milk and reconstituted milk;
4) liquid beverages, including bottled water, wines, juices, and carbonated beverages.
The manganese metal organic framework biological composite material is an impedance type electrochemical immunosensor which has the advantages of sensitive electrochemical impedance signal response, stable structure and specificity identification of target pathogenic bacteria, namely Listeria monocytogenes, wherein the size of Mn-MOF-74 crystals is uniform (about 150-200 nm), the biological affinity is good, the modification is easy, the prepared manganese metal organic framework biological composite material has the specificity marking capability on target bacteria, and is easy to be reduced by the target bacteria to generate manganese hydrogen peroxide ions with good conductivity, and the electrochemical detection technology for the Listeria monocytogenes developed by using the biological composite material has the advantages of simple operation, low detection limit and high efficiency and rapidness in the detection process.
Drawings
FIG. 1 is a schematic diagram of the preparation of the metal organic framework biological composite material of the present invention.
FIG. 2 is a representation of the morphology and structural composition of the metal organic framework biocomposite prepared in example 1 of the present invention, wherein FIG. 2(a) is a TEM image of Mn-MOF-74 microspheres; FIG. 2(b) is an XPS characterization of Mn-MOF-74 microspheres; FIG. 2(c) is an XRD representation of Mn-MOF-74 microspheres; FIG. 2(d) is FTIR characterization of Mn-MOF-74 microspheres.
Fig. 3 is a graph representing physical properties and elemental composition of a metal-organic framework biocomposite prepared in example 1 of the present invention, wherein fig. 3(a) is an SEM image of the composite; FIG. 3(b) is a diagram showing the distribution of the elements of the composite de; FIG. 3(c) thermogravimetric analysis of the composite material; FIG. 3(d) is a graph showing the specific surface area and micropore analysis of the composite material.
FIG. 4 is a schematic diagram of electrochemical detection technology of Listeria monocytogenes in water and milk based on metal-organic framework biocomposites.
FIG. 5 is a transmission electron microscope image of an immunosensor in which an immunomagnetic bead captures Listeria monocytogenes and labels manganese metal organic frameworks.
Detailed Description
The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.
Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
Example 1 preparation of manganese Metal organic framework Biocomposite Material
The flow chart of the preparation method is shown in figure 1.
(1) Preparation of MnCl2·4H2O (439.2mg,2.22mM,3.3Equiv) and 2, 5-dihydroxyterephthalic acid (DHTP,133.2mg,0.672mM,1Equiv) were mixed in 60mL of a DMF-methanol-water (1:1:15, V/V/V) mixed solvent for 20 min. Transferring the mixed solution into a 100mL polytetrafluoroethylene reaction kettle sleeve, crystallizing at 140 ℃ for 24h, centrifugally purifying and drying to obtain a product which is brown yellow solid powder (shown as a spherical structure under an electron microscope),
the TEM image, the XPS characterization image, the XRD characterization image and the FTIR characterization image of the Mn-MOF-74 nano-microsphere prepared in the example are respectively shown in FIG. 2(a), FIG. 2(b), FIG. 2(c) and FIG. 2(d), and it can be seen that the particle size of the Mn-MOF-74 nano-microsphere is 200-300 nm; the microsphere mainly comprises three elements of manganese (Mn), carbon (C) and oxygen (O); the crystallinity of the Mn-MOF-74 microspheres is good, and the crystal faces of the Mn-MOF-74 microspheres corresponding to the characteristic diffraction angles (2 theta) of the crystals at 6.85 degrees and 11.90 degrees are (2, -1,0) and (3,0, 0); the infrared spectrum of the Mn-MOF-74 microsphere is 1591cm-1The absorption peak corresponds to the tensile vibration absorption peak of carboxylate (-COO) in organic ligand DHTP; at 1470cm-1And 1440cm-1The absorption peak at (a) is infrared absorption due to stretching vibration of C ═ C in the benzene ring, and belongs to skeleton vibration of the benzene ring, indicating that the organic ligand DHTP participates in the synthesis of Mn-MOF-74.
(2) 10.0mg of Mn-MOF-74 nanospheres were weighed into 1.0ml PBS buffer (pH 7.4), dispersed by sonication, 20mM (3.834g) EDC solid was added, vortexed for 10 minutes, washed by centrifugation, and the unreacted EDC was washed with PBS buffer and resuspended in 1.0ml PBS buffer.
(3) To 1.0mL of PBS buffer containing 10mM (1.151g) NHS (pH 7.4) was added 100. mu.g of monoclonal antibody (MAb) against Listeria monocytogenes.
(4) The MAb-NHS solution was vortexed with the carboxyl-activated Mn-MOF-74 nanosphere solution at 37 ℃ for 2 hours. The mixed product was centrifuged and washed with PBS buffer. The product was resuspended in 0.1% BSA-PBS blocking solution and stored at 4 ℃ at low temperature.
The SEM image, the elemental distribution map, the thermogravimetric analysis image, the specific surface area and the micropore analysis image of the manganese metal organic framework bio-composite material prepared in this example are shown in fig. 3(a), fig. 3(b), fig. 3(c) and fig. 3(d), respectively, and it can be seen that the surface of the Mn-MOF-74 microsphere is relatively flat and has good structural regularity (particle size 200-; the scanning electron microscope tandem energy spectrometer (EDS) shows the specific distribution condition of Mn, O and C elements in Mn-MOF-74; thermogravimetric analysis (TGA, FIG. 3(c)) showed that the Mn-MOF-74 mass decreased between 25 ℃ and 193 ℃ with a material weight loss of 26%. The mass loss caused by the removal of solvents (water and ethanol) in the surface and internal pores of the Mn-MOF-74 caused by high temperature is shown, and the pore structure characteristics and integrity of the material are still stable at the moment, which shows that the thermal stability of the Mn-MOF-74 is good within 193 ℃; the adsorption type of the nitrogen adsorption isotherm curve of Mn-MOF-74 is the combination of the type I and type IV adsorption isotherms. At a relative pressure (P/P)0) Below 0.01, there is a sharp increase in the adsorption curve, indicating the presence of a microporous structure in the Mn-MOF-74 structure.
Example 2 establishment of impedance detection mode of manganese metal organic framework biological composite material as immunosensor
Weighing 10mg manganese metal organic framework (Mn-MOF-74) nano microspheres, performing 10-fold gradient dilution by using deionized water to prepare an aqueous solution with the concentration range of 10 ng/mL-10 mu g/mL, adding a hydrogen peroxide solution with the concentration of 100 mu M, centrifuging, taking 10 mu L of a supernatant solution, dropwise adding the supernatant solution into a working electrode working area of an electrochemical workstation, and testing impedance signals of electrochemical sensors with different concentrations.
Through detection, the manganese metal organic framework biological composite material deionization with the concentration range of 10 ng/mL-10 mu g/mL can be knownThe impedance signal range in the aqueous solution is 1116-10493 omega (see table 1), the linear equation is Y1378X-738.68, R2＝0.9797。
The above results indicate that the impedance sensor is an electrochemical impedance probe with sensitive and stable signal response.
TABLE 1 reduction impedance test values of hydrogen peroxide solution for manganese metal organic framework biological composite materials with different concentrations
Example 3 detection application of manganese Metal organic framework immunosensor to Listeria monocytogenes in Water and milk
A schematic diagram of the electrochemical detection technology of the metal-organic framework-based biological composite material for listeria monocytogenes in water and milk is shown in FIG. 4.
To a concentration of 108The CFU/mL Listeria monocytogenes is added into water and milk samples in a pure culture way, and then the added samples are diluted in the water and milk samples in a gradient of 10 times to prepare the Listeria monocytogenes with the concentration range of 100～104CFU/mL of bacterial test solution. After adding 100. mu.g of immunomagnetic beads to each diluted sample and incubating for 20 minutes (40rpm, room temperature) on a shaker, the mixture was magnetically separated for 1 minute and then washed 3 times with PBS buffer and PBST buffer (0.05% Tween-20 in PBS buffer, V/V). Then 100 μ g of manganese metal organic framework biocomposite was added to the magnetically separated sample, followed by incubation on a shaker for 20 minutes (at 40rpm, room temperature) to form a sandwich complex of immunomagnetic beads-listeria monocytogenes-manganese metal organic framework immunosensor, whose tem image is shown in fig. 5. The complex was washed three times with PBST and deionized water, respectively.
Finally, the treated sandwich complexes were resuspended in a hydrogen peroxide solution (100 μ L, 0.1 mM). In 5 seconds, the Mn-MOF-74 in the compound is reduced and decomposed by the hydrogen peroxide solution, and manganese ions are released into the solution. mu.L of the supernatant after magnetic separation was used for the impedance test (sampling frequency 10 KHz).The impedance measurements were counted and analyzed (see Table 2 for the concentration of each different bacterium in the water and milk samples, 1.0X 10, respectively0～1.0×105CFU/mL)。
The test results show that the concentration range of the method in drinking water and milk is 1.0 x 100～1.0×104The minimum detection limit of the CFU/mL Listeria monocytogenes sample is 6.6CFU/mL and 4.1CFU/mL respectively, the addition recovery rate of the water sample is 90.2-95.0%, and the addition recovery rate of the milk sample is 87.9-92.7%.
TABLE 2 impedance test results for different Listeria monocytogenes bacterial concentrations in Water and milk
Claims (10)
1. A manganese metal organic framework biological composite material is formed by covalent coupling of Mn-MOF-74 nano microspheres and a specific biological recognition material.
2. The manganese metal-organic framework biocomposite material of claim 1, wherein: the particle size of the Mn-MOF-74 nano microspheres is 200-300 nm.
3. The manganese metal-organic framework biocomposite material of claim 1 or 2, wherein: the specific biological recognition material is a monoclonal antibody, a polyclonal antibody, a genetic engineering antibody or a nucleic acid aptamer.
4. A method for preparing the manganese metal organic framework biological composite material as claimed in any one of claims 1 to 3, comprising the following steps:
coupling the Mn-MOF-74 nano microspheres and the specific biological recognition material by adopting an active ester method.
5. The method of claim 4, wherein: the active ester method comprises the following steps:
1) dispersing the Mn-MOF-74 nano microspheres in a PBS buffer solution, adding EDC for reaction to obtain the Mn-MOF-74 nano microspheres activated by carboxyl;
2) dispersing the specific biological recognition material in PBS (phosphate buffer solution) of NHS (polyethylene glycol succinate), mixing with the system obtained in the step 1), reacting, and centrifuging to obtain the manganese metal organic framework biological composite material.
6. The production method according to claim 4 or 5, characterized in that: preparing the Mn-MOF-74 nano microspheres from manganese chloride tetrahydrate and 2, 5-dihydroxy terephthalic acid by a solvothermal reaction method;
the molar ratio of the manganese chloride tetrahydrate to the 2, 5-dihydroxy terephthalic acid is 2-4: 1.
7. the method of claim 6, wherein: the solvent thermal reaction method comprises the following steps:
dissolving the manganese chloride tetrahydrate and the 2, 5-dihydroxy terephthalic acid in a mixed solvent of DMF, methanol and water to obtain a mixed solution; and transferring the mixed solution into a reaction kettle sleeve, heating at 130-140 ℃ for 20-24 h for crystallization, and centrifuging to obtain the Mn-MOF-74 nano microspheres.
8. The use of the manganese metal organic framework biocomposite material of any one of claims 1-3 as or in the preparation of impedance immunosensors;
in the presence of reducing substances, the manganese metal organic framework biological composite material releases divalent manganese ions;
the reducing substance is hydrogen peroxide, vitamin C and/or glutathione.
9. The use of the manganese metal-organic framework biocomposite material as claimed in any one of claims-3 as an impedance immunosensor in the detection of food-borne pathogenic bacteria in any one of the following products:
1) ambient water;
2) domestic water and drinking water;
3) a dairy product;
4) a liquid beverage.
10. Use according to claim 9, characterized in that: the food-borne pathogenic bacteria comprise salmonella, staphylococcus aureus, escherichia coli and listeria monocytogenes;
the dairy product comprises milk, yogurt, fruit milk and reconstituted milk;
the liquid beverage comprises bottled water, wine, fruit juice and carbonated beverage.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110642668.6A CN113372439A (en) | 2021-06-09 | 2021-06-09 | Manganese metal organic framework biological composite material and application thereof in detection of food-borne pathogenic bacteria |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110642668.6A CN113372439A (en) | 2021-06-09 | 2021-06-09 | Manganese metal organic framework biological composite material and application thereof in detection of food-borne pathogenic bacteria |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN113372439A true CN113372439A (en) | 2021-09-10 |
Family
ID=77573400
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110642668.6A Pending CN113372439A (en) | 2021-06-09 | 2021-06-09 | Manganese metal organic framework biological composite material and application thereof in detection of food-borne pathogenic bacteria |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113372439A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114740067A (en) * | 2022-04-12 | 2022-07-12 | 济南大学 | Preparation and application of photoelectric sensor for non-immobilized ATP detection |
| CN115532295A (en) * | 2022-09-27 | 2022-12-30 | 苏州大学 | Application of nano material containing Zn-N-C active center in removing bacterial biofilm |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104749366A (en) * | 2013-12-31 | 2015-07-01 | 中国科学院上海微系统与信息技术研究所 | Method for rapidly detecting pathogenic bacteria |
| CN110006971A (en) * | 2019-03-12 | 2019-07-12 | 宁波大学 | A kind of preparation method and applications of the aptamer sensor of binary channels output detection food-borne pathogens |
| CN110161100A (en) * | 2019-05-23 | 2019-08-23 | 闽南师范大学 | The label-free electrochemical sensor preparation method of cardiac muscle troponin I and the detection method to cTnI |
| 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 |
-
2021
- 2021-06-09 CN CN202110642668.6A patent/CN113372439A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104749366A (en) * | 2013-12-31 | 2015-07-01 | 中国科学院上海微系统与信息技术研究所 | Method for rapidly detecting pathogenic bacteria |
| CN110006971A (en) * | 2019-03-12 | 2019-07-12 | 宁波大学 | A kind of preparation method and applications of the aptamer sensor of binary channels output detection food-borne pathogens |
| CN110161100A (en) * | 2019-05-23 | 2019-08-23 | 闽南师范大学 | The label-free electrochemical sensor preparation method of cardiac muscle troponin I and the detection method to cTnI |
| 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 |
Non-Patent Citations (3)
| Title |
|---|
| SIHAN WANG ET AL: "Gold interdigitated micro-immunosensor based on Mn-MOF-74 for the detection of Listeria monocytogens", 《BIOSENSORS AND BIOELECTRONICS》 * |
| 姚稼灏等: "多孔纳米材料固定化酶研究进展", 《微生物学通报》 * |
| 翟睿等: "金属有机框架材料的研究进展", 《色谱》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114740067A (en) * | 2022-04-12 | 2022-07-12 | 济南大学 | Preparation and application of photoelectric sensor for non-immobilized ATP detection |
| CN115532295A (en) * | 2022-09-27 | 2022-12-30 | 苏州大学 | Application of nano material containing Zn-N-C active center in removing bacterial biofilm |
| CN115532295B (en) * | 2022-09-27 | 2023-07-25 | 苏州大学 | Application of nanomaterial containing Zn-N-C active center in removal of bacterial biofilm |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Yang et al. | Heterogeneous nucleation and synthesis of carbon dots hybrid Zr-based MOFs for simultaneous recognition and effective removal of tetracycline | |
| Liu et al. | A nanosized metal–organic framework of Fe-MIL-88NH 2 as a novel peroxidase mimic used for colorimetric detection of glucose | |
| Varshney et al. | Magnetic nanoparticle-antibody conjugates for the separation of Escherichia coli O157: H7 in ground beef | |
| CN113372439A (en) | Manganese metal organic framework biological composite material and application thereof in detection of food-borne pathogenic bacteria | |
| CN110586052B (en) | Preparation and application of magnetic composite porous network adsorption material | |
| Wang et al. | Rapid and ultrasensitive detection of Salmonella typhimurium using a novel impedance biosensor based on SiO2@ MnO2 nanocomposites and interdigitated array microelectrodes | |
| CN110108881B (en) | Preparation method and application of bifunctional biosensor HRP @ ZIF-8/DNA | |
| CN103308373A (en) | Magnetic bead separation method of escherichia coli O157 | |
| CN110841612A (en) | Magnetic NH2Preparation and application of-MOFs nano material | |
| Deng et al. | Poly-l-lysine-functionalized magnetic beads combined with polymerase chain reaction for the detection of Staphylococcus aureus and Escherichia coli O157: H7 in milk | |
| Ma et al. | Copper (II) ions-immobilized virus-like hollow covalent organic frameworks for highly efficient capture and sensitive analysis of amyloid beta-peptide 1–42 by MALDI-MS | |
| CN103293297A (en) | Method for quickly separating salmonella | |
| Yan et al. | Dual recognition strategy for the rapid and precise detection of Bacillus cereus using post-modified nano-MOF and aptamer | |
| Yue et al. | Rapid detection of Salmonella in milk by a nuclear magnetic resonance biosensor based on the streptavidin-biotin system and O-carboxymethyl chitosan target gadolinium probe | |
| Duan et al. | Preparation of immunomagnetic iron-dextran nanoparticles and application in rapid isolation of E. coli O157: H7 from foods | |
| Li et al. | A functionalized magnetic graphene-based MOFs platform as the heterogeneous mimic enzyme sensor for glucose detection | |
| Yang et al. | Mesoporous yolk–shell structure molecularly imprinted magnetic polymers for the extraction and detection of 17β-estradiol | |
| Xu et al. | Sandwich-type colorimetric assay based on molecularly imprinted polymers and boronic acid functionalized Ni-Fe-MOF nanozyme for sensitive detection of allergen ovalbumin | |
| CN106947756A (en) | The method of quick Magneto separate bacillus cereus | |
| Hu et al. | A robust and facile colorimetric aptasensor for the detection of Salmonella Typhimurium based on the regulation of Fe3O4@ Cu@ PCPy yolk-shell nanozyme activity | |
| KR101372249B1 (en) | Method for manufacturing the magnetic mesoporous silica, the high efficient bacteria separation and detection method using thereof | |
| CN114774120B (en) | Based on iron-doped carbon dot combined magnetic Fe 3 O 4 Preparation method and application of @ PDA biosensor | |
| CN114487082B (en) | Pretreatment method for sample for measuring trace elements in milk powder | |
| CN106967709A (en) | The method that the magnetic nano-particle fast enriching of antibiotics modification separates Listeria monocytogenes | |
| Rashidiani et al. | Application of gold core-shell magnetic nanoparticles immunosensor for detection of vibrio cholera |
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 | ||
| WD01 | Invention patent application deemed withdrawn after publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20210910 |