CN103197070A - Quick detection method of NMR (nuclear magnetic resonance) food-borne pathogenic bacteria based on gamma-Fe2O3 nano-particle indirect inrichment - Google Patents
Quick detection method of NMR (nuclear magnetic resonance) food-borne pathogenic bacteria based on gamma-Fe2O3 nano-particle indirect inrichment Download PDFInfo
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Abstract
The invention provides a quick detection method of NMR (nuclear magnetic resonance) food-borne pathogenic bacteria based on gamma-Fe2O3 nano-particle indirect enrichment, and belongs to the technical field of food safety pathogenic bacteria quick detection. Depending on an established nuclear magnetic resonance detection method for pathogenic bacteria in a food liquid sample, anti-immunomagnetic beads 2 are prepared by utilizing a gamma-Fe2O3 nano-particle material to specifically enrich an anti-mark target strain 1, whether a target pathogenic bacterium exists in the sample or not can be detected by utilizing the influence of paramagnetic and superparamagnetic characteristics of gamma-Fe2O3 nano-particles to the relaxation time of a nuclear magnetic resonance deamplification signal. The gamma-Fe2O3 nano-particles are enrichment means or a detecting probe. Different particular corresponding relationships are that a linear relationship is indicated by the paramagnetic immunomagnetic beads under a certain condition, namely the value of T2/T1 of the sample is smaller while the content of immunomagnetic beads is larger, so that a target strain can be quantitatively detected in a certain range. The method can be used for quickly detecting hazard pathogenic bacteria in a food sample, and can be used for quickly screening large-scale samples to be detected.
Description
Technical field
The present invention relates to the method for quick of a kind of pathogenic bacteria, relate in particular to a kind of based on γ-Fe
2O
3The NMR food-borne pathogens method for quick of the indirect enrichment of nano particle.
Background technology
Ultimate principle: monoclonal antibody or antigen molecule and enzyme molecule are by covalent bonds, and this combination can not change immunological characteristic and the biologically active of monoclonal antibody, antigen and enzyme, and specific monoclonal antibody only can be combined with specific antigen.The principle steps that it is main: 1. in sample to be checked, add the 1st antibody of object bacteria, if there is object bacteria, then closes with 1 resistive connection and form 1 anti-compound; 2. utilize magnetic
γ-Fe
2O
3Nano particle, surface-functionalized by the modified antibodies realization, be the 2 anti-magnetic bead surfaces that are coupled at 1 anti-antibody, form the specific immunity magnetic bead, and unnecessary avtive spot is sealed.2. the specificity 1 anti-monoclonal antibody with another part object bacteria adopts certain method to be fixed on surface of solid phase carriers, and with unnecessary avtive spot sealing.3. 2 anti-specific immunity magnetic beads with the preparation of the 2nd step join in the sample to be checked in the 1st step, be used for grasping the enrichment object bacteria, by externally-applied magnetic field magnetic bead is separated, at this moment, combine 1 anti-compound of object bacteria and do not have excessive 1 of combining target bacterium to resist and all can close with 2 resistive connections of magnetic bead surfaces, still mix.4. with the above-mentioned magnetic bead that mixes, be added to the surface of solid phase carriers in the 2nd step, the magnetic bead that has then grasped object bacteria will be combined with surface of solid phase carriers 1 anti-generation specificity, form double antibodies sandwich, can will the magnetic bead wash-out of combination not take place with aseptic washed with de-ionized water.5. adopt eluant, eluent that the specific nano immunomagnetic beads of the combination on the immobilization carrier is washed again, the magnetic separation washes ion, solvent.If this part magnetic bead exists, grasped the magnetic bead of object bacteria exactly.Because
γ-Fe
2O
3Nano particle has paramagnetic and super paramagnetic characteristic, and is very responsive for magnetic resonance tool, for other molecules, trace
γ-Fe
2O
3Nano particle can significantly reduce spin-lattice relaxation parameter T1 and the spin spin relaxation time T2 of aseptic deionized water, and aseptic deionized water is under certain even field intensity, and T1/T2 fixes.Eluent is placed nuclear magnetic resonance analyser, contrast with aseptic deionized water control group.The explanation that the reduction of T1/T2 value significantly takes place has magnetic bead to exist, thereby has pathogenic bacteria to detect in the side light food samples.Magnetic bead content and T1/T2 value reduce proportional.By mark-on, can quantitatively detect object bacteria.In this method
γ-Fe
2O
3The nano particle magnetic bead is the means of separation and concentration, simultaneously
γ-Fe
2O
3The paramagnetic that nano particle has and super paramagnetic characteristic can be used as the probe of quantitative detection again.The major advantage of this method is exactly quick, highly sensitive.Cultivate time of 2-3 days even several days with respect to the microorganisms of pathogenic bacteria, the method depends primarily on the The pretreatment time, and magnetic resonance detection only needs a few minutes.Therefore, can do the positive-selecting of extensive sample to be checked with this method, the positive sample that detects also needs to cultivate with microorganism and prove conclusively.At present, also do not have this method of bibliographical information both at home and abroad, but the report that adopts immunomagnetic beads to carry out the enrichment of pathogenic bacteria, the enrichment of object etc. still is a lot, but does not all adopt nuclear magnetic resonance to do further detection.
Summary of the invention
The purpose of this invention is to provide a kind of based on γ-Fe
2O
3The NMR food-borne pathogens method for quick of the indirect enrichment of nano particle, be used for various food samples is estimated, this method is a kind of objective method that effectively detects harmful pathogenic bacteria in the food, thereby has reduced the screening time of the harmful pathogenic bacteria of food samples to a certain extent greatly.
A kind of based on
γ-Fe
2O
3The nuclear magnetic resonance technique fast detecting of the indirect enrichment of nano particle goes out the method for the harmful pathogenic bacteria in the food.Utilize nuclear magnetic resonance analyser to the response susceptibility of paramagnetic, super paramagnetic material, propose the NMR (Nuclear Magnetic Resonance) relaxation parameter change with
γ-Fe
2O
3The correlativity index of the super paramagnetic immunomagnetic beads of nano particle content.Different pathogenic bacteria detect the lower limit difference.
This method depends on harmful pathogenic bacteria characteristic immunomagnetic beads enrichment, the separation in the food samples of can be used for of foundation, and the angle from the NMR (Nuclear Magnetic Resonance) relaxation signal parameter changes detects the harmful pathogenic bacteria in the sample; Adopt 1 anti-mark of object bacteria to form 1 anti-compound; Adopt the i.e. 2 super paramagnetic immunomagnetic beadses that resist of coupling 1 antiantibody, the specificity pathogenic bacteria in the sample can be carried out enrichment and separation; Because nuclear magnetic resonance analyser SPIN-LATTICE RELAXATION efficient and spin-spin relaxation efficient are right
γ-Fe
2O
3Nano particle is very responsive, namely in deionized water, has the super paramagnetic of trace
γ-Fe
2O
3Nano particle, then the spin-lattice relaxation time of water (T1) and/spin-spin relaxation (T2) will significantly descend.Under certain condition, the paramagnetic characteristic of super paramagnetic immunomagnetic beads reduces the relaxation decay signal T2/T1 generation linearity of nuclear magnetic resonance.By quantitatively detecting the immunomagnetic beads content in the sample, thereby can quantitatively go out the harmful pathogenic bacteria content in the food samples indirectly.Detected pathogenic bacteria content and magnetic bead content linear dependence, degree of fitting is better.Final is tie with the corresponding relation between super paramagnetic immunomagnetic beads and pathogenic bacteria, determines the pathogenic bacteria clump count in the food samples.
The present invention is achieved in that step is as follows:
1) in sample to be checked, adds the 1st antibody of object bacteria, if there is object bacteria, then closes with 1 resistive connection and form 1 anti-compound;
2) the 2 anti-i.e. preparations of the antibody immune magnetic beads of the 1st antibody;
3) object bacteria specificity 1 anti-monoclonal antibody is fixed on surface of solid phase carriers;
4) immunomagnetic beads enrichment aimed strain, and separate: the 2 anti-immunomagnetic beadses that the 2nd step was made add the 1st sample to be checked that goes on foot, fully mix concussion, grasp after the object bacteria by applying externally-applied magnetic field, then magnetic bead just is pooled to magnetic field on one side, siphon away supernatant and then can isolate magnetic bead if in the sample to be checked object bacteria is arranged, then by the enrichment of magnetic bead institute, add the magnetic bead suspension that a small amount of aseptic deionized water then forms object bacteria; At this moment, combine 1 anti-compound of object bacteria and do not have excessive 1 of combining target bacterium to resist and all can close with 2 resistive connections of magnetic bead surfaces, still mix.
4) the magnetic bead suspension with enrichment is added on the solid phase carrier of having fixed 1 anti-monoclonal antibody of the 3rd step making, if exist object bacteria then to form double antibodies sandwich; With aseptic washed with de-ionized water, the magnetic bead that does not then grab object bacteria is just washed off, if there is not object bacteria, then all magnetic beads are all washed off;
5) afterwards, washing with the magnetic bead of eluant, eluent with the double antibodies sandwich on the fixed head, separate the method for magnetic bead with externally-applied magnetic field and fall ion and solvent with aseptic washed with de-ionized water, is exactly the magnetic bead of catching object bacteria if also there is magnetic bead;
6) this part magnetic bead, the suspension that adds aseptic deionized water formation magnetic bead, carrying out the relaxation time of nuclear magnetic resonance measures, be blank with aseptic deionized water, the relaxation time T1/T2 of the suspension that records compares aseptic deionized water remarkable reduction, and then explanation contains magnetic bead, thereby in the indirect proof sample target pathogenic bacteria is arranged, the amount of magnetic bead and the decline of T1/T2 are proportional, can quantitatively go out the amount of object bacteria by quantitative magnetic bead indirectly.
Described NMR nano-probe is
γ-Fe
2O
3Nano particle, nanometer particle size is less than 1000 nanometers.
Described object bacteria finally detect evaluation method based on the variation of the relaxation time characterisitic parameter of nuclear magnetic resonance technique.
Described relaxation time characteristic refers to spin-lattice relaxation time (T1) and spin spin relaxation time (T2).
Described relaxation time characteristic, T1 adopts 180 °-τ-90 ° of pulse method to measure, and T2 adopts cpmg sequence row method to measure.
The monoclonal antibody of described 1 anti-finger object bacteria, the antibody of 2 anti-finger the 1st antibody can be that monoclonal antibody also can be to resist more.
Beneficial effect of the present invention: the invention provides a kind of objective fast detecting and go out the method for the harmful pathogenic bacteria in the food, it is characterized in that depending on the magnetic resonance detection method that can be used for the detection of superparamagnetic immunomagnetic beads of foundation.This method can objectively detect harmful pathogenic bacteria in the food effectively, confirms that than the biological culture of pathogenic bacteria this method has the advantage of fast detecting, can be used for the rapid screening of extensive sample.
Embodiment
Example 1
Measure it in the check food samples and whether contain harmful pathogenic bacteria---Listeria.
1. 2 anti-immunomagnetic beads preparations: the anti-IgG monoclonal antibody of the listerial rabbit of 1 anti-employing, 2 is anti-for listerial goat anti-rabbit igg, can be that monoclonal antibody also can be to resist more.2 anti-immunomagnetic beads preparations: the coupling of magnetic bead and monoclonal antibody.Adopt commercial carboxylated magnetic bead, or adopt microemulsion method to prepare γ-Fe voluntarily
2O
3Nano microsphere is modified to carboxylated magnetic bead or amination magnetic bead by bag.Be example with carboxylated magnetic bead, itself and 2 anti-coupling steps: 1. clean: get the carboxylated magnetic bead of 5mg respectively in the 5mL centrifuge tube, 2mL PB(0.02M pH=6.0) washes 2 times, and 2mL MES(0.05M is used in the washing back, and is pH=6.0) resuspended; 2. activation: the ultrasonic magnetic bead that makes disperses, and adds 1mg EDC more respectively, 1mg NHSS(0.05M, pH=6.0 MES dissolving), 25 ℃ of activation 2h, gyroscope 15r/min keeps suspended state; 3. coupling: magnetic separates, inhale and remove supernatant, 2mL PB(0.02M, pH=7.4) washing once and be transferred to new centrifuge tube, 3mL PB(0.02M, pH=7.4) resuspended, the ultrasonic magnetic bead that makes disperses.Respectively 275 μ g the 2nd antibody goat anti-rabbit igg is joined again and activate in the magnetic bead, 25 ℃ of coupling 3h, gyroscope 15r/min keeps suspended state; 4. sealing: magnetic separates, and takes out supernatant (protein content in the supernatant is to be checked), adding 1mg/mL monoethanolamine and 1mg/mL BSA sealing (0.02M, the PB dissolving of pH=7.4, and regulate pH to neutrality), 25 ℃ of sealing 2h, gyroscope 15r/min keeps suspended state; 5. preserve: 0.01M PBS washing magnetic bead 2 times, magnetic separates, and removes supernatant, and is 0.5%BSA) resuspended with 3mL pH7.4 PB(0.02M, 0.02% NaN3, is stored in 4 ℃.The immunomagnetic beads of preparation is standby.
2. 1 anti-monoclonal antibody is fixed: can adopt conventional ELISA Plate fixing means, also can adopt following method.With clean cover glass 5 * 5mm
2Square, coating machine spray one deck Cr (2 –, 4 nm) earlier are fixing golden in order to help.Be used in surface sputtering again and spray one deck nm of gold, adopt 200 microlitres, 2 mmol disulfide group-succinimide-propionic esters (DSP) that nm of gold is modified (DMSO, dimethyl sulfoxide (DMSO) dilution DSP) again.Add the anti-IgG monoclonal of listerial rabbit monoclonal antibody, be about to 100
L100
The g/mL monoclonal antibody be fixed on the glass plate by e and 37 ℃ hatch 45 min.Add 1% bovine serum albumin (BSA), 22 ℃, 1 hour, remaining avtive spot on the plate is sealed and drying.
3. food samples is carried out pre-service, adopt the FDA enrichment in case of necessity, sample is filtered, increases pre-service such as bacterium activation, obtain sample to be checked.The 1st antibody, the anti-IgG of listerial rabbit will be added in the sample to be checked.If have the Listeria in the sample to be checked, will with 1 anti-formation 1 anti-compound.Fully shake after the listerial goat anti-rabbit igg magnetic bead of the 1st the 2nd antibody that makes of step added.Last magnetic force frame separates magnetic bead, adds the suspension that a small amount of aseptic deionized water obtains magnetic bead.At this moment, if the target Listeria is arranged in the sample to be checked, then pass through the interaction of the 2nd antibody and the 1st antibody, thereby catch this compound, reached the purpose of object bacteria enrichment.At this moment, combine 1 anti-compound of object bacteria and do not have excessive 1 of combining target bacterium to resist and all can close with 2 resistive connections of magnetic bead surfaces, still mix.The magnetic bead suspension of this enrichment is added on the prepared 1 anti-fixed head of the 2nd step, then combine in the magnetic bead emulsion listerial magnetic bead further the monoclonal antibody generation specificity on fixed head be combined, form the double antibodies sandwich structure.Just can will do not have in conjunction with listerial magnetic bead flush away with aseptic washed with de-ionized water this moment.Just only combine listerial magnetic bead what fixed head was left.
4. with eluent (methyl alcohol etc.) the listerial magnetic bead that combines on the fixed head is eluted.Last magnetic force frame, the separation magnetic bead also cleans 1-2 time, with ion, solvent flush away.The solution that obtains is with T1 and the T2 of nuclear magnetic resonance analyser (NMR20, Niu Mai company) mensuration solution.Be blank with aseptic deionized water, the T1/T2 that solution records compares with blank, and there were significant differences, and illustrating has magnetic bead to exist in the solution, thereby in the explanation sample Listeria is arranged, and the amount of magnetic bead and the drop-out value of T1/T2 are proportional.The bright magnetic bead of more speaking more that descends is more many, thereby the side light Listeria is more many, can quantitatively detect the number of object bacteria in the sample by the mark-on checking.
Embodiment
Example 2
Measure food samples and whether contain harmful pathogenic bacteria Escherichia coli O 157: H7.
1.2 anti-immunomagnetic beads preparation: the anti-IgG monoclonal antibody of rabbit of 1 anti-employing O157:H7,2 anti-ly are the goat anti-rabbit igg of O157:H7, can be that monoclonal antibody also can be to resist more.2 anti-immunomagnetic beads preparations: the coupling of magnetic bead and monoclonal antibody.Adopt commercial carboxylated magnetic bead, or adopt microemulsion method to prepare γ-Fe voluntarily
2O
3Nano microsphere is modified to carboxylated magnetic bead or amination magnetic bead by bag.Be example with carboxylated magnetic bead, itself and 2 anti-coupling steps: 1. clean: get the carboxylated magnetic bead of 5mg respectively in the 5mL centrifuge tube, 2mL PB(0.02M pH=6.0) washes 2 times, and 2mL MES(0.05M is used in the washing back, and is pH=6.0) resuspended; 2. activation: the ultrasonic magnetic bead that makes disperses, and adds 1mg EDC more respectively, 1mg NHSS(0.05M, pH=6.0 MES dissolving), 25 ℃ of activation 2h, gyroscope 15r/min keeps suspended state; 3. coupling: magnetic separates, inhale and remove supernatant, 2mL PB(0.02M, pH=7.4) washing once and be transferred to new centrifuge tube, 3mL PB(0.02M, pH=7.4) resuspended, the ultrasonic magnetic bead that makes disperses.Respectively 275 μ g the 2nd antibody is joined again and activate in the magnetic bead, 25 ℃ of coupling 3h, gyroscope 15r/min keeps suspended state; 4. sealing: magnetic separates, and takes out supernatant (protein content in the supernatant is to be checked), adding 1mg/mL monoethanolamine and 1mg/mL BSA sealing (0.02M, the PB dissolving of pH=7.4, and regulate pH to neutrality), 25 ℃ of sealing 2h, gyroscope 15r/min keeps suspended state; 5. preserve: 0.01M PBS washing magnetic bead 2 times, magnetic separates, and removes supernatant, and is 0.5%BSA) resuspended with 3mL pH7.4 PB(0.02M, 0.02% NaN3, is stored in 4 ℃.The immunomagnetic beads of preparation is standby.
2. 1 anti-monoclonal antibody is fixed: can adopt conventional ELISA Plate fixing means, also can adopt following method.With clean cover glass 5 * 5mm
2Square, coating machine spray one deck Cr (2 –, 4 nm) earlier are fixing golden in order to help.Be used in surface sputtering again and spray one deck nm of gold, adopt 200 microlitres, 2 mmol disulfide group-succinimide-propionic esters (DSP) that nm of gold is modified (DMSO, dimethyl sulfoxide (DMSO) dilution DSP) again.Add the anti-IgG monoclonal antibody of O157:H7 rabbit, be about to 100
L100
The g/mL monoclonal antibody be fixed on the glass plate by e and 37 ℃ hatch 45 min.Adding bovine serum albumin seals remaining avtive spot on the plate and drying.
3. food samples is carried out pre-service, sample is filtered, increases pre-service such as bacterium activation, obtain sample to be checked.The 1st antibody, the anti-IgG of the rabbit of O157:H7 will be added in the sample to be checked.If have O157:H7 in the sample to be checked, will with 1 anti-formation 1 anti-compound.Fully shake after the goat anti-rabbit igg antibody magnetic bead of the 1st the 2nd antibody O157:H7 that makes of step added.Last magnetic force frame separates magnetic bead, adds the emulsion that low amounts of water obtains magnetic bead.At this moment, if target Escherichia coli O 157: H7 is arranged in the sample to be checked, then pass through the interaction of the 2nd antibody and the 1st antibody, thereby catch this compound, reached the purpose of object bacteria enrichment.At this moment, combine 1 anti-compound of object bacteria and do not have excessive 1 of combining target bacterium to resist and all can close with 2 resistive connections of magnetic bead surfaces, still mix.The magnetic bead suspension of this enrichment is added on the prepared 1 anti-fixed head of the 2nd step, and the magnetic bead that then combines Escherichia coli O 157: H7 in the magnetic bead suspension further monoclonal antibody generation specificity on fixed head is combined, and forms the double antibodies sandwich structure.This moment is with aseptic washed with de-ionized water, just can be with not in conjunction with Escherichia coli O 157: the magnetic bead flush away of H7.At the remaining magnetic bead that just only combines Escherichia coli O 157: H7 of fixed head.
With eluent (methyl alcohol etc.) with the Escherichia coli O 157 that combines on the fixed head: the magnetic bead of H7 elutes.Last magnetic force frame separates magnetic bead and with aseptic washed with de-ionized water 1-2 time, with ion, solvent flush away.The solution that obtains, with nuclear magnetic resonance analyser (NMR20, Niu Mai company) T1 and the T2 of mensuration solution, be blank with the deionized water, the T1/T2 that solution records is with relatively blank, and there were significant differences, and illustrating has magnetic bead to exist in the solution, thereby Escherichia coli O 157: H7 is arranged in the explanation sample, and the amount of magnetic bead and the drop-out value of T1/T2 are proportional.The bright magnetic bead of more speaking more that descends is more many, thereby side light Escherichia coli O 157: H7 is more many, can quantitatively detect the number of object bacteria in the sample by the mark-on checking.
Claims (7)
1. one kind based on γ-Fe
2O
3The NMR food-borne pathogens method for quick of the indirect enrichment of nano particle, its characterization step is as follows:
1) 1 resistive connection that will detect object bacteria and object bacteria closes formation 1 anti-compound;
2) 1 antiantibody of detection object bacteria, i.e. the preparation of 2 anti-specific immunity magnetic beads;
3) with object bacteria 1 anti-monoclonal antibody specific fixing on solid phase carrier;
4) immunomagnetic beads enrichment aimed strain, and separate: the 2 anti-immunomagnetic beadses that the 2nd step was made joined for the 1st step and combine 1 anti-sample to be checked, fully mix concussion, grasp after the object bacteria by applying externally-applied magnetic field, then magnetic bead just is pooled to magnetic field on one side, siphon away supernatant and then can isolate magnetic bead, if in the sample to be checked object bacteria is arranged, object bacteria is at first closed the formation compound with 1 resistive connection, after adding 2 diamagnetic pearls, by 2 anti-and 1 anti-interactions, thereby catch 1 anti-compound, by the enrichment of magnetic bead institute, add the magnetic bead suspension that a small amount of aseptic deionized water then forms object bacteria;
4) the magnetic bead suspension of enrichment being added to the 3rd step has fixed on the solid phase carrier of 1 anti-monoclonal antibody, if exist object bacteria then to form double antibodies sandwich, with aseptic washed with de-ionized water, the magnetic bead that does not then grab object bacteria is just washed off, if there is not object bacteria, then all magnetic beads are all washed off;
5) afterwards, washing with the magnetic bead of eluant, eluent with the double antibodies sandwich on the fixed head, with the method that externally-applied magnetic field separates magnetic bead, fall ion and solvent with aseptic washed with de-ionized water, is exactly the magnetic bead of catching object bacteria if also there is magnetic bead;
6) this part magnetic bead, the suspension that adds aseptic deionized water formation magnetic bead, carrying out the relaxation time of nuclear magnetic resonance measures, under fixing field intensity, the T1/T2 of aseptic deionized water is a steady state value, be blank with aseptic deionized water, the relaxation time T1/T2 of the suspension that records compares aseptic deionized water remarkable reduction, then explanation contains magnetic bead, thereby the target pathogenic bacteria are arranged in the indirect proof sample, the amount of magnetic bead and the decline of T1/T2 are proportional, can quantitatively go out the amount of object bacteria by adding the scalar quantity magnetic bead indirectly.
2. according to claim 1 based on γ-Fe
2O
3The NMR food-borne pathogens method for quick of the indirect enrichment of nano particle is characterized in that described NMR nano-probe is
γ-Fe
2O
3Nano particle, nanometer particle size is less than 1000 nanometers.
3. according to claim 1 based on γ-Fe
2O
3The NMR food-borne pathogens method for quick of the indirect enrichment of nano particle, what it is characterized in that described object bacteria finally detects the variation of evaluation method based on the relaxation time of nuclear magnetic resonance technique.
4. according to claim 1 based on γ-Fe
2O
3The NMR food-borne pathogens method for quick of the indirect enrichment of nano particle is characterized in that described NMR relaxation time characteristic, refers to spin-lattice relaxation time (T1) and spin spin relaxation time (T2).
5. according to claim 1 based on γ-Fe
2O
3The NMR food-borne pathogens method for quick of the indirect enrichment of nano particle is characterized in that described relaxation time characteristic, and T1 adopts 180-τ-90 pulse method to measure, and T2 adopts cpmg sequence row method to measure.
6. according to claim 1 based on γ-Fe
2O
3The NMR food-borne pathogens method for quick of the indirect enrichment of nano particle is characterized in that object bacteria earlier and 1 resistive connection closes the formation compound, and with 2 anti-specific immunity enrichment with magnetic bead, separation, 1 has resisted the signal amplification.
7. according to claim 1 based on γ-Fe
2O
3The NMR food-borne pathogens method for quick of the indirect enrichment of nano particle is characterized in that 1 anti-finger object bacteria monoclonal antibody, and the antibody of 2 anti-finger the 1st antibody can be that monoclonal also can be polyclonal antibody.
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