CN103245686A - Rapid NMR (nuclear magnetic resonance) food-borne pathogenic bacteria detection method based on indirect enrichment of paramagnetic nano-Ni probe - Google Patents
Rapid NMR (nuclear magnetic resonance) food-borne pathogenic bacteria detection method based on indirect enrichment of paramagnetic nano-Ni probe Download PDFInfo
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Abstract
The invention relates to a rapid NMR food-borne pathogenic bacteria detection method based on indirect enrichment of a paramagnetic nano-Ni probe, and belongs to the field of food pathogenic bacteria rapid detection technologies. Depending on a constructed NMR method capable of being used for detecting pathogenic bacteria in a food liquid sample, the rapid detection method comprises the following steps: capturing a target bacterium by using a first antibody; coating the paramagnetic nano-Ni probe with a second antibody which is an antibody of the first antibody, and performing enrichment and separation on the target bacterium; and detecting whether the sample contains the target pathogenic bacterium according to the influences of a paramagnetic property of nano-Ni on relaxation time of an NMR deamplification signal. The different concrete corresponding relationship is that the paramagnetic nano-Ni probe shows a linear relationship under a certain condition, namely, the larger the nano-Ni content is, the smaller the T2/T1 value of the sample is, so that the target bacterium can be detected quantitatively within a certain range. The method provided by the invention can be used for rapid detection on harmful pathogenic bacteria in food samples, and can be used for rapidly screening a large amount of to-be-detected samples.
Description
Technical field
The present invention relates to the fast detecting side of a kind of pathogenic bacteria, relate in particular to a kind of NMR food-borne pathogens method for quick based on the indirect enrichment of paramagnetic nano Ni probe.
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 Ni material has ferromagnetism, when particle diameter little of to a certain degree the paramagnetic characteristic occurring.Do not have magnetic when namely not having externally-applied magnetic field, and when externally-applied magnetic field is arranged, show certain magnetic, can be used for magnetic and separate.Simultaneously, paramagnet is very remarkable to the influence of NMR signal, and the paramagnet of trace will make NMR signal show variation.Therefore can make up paramagnetic specificity Ni nano-probe biology sensor, do detection from the angle of magnetic resonance.
The principle steps that it is main: 1. add the 1st antibody that detects object bacteria in sample, if having object bacteria in the sample, then the combination by antibody antigen forms 1 anti-compound, and resisted the signal amplification this moment 1.2. buy paramagnetic nano Ni material from the market, also can prepare nano level Ni by additive method.Use silane coupling agent, its general formula is: Y (CH
2) nSiX
3Herein, n is 0-3; X is hydrolyzable group; Y is organo-functional group.X is chloro, methoxyl, ethoxy, acetoxyl group etc. normally, generates silanol (Si (OH) during these group hydrolysis
3), and be combined with dead matter, form siloxane.Y is vinyl, amino, epoxy radicals, methacryloxy, sulfydryl.These reactive groups can the combination with the organic substance reaction.Therefore, by using silane coupling agent, can between the interface of dead matter and organic substance, erect " molecule bridge ", the material of two kinds of character great disparities is linked together improve the effect of performance of composites and increase bonding strength.Can realize surface-functionalizedly by modified antibodies, form the specific immunity probe, seal unnecessary avtive spot again.Because nanometer Ni probe has the paramagnetic characteristic, therefore, can separate the antibody that does not have on the connection by externally-applied magnetic field.What nanometer Ni material was modified is the antibody of the 1st antibody, and namely 2 is anti-.3. adopt certain method to be fixed on surface of solid phase carriers object bacteria specificity the 1st antibody, and unnecessary avtive spot sealing is standby.4. the sample of handling in the 1st step adds the paramagnetic nano Ni probe that the 2nd step made, and fully mixes concussion reaction a period of time, grasp the after-applied externally-applied magnetic field of object bacteria, because nanometer Ni has the paramagnetic characteristic, the Ni probe can gather magnetic field on one side, siphons away supernatant and then can isolate probe.If object bacteria is arranged in the sample to be checked, then at first can be when the 1st step and 1 anti-formation 1 anti-compound, 1 anti-compound can be again with 2 of detecting probe surface resist compound, by externally-applied magnetic field by enrichment, separation.After separating, washing, magnetic adds the paramagnetic nano Ni probe suspension that a small amount of aseptic deionized water then forms object bacteria.At this moment, the excess probe of catching object bacteria and not catching object bacteria still mixes.5. with the above-mentioned probe that mixes, be added to the surface of solid phase carriers in the 3rd step, the probe that has then grasped object bacteria will be combined with surface of solid phase carriers monoclonal antibody generation specificity, form double antibodies sandwich, can will the probe wash-out of combination not take place with aseptic washed with de-ionized water.6. adopt eluant, eluent that the specific nano immunological probe of the combination on the immobilization carrier is washed again, the method for separating with magnetic washes ion, solvent.If this part probe exists, grasped the probe of object bacteria exactly.Because Ni has the paramagnetic characteristic, very responsive for the resonance instrument, for other molecules, the Ni of trace 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 probe to exist, thereby has pathogenic bacteria to detect in the side light food samples.Probe content and T1/T2 value reduce proportional.By mark-on, can quantitatively detect object bacteria.The Ni probe is the means of separation and concentration in this method, and the paramagnetic characteristic that has of Ni simultaneously can be used as the probe of quantitative detection again.The major advantage of this method is exactly quick, highly sensitive.Cultivate the 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.All food standards, pathogenic bacteria all must not detect.Therefore, can do the positive-selecting of extensive sample to be checked with this method, can quantitatively detect to a certain extent.At present, also there is not this method of bibliographical information both at home and abroad.
Summary of the invention
A kind of NMR food-borne pathogens method for quick based on the indirect enrichment of paramagnetic nano Ni probe is 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 NMR food-borne pathogens method for quick based on the indirect enrichment of paramagnetic nano Ni probe, utilize nuclear magnetic resonance analyser to the response susceptibility of paramagnetic material, propose the correlativity index of the variation of NMR (Nuclear Magnetic Resonance) relaxation parameter and Ni nano particle paramagnetic immunological probe content.Different pathogenic bacteria detect the lower limit difference.
This method depends on harmful pathogenic bacteria specificity paramagnetic nano Ni probe 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.The paramagnetic nano Ni probe of monoclonal antibody specific that adopted coupling can carry out enrichment with the specificity pathogenic bacteria in the sample.Because nuclear magnetic resonance analyser SPIN-LATTICE RELAXATION efficient and spin-spin relaxation efficient are very responsive to the Ni nano particle, namely in deionized water, the paramagnetic Ni nano particle that has trace, then the spin-lattice relaxation time of water (T1) and/spin-spin relaxation (T2) will significantly descend.Under certain condition, the paramagnetic characteristic of paramagnetic immunological probe reduces the relaxation decay signal T2/T1 generation linearity of nuclear magnetic resonance.By quantitatively detecting the immunological probe content in the sample, thereby can quantitatively go out the harmful pathogenic bacteria content in the food samples indirectly.Detected pathogenic bacteria content and probe content linear dependence, degree of fitting is better.Final is tie with the corresponding relation between paramagnetic immunological probe and pathogenic bacteria, determines the pathogenic bacteria clump count in the food samples.And all food standards, pathogenic bacteria all must not detect.Therefore, this method can be done the positive-selecting of extensive sample to be checked, can quantitatively detect to a certain extent.
The present invention is achieved in that step is as follows:
1) add the 1st antibody that detects object bacteria in sample, if having object bacteria in the sample, then the combination by antibody antigen forms 1 anti-compound.
2) antibody of the 1st antibody of detection object bacteria, namely 2 anti-bags are by the preparation of paramagnetic nano Ni probe;
3) with object bacteria specificity the 1st antibody fixing standby on solid phase carrier.
4) enrichment aimed strain, and separate: the sample of handling in the 1st step, add the paramagnetic nano Ni probe that the 2nd step made, fully mix concussion reaction a period of time, grasp the after-applied externally-applied magnetic field of object bacteria, because the paramagnetic characteristic of nanometer Ni, then the Ni probe just is pooled to magnetic field on one side, siphons away supernatant and then can isolate probe.If object bacteria is arranged in the sample to be checked, then at first can be when the 1st step and 1 anti-formation 1 anti-compound, 1 anti-compound can be again with 2 of detecting probe surface resist compound, by externally-applied magnetic field by enrichment, separation.After separating, washing, magnetic adds the paramagnetic nano Ni probe suspension that a small amount of aseptic deionized water then forms object bacteria.At this moment, the excess probe of catching object bacteria and not catching object bacteria still mixes.
5) the probe suspension with enrichment is added on the solid phase carrier of having fixed 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 probe that does not then grab object bacteria is just washed off, if there is not object bacteria, then all probes are all washed off.
6) afterwards, washing with the probe of eluant, eluent with the double antibodies sandwich on the fixed head, fall ion and solvent with the method for externally-applied magnetic field separate probe with aseptic washed with de-ionized water, is exactly the probe of catching object bacteria if also there is probe.This part probe, the suspension that adds aseptic deionized water formation probe, 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 probe, thereby in the indirect proof sample target pathogenic bacteria is arranged, the amount of probe and the decline of T1/T2 are proportional, can quantitatively go out the amount of object bacteria by quantitative probe to a certain extent indirectly.
Described NMR probe is the nanoscale Ni material with paramagnetic characteristic, and 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.
Described 1 is anti-for detecting the specific antibody of object bacteria, preferably monoclonal antibody.2 anti-be the antibody of the 1st antibody.
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 detecting the indirect enrichment of paramagnetic nano Ni probe 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 monocytogenes.
1. nanometer Ni immunological probe preparation: the 1st antibody adopts the anti-IgG monoclonal antibody of rabbit of Listeria monocytogenes, and 2 anti-ly are the goat anti-rabbit igg of Listeria monocytogenes.Buy nanometer Ni material from the market, such as Deco island, Beijing gold Science and Technology Co., Ltd. or the super prestige nano material in Shanghai company, 20nm, purity 99.9%.
Silicon dioxide coated nanometer Ni: getting the 47.5g sodium silicate, be dissolved in the beaker with deionized water, is 12-13 with the salt acid for adjusting pH value.Get 5.0g nanometer Ni and join in this beaker, mechanical raking (using glass bar) 5min.With the ultrasonic 30min of mixed liquor, stir in good time.Be warmed up to 85 ℃, dropwise add salt acid for adjusting pH value 6-7, generate precipitation.Limit magnetic separates the limit and spends the deionised water precipitation, washs 3-4 time.Then, precipitation is scattered in the 250mL methyl alcohol.Above process triplicate guarantees that silicon is attached on the Ni.
Amino containing silane Ni nano material: the silicon dioxide coated nanometer Ni that will make joins in the 25mL methyl alcohol, uses 1mLH
2O and methyl alcohol are diluted to 150mL.Adding 150mL glycerine then mixes.Ultrasonic 30min transfers in the 500mL there-necked flask of stirring apparatus.Add 10mL amino silicane coupling agent (AEAPS), after stirring 3h fast under 80-90 ℃, migrate out product.Product spends deionised water 3 times, methanol wash 2 times (using the Buchner funnel suction filtration).Vacuum drying.It should be noted that suction filtration because glycerine is arranged, thus slow, to take out can inhale with suction pipe after a period of time in good time and remove supernatant liquid, the suction filtration process needs 6-8h approximately.At last with the amino containing silane Ni nano material vacuum drying 12h that obtains.
Antibody modification: get a certain amount of amination Ni nano particle, add the goat anti-rabbit igg antibody of excessive Listeria monocytogenes, 26 ℃ hatch, wash after, add excessive 1% bovine serum albumin (BSA), 22 ℃, the active room of confining surface, washing, resuspension.Because nanometer Ni has the paramagnetic characteristic, externally-applied magnetic field separates, and nanometer Ni will be pooled to magnetic field on one side, siphons away supernatant, and washing is then with unnecessary antibody, BSA flush away.The preparation the paramagnetic nano immunological probe be kept at 4 ℃ stand-by.
2. 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 Listeria monocytogenes first antibody, the anti-IgG monoclonal antibody of rabbit is about to 100
L 100
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 the rabbit of Listeria monocytogenes will be added in the sample to be checked.If have Listeria monocytogenes in the sample to be checked, will with 1 anti-formation 1 anti-compound.Fully shake after the goat anti-rabbit igg probe of the 1st the 2nd antibody Listeria monocytogenes that makes of step added.Last magnetic force frame separate probe adds the suspension that a small amount of aseptic deionized water obtains probe.At this moment, if the target Listeria monocytogenes 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 detecting probe surface, still mix.The probe suspension of this enrichment is added on the prepared 1 anti-fixed head of the 2nd step, and the probe that then combines Listeria monocytogenes in the probe 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 less than the probe flush away in conjunction with Listeria monocytogenes.At the remaining probe that just only combines Listeria monocytogenes of fixed head.
4. with eluent (methyl alcohol etc.) probe that combines Listeria monocytogenes on the fixed head is eluted.Last magnetic force frame, separate probe is also cleaned 1-2 time, with the ion 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 probe to exist in the solution, thereby in the explanation sample Listeria monocytogenes is arranged.The amount of probe and the drop-out value of T1/T2 are proportional.The bright probe of more speaking more that descends is more many, thereby the side light Listeria monocytogenes is more many, can quantitatively detect the number of object bacteria in the sample by the mark-on checking.All food standards all must not detect Listeria monocytogenes, and the method can fast detecting go out whether contain Listeria monocytogenes in the sample.
Embodiment
Example 2
Measure food samples and whether contain harmful pathogenic bacteria---Escherichia coli O 157: H7.
1. nanometer Ni immunological probe 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.The Ni nano particle can be bought (as the super prestige nano material company in Shanghai) from the market.
Silicon dioxide coated nanometer Ni: getting the 47.5g sodium silicate, be dissolved in the beaker with deionized water, is 12-13 with the salt acid for adjusting pH value.Get 5.0g nanometer Ni and join in this beaker, mechanical raking (using glass bar) 5min.With the ultrasonic 30min of mixed liquor, stir in good time.Be warmed up to 85 ℃, dropwise add salt acid for adjusting pH value 6-7, generate precipitation.Limit magnetic separates the limit and spends the deionised water precipitation, washs 3-4 time.Then, precipitation is scattered in the 250mL methyl alcohol.Above process triplicate guarantees that silicon is attached on the Ni.
Amino containing silane Ni nano material: the silicon dioxide coated nanometer Ni that will make joins in the 25mL methyl alcohol, uses 1mLH
2O and methyl alcohol are diluted to 150mL.Adding 150mL glycerine then mixes.Ultrasonic 30min transfers in the 500mL there-necked flask of stirring apparatus.Add 10mL amino silicane coupling agent (AEAPS), after stirring 3h fast under 80-90 ℃, migrate out product.Product spends deionised water 3 times, methanol wash 2 times (using the Buchner funnel suction filtration).Vacuum drying.It should be noted that suction filtration because glycerine is arranged, thus slow, to take out can inhale with suction pipe after a period of time in good time and remove supernatant liquid, the suction filtration process needs 6-8h approximately.At last with the amino containing silane Ni nano material vacuum drying 12h that obtains.
Antibody modification: get a certain amount of amination Ni nano particle, add excessive 2 anti-, i.e. the goat anti-rabbit igg antibody of Escherichia coli O 157: H7,26 ℃ hatch, wash after, add excessive 1% bovine serum albumin (BSA), 22 ℃, the active room of confining surface, washing, resuspension.Because nanometer Ni has the paramagnetic characteristic, externally-applied magnetic field separates, and nanometer Ni will be pooled to magnetic field on one side, siphons away supernatant, and washing is then with unnecessary antibody, BSA flush away.The preparation the paramagnetic nano immunological probe be kept at 4 ℃ stand-by.
2. 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 1st antibody, namely the anti-IgG monoclonal antibody of rabbit is about to 100
L 100
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, 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 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.With the 2nd antibody that the 1st step made, the goat anti-rabbit igg probe of O157:H7 fully shakes after adding.Last magnetic force frame separate probe adds the suspension that a small amount of aseptic deionized water obtains probe.At this moment, if the target Listeria monocytogenes 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 detecting probe surface, still mix.The probe suspension of this enrichment is added on the prepared 1 anti-fixed head of the 2nd step, and the probe that then combines O157:H7 in the probe 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 less than the probe flush away in conjunction with O157:H7.At the remaining probe that just only combines O157:H7 of fixed head.
With eluent (methyl alcohol etc.) with the Escherichia coli O 157 that combines on the fixed head: the probe of H7 elutes.Last magnetic force frame, separate probe 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 or miniNMR East China Normal University) measure T1 and the T2 of 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 probe to exist in the solution, thereby Escherichia coli O 157: H7 is arranged in the explanation sample, and the amount of probe and the drop-out value of T1/T2 are proportional.The bright probe 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 (6)
1. NMR food-borne pathogens method for quick based on the indirect enrichment of paramagnetic nano Ni probe, its characterization step is as follows:
1) add specificity the 1st antibody that detects object bacteria in sample, if having object bacteria in the sample, then the combination by antibody antigen forms 1 anti-compound;
2) antibody of the 1st antibody of detection object bacteria, namely 2 anti-bags are by the preparation of paramagnetic nano Ni probe;
3) with object bacteria specificity the 1st antibody fixing standby on solid phase carrier;
4) enrichment aimed strain, and separate: the sample of handling in the 1st step, add the paramagnetic nano Ni probe that the 2nd step made, fully mix concussion reaction a period of time, grasp the after-applied externally-applied magnetic field of object bacteria, because nanometer Ni has the paramagnetic characteristic, the Ni probe can gather magnetic field on one side, siphons away supernatant and then can isolate probe; If in the sample to be checked object bacteria is arranged, then at first can be at the 1st when step and 1 anti-formation 1 anti-compound, 1 anti-compound can resist compound with 2 of detecting probe surface again, by enrichment, separation, after separating, washing, magnetic adds the paramagnetic nano Ni probe suspension that a small amount of aseptic deionized water then forms object bacteria by externally-applied magnetic field; At this moment, catch and the Ni probe of not catching 1 anti-compound still mixes;
5) the Ni probe suspension of enrichment being added to the 2nd step has fixed on the solid phase carrier of monoclonal antibody, if exist object bacteria then to form double antibodies sandwich, with aseptic washed with de-ionized water, the probe that does not then grab object bacteria is just washed off, if there is not object bacteria, then all probes are all washed off;
6) afterwards, washing with the probe of eluant, eluent with the double antibodies sandwich on the fixed head, fall ion and solvent with the method for externally-applied magnetic field separate probe with aseptic washed with de-ionized water, is exactly the probe of catching object bacteria if also there is probe;
7) this part probe, the suspension that adds aseptic deionized water formation probe, 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 probe, thereby the target pathogenic bacteria are arranged in the indirect proof sample, the amount of probe and the decline of T1/T2 are proportional, can quantitatively go out the amount of object bacteria by adding the scalar quantity probe indirectly.
2. the NMR food-borne pathogens method for quick based on the indirect enrichment of paramagnetic nano Ni probe according to claim 1 is characterized in that described NMR nano-probe is the nanometer Ni material with paramagnetic characteristic.
3. the NMR food-borne pathogens method for quick based on the indirect enrichment of paramagnetic nano Ni probe according to claim 1, what it is characterized in that described object bacteria finally detects evaluation method based on the variation of the relaxation time characterisitic parameter of nuclear magnetic resonance technique.
4. the NMR food-borne pathogens method for quick based on the indirect enrichment of paramagnetic nano Ni probe according to claim 1 is characterized in that described NMR relaxation time characteristic, refers to spin-lattice relaxation time (T1) and spin spin relaxation time (T2).
5. the NMR food-borne pathogens method for quick based on the indirect enrichment of paramagnetic nano Ni probe according to claim 1 is characterized in that described relaxation time characteristic, and T1 adopts 180 °-τ-90 ° of pulse method to measure, and T2 adopts cpmg sequence row method to measure.
6. the NMR food-borne pathogens method for quick based on the indirect enrichment of paramagnetic nano Ni probe according to claim 1 is characterized in that described 1 is anti-for detecting the specific antibody of object bacteria, preferably monoclonal antibody; 2 anti-be the antibody of the 1st antibody.
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| CN101726535A (en) * | 2008-10-24 | 2010-06-09 | 北京朔望科技有限公司 | Time-resolved remanence relaxation detector and application thereof in detection of superparamagnetic nanomaterial |
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| CN101726535A (en) * | 2008-10-24 | 2010-06-09 | 北京朔望科技有限公司 | Time-resolved remanence relaxation detector and application thereof in detection of superparamagnetic nanomaterial |
| WO2011150212A2 (en) * | 2010-05-26 | 2011-12-01 | The General Hospital Corporation | Magnetic nanoparticles |
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