CN102784628A - Solid-phase micro extraction fiber extraction head and preparation method thereof - Google Patents
Solid-phase micro extraction fiber extraction head and preparation method thereof Download PDFInfo
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Abstract
The invention discloses a preparation method for a solid-phase micro extraction fiber extraction head. The preparation method comprises the following steps of: firstly, carrying out silane treatment on a bare quartz fiber; secondly, adding an acrylamide function monomer, a cross-linking agent and an initiator in a polymerization solution and fully mixing to obtain a prepolymerization solution; thirdly, coating a porous polymer coating on the surface of the quartz fiber subjected to silane treatment; fourthly, carrying out ageing treatment on the quartz fiber; fifthly, repeating the second step to the fourth step for multiple times and repeatedly coating the porous polymer coating on the surface of the quartz fiber for multiple times; sixthly, carrying out bonding reaction on the porous polymer coating and the quartz fiber; and seventhly, adding the quartz fiber treated in the sixth step into an aptamer solution and carrying out bonding reaction at room temperature. According to the preparation method disclosed by the invention, the quartz fiber is treated by the steps, and thus the bonded amount of the aptamer on the surface of the fiber can be remarkably improved; and an SPME (Solid-Phase Micro Extraction) head with a novel aptamer/ porous polymer coating can be prepared.
Description
Technical field
The invention belongs to the chemical analysis test instrument field, relate to the solid-phase micro-extraction coating fiber preparation method, this fiber is applicable to the separation and the enrichment of trace biology alkali in the complex biological sample, antibiotic or ucleotides material.
Background technology
Efficiently, the sample pre-treatments technology of high selectivity is the important step of Analysis of Complex matrix like trace, ultra trace material in biological, medicine, environmental sample and the food.In the chromatography process, sample pretreatment normally the most consuming time be difficult for the step of automation, also be the most key step of generally acknowledging at present.But with respect to the development of instrument analysis technology, the sample pre-treatments Progress in technique is slower always.Shortcomings such as traditional sample pre-treatments technology is like liquid-liquid extraction, Suo Shi extraction, column chromatography etc., and ubiquity length consuming time, efficient are low, complex operation, consumption of organic solvent are big.The existence of these problems makes sample pre-treatments work become the most time-consuming in the whole assay determination process, effort, also introduces a link of error the most easily.
SPME (Solid-Phase Microextraction; SPME) technology is proposed in 1989 by Belardi and Pawliszyn; In flowing phase and be fixed between the macromolecular solid phasing of fused silica fiber surface two principles of distributing mutually, realize the extraction and the enrichment of analyte in the sample, directly desorb in the coupling instrument, sample introduction and analysis then based on analyte; The sample pretreatment process is greatly simplified, improved analysis speed and sensitivity.The core of SPME technology is its extracting head coating surfaces, but commercialization coating kind is limited, and a few is only arranged, and it is relatively poor to be coated with layer-selective, and is not suitable for handling polarity or alkaline matter.Though some bibliographical informations the development work of all kinds of New type of S PME coatings, like polysiloxanes-fullerene (Xiao C.H., Han S.Q.; Wang Z.Xing Y., J., and Wu C.Y.2001.J.Chromatogr.A 927:121-130), crown ether (Zeng; Z.R., W.L.Qiu, and Z.F.Huang.2001.Anal.Chem.73:2429-2436), calixarenes (Li; X.J., Z.R.Zeng, S.Z.Gao; And H.B.Li.2004.J.Chromatogr.A 1023:15-25) etc., selectivity improves with respect to the commercialization coating, but the analytic target scope is narrower; Be mainly used in and measure volatilization or half volatile organic environment pollutant, and selectivity is still strong inadequately.Since nineteen ninety, molecular imprinting in the analytical chemistry field, has especially obtained extensive use in research fields such as SPE, chromatograph packing material, chemical sensors because of its " key-lock " interaction recognition principle that is similar to enzyme-substrate.Molecularly imprinted polymer (the Molecularly imprinted polymer of this technology preparation; MIP) have characteristics such as selectivity height, chemical stability is good, preparation is simple; Be particularly suitable for selective coating material (Hu X.G. as SPME; Hu Y.L., and Li G.K.2007.J.Chromatogr.A 1147:1-9.).But because the limitation of molecular imprinting itself, MIP remains synthetic need of following weak point: MIP as the SPME coating material and adds a large amount of template molecules, is difficult to complete wash-out, possibly occur template molecule seepage phenomenon during extraction; The MIP selectivity receives serious interference mainly based on hydrogen bond action during analysis of biological samples (aqueous solution); Benzene, toluene, chloroform or other organic solvent are used in the extraction of MIP coating often, are prone to contaminated environment; The rigidity of MIP coating identification " hole " is destroyed easily in the preparation process or during extraction or is out of shape, and its specificity and affinity are not as bio-identification system such as interactions such as enzyme-substrate, antibody-acceptor.Therefore, development selectivity SPME coating material stronger, that be more suitable for trace materials analysis in complex biological sample is very necessary, and the more suitable selection beyond doubt of bio-identification system.
Aptamer (Aptamer) is the single stranded oligonucleotide sequence of one section weak point obtaining through in-vitro screening of the phyletic evolution technology (SELEX) through the index concentration aglucon; It is folding that it through pairing between some complementary base in the chain and electrostatic interaction, hydrogen bond action etc. self adaptability takes place; Form some stable three-D space structures; Like hair clip, false knot, bulge loop, G2 tetrad etc., complementary with the ligand molecular high-affinity, combine with high specificity through steric configuration.For example, theophylline is commonly used in asthma, bronchitis and emophysematous treatment makes bronchiectasis medicine, is prone to cause poisoning, is difficult to distinguish with caffeine (only differing a methyl on both structures) in the serum.Jenison etc. separate from the RNA storehouse and obtain the theophylline aptamers, with the affinity of theophylline than caffeine high more than 10000 times (Pardi A., and Polisky is 263:1425-1429. B.1994.Science for Jenison R.D., Gill S.C.); The atriphos that screenings such as Sazani obtain (ATP) RNA aptamers is higher 64 times than adenosine diphosphate (ADP) (ADP) with the affinity of ATP; Than high 1100 times of (the Sazani P.L. of single AMP (AMP); Larralde R., and Szostak J.W.2004.J.Am.Chem.Soc.126:8370-8371.).Aptamer except that have the same with antibody can efficient with respective ligand, the single-minded characteristics that combine, also have the following advantages: be prone to chemistry and modify; In-vitro screening, chemical synthesis is obtained easily, and cost is low; Good stability, recoverability; The target molecule scope is wide.Aptamers obtains from random oligonucleotide storehouse screening through the SELEX technology, and single-chain nucleic acid forms the diversity of interaction force between diversity and the aptamers and the part of three-dimensional structure in addition, and most materials of occurring in nature can both screen corresponding aptamers (Burgstaller P. in theory; Girod A., and Blind is Discovery Today 7:1221-1228. M.2002.Drug), the part scope of having reported at present is quite extensive; Little molecule (Ferapontova E.E., Olsen E.M., and Gothelf K.V.2008.J.Am.Chem.Soc.130:4256-4258.), carbohydrate (Jeong S. are arranged; Eom T., Kim S., Lee S.; And Yu is J.2001.Biochem.Biophys.Res.Commun.281:237-243.), amino acid (Famulok M.1994.J.Am.Chem.Soc.116:1698-1706.), nucleotides (Zhang S.S.; Xia J.P., and Li X.M.2008.Anal.Chem.80:8382-8388.), peptide (Davis J.J., Tkac J.; Laurenson S.; And Ferrigno P.K.2007.Anal.Chem.79:1089-1096.), protein (and Lu is Chem.19:412-417. Y.2008.Bioconjugate for Yigit M.V., Mazumdar D.) even whole cell (Hamula C.L.A.; Zhang H.Q.; Guan L.L., Li X.F., and Le X.C.2008.Anal.Chem.80:7812-7819.).
The application of aptamer in the analytical chemistry field become the focus that domestic and international researcher pays close attention to.Aptamers can be fixed on various material surfaces as aglucon after chemical modification; Like metal, silica gel, glass, magnetic microsphere, quantum dot etc.; Be applied to all kinds of isolation technics, comprise liquid chromatogram, affinity chromatography, Capillary Electrophoresis, capillary electric chromatogram, biology sensor and AFM etc.For example; People such as Brumb are modified at the arginic RNA aptamers of L-on the affinity chromatograph filling, realize L-arginine and the arginic high efficiency separation of D-(Brumbt A., Ravelet C.; Grosset C.; Ravel A., Villet A., and Peyrin are E.2005.Anal.Chem.77:1993-1998.); Chris Le etc. is fixed on aptamers in the capillary chromatography integral post, is used for the separation and the detection (Zhao Q., Li X.F., and Chris Le are X.2008.Anal.Chem.80:3915-3920.) of protein mixture cromoci.The application of aptamers in the sample pre-treatments field also has very big potentiality; Tan etc. are fixed on the magnetic nanoparticle surface with aptamers; The selective separation enrichment (Herr J.K., Smith J.E., the Medley C.D. that are used for the complex biological sample cancer cell; Shangguan D.H., and Tan W.H.2006.Anal.Chem.78:2918-2924.); Oktem etc. are fixed on magnetic bead surfaces with aptamers; The purifying that is used for bacterium crude extract reorganization Taq archaeal dna polymerase; Once extract the operation extraction yield and reach 93% (Oktem H.A.; Bayramoglu G., Ozalp V.C., and Arica M.Y.2007.Biotechnol.Prog.23:146-154.).
Summary of the invention
The present invention is directed to that the described SPME coating of preamble kind is less, selectivity is not high and the compatible problem such as relatively poor of biological sample; Characteristics such as aptamer affinity height, high specificity, biological sample compatibility are good and SPME technology is consuming time less, efficient is high, simple operation and other advantages combines; Porous polymer coating at silanization quartz fibre surface preparation one deck high-permeability; Through the chemical bonding method aptamer is fixed in the coating loose structure; Develop a kind of New type of S PME fibre abstraction head based on aptamer/porous polymer coating, the high selectivity that is used for complex biological sample trace, ultra trace alkaloid, antibiotic or ucleotides material is separated and enrichment.
The present invention realizes through following technical scheme:
A kind of preparation method of solid-phase micro-extraction fibre extracting head, carry out according to the following steps:
(1) naked quartz fibre is carried out the alkali cleaning of hydrofluoric acid burn into, pickling and activation processing, adopt silylating reagent to carry out silanization then and handle with unsaturated double-bond;
(2) in polymer solvent, add acrylamide function monomer, crosslinking agent and initator, fully mix, obtain pre-polymer solution;
(3) get a certain amount of pre-polymer solution in test tube, put into the prepared silanization quartz fibre of step 1, logical nitrogen deoxygenation, sealing test tube mouth, heat causes copolyreaction in the water-bath, makes the surface-coated polymer coating of quartz fibre;
(4) behind the polymerization certain hour, extract quartz fibre, put into another clean tube, logical nitrogen deoxygenation, sealing test tube mouth places drying box aging under uniform temperature;
(5) (2) to (4) step several times more than repeating on the same quartz fibre repeat to be coated with stain several times porous polymer coating on the quartz fibre surface;
(6) the porous polymer coating quartz fibre that obtains at last in (5) step is placed bonding liquid, react certain hour under the room temperature, take out quartz fibre, clean with distilled water, nitrogen dries up;
(7) will place aptamer solution through the quartz fibre after (6) step handled, bonding reaction certain hour under the room temperature takes out quartz fibre, uses buffer solution for cleaning, and nitrogen dries up, and low temperature is preserved.
Specifically, said silylating reagent is a trimethylol-propane trimethacrylate.
Specifically, said polymer solvent is an acetone, and crosslinking agent is a trimethylol-propane trimethacrylate; Said initator is an azodiisobutyronitrile.
Preferably, acetone and trimethylol-propane trimethacrylate volume ratio are 9:1, and acrylamide and trimethylol-propane trimethacrylate mol ratio are 1:4.
The porous polymer coating repeats to be coated with stain and reaches 2 ~ 5 layers on the quartz fibre surface.
Specifically, the bonding agent that is adopted is a carbonyl dimidazoles, utilizes carbonyl dimidazoles to pass through the amino of modifying on amino and the aptamer end group of chemical bonding link porous polymer coating surface.
Further, the bonding conditions that is adopted: carbonyl dimidazoles concentration is 0.7mol/L, and the bonding time is 1.5 hours.
The used buffer solution of aptamer bonding is the TE buffer solution, and the aptamer solution concentration is 6.25 μ g/mL, and the aptamers bonding time is 24 hours.
The quartz fibre surface is evenly level and smooth, and surface area is little, is unfavorable for the bonding of aptamer.After the present invention carries out chemical modification and silanization processing to the quartz fibre surface; Through the radical copolymerization method and repeat to be coated with the stain mode and have certain thickness porous polymer coating in the quartz fibre surface preparation; Utilize the three-dimensional porous structure and the high-specific surface area of porous polymer coating, significantly improve the bonded amount of aptamer at fiber surface.
Aptamer is immobilized to have polymer embedding, physical absorption, directly self assembly, methods such as self assembly indirectly, but lower, immobilized insecure, the aptamers of the immobilized rate of ubiquity is prone to problems such as loss.The present invention adopts carbonyl dimidazoles as the link agent; Link the amino group and the end modified amino group of aptamer of porous polymer coating surface function monomer acrylamide through the chemical bonding mode; The aptamer covalent bonding is immobilized in the porous polymer coating surface, prepare the SPME extracting head of novel nucleic acids aptamers/porous polymer coating.
Description of drawings
Fig. 1. aptamer/porous polymer coating SPME extracting head prepares the process sketch map.(A. quartz fibre chemical surface treatment and silanization; B. porous polymer coated fiber preparation; C. porous polymer coating surface bonding aptamer)
Fig. 2. aptamer/porous polymer coating SPME extracting head electron micrograph.(Fig. 2 A is 1000 multiplication factors; Fig. 2 B is 18000 multiplication factors)
Fig. 3 .ATP aptamer/porous polymer coating SPME extracting head (Apt-Pol-fiber); Double alkali yl mispairing ATP aptamer/porous polymer coating SPME extracting head (TMApt-Pol-fiber); Out of order ATP aptamer/porous polymer coating SPME extracting head (SApt-Pol-fiber) extracts the 0.1mg/L adenosine triphosphate atp respectively; Adenosine diphosphate (ADP) ADP; Single AMP AMP; Adenosine A denosine; GTP (GTP); Uridine triphosphate (UTP); Cytidine (CTP); Isopropyl methoxalamine Metolachlor; The single mark of theophylline Theophylline standard liquid extraction quantity comparison diagram.
Fig. 4 .ATP aptamer/porous polymer coating SPME extracting head extraction variable concentrations ATP, ADP, AMP mix mark standard liquid extraction quantity curve.
The specific embodiment
This enforcement is prepared as example, describes the present invention, but do not limit protection scope of the present invention with this with ATP aptamer/porous polymer coating SPME extracting head.
As shown in Figure 1, the preparation process of ATP aptamer/porous polymer coating New type of S PME extracting head is following:
(1) cut-off directly is that 125 μ m, length are the naked quartz fibre of 6.0cm, with 10% hydrofluoric acid solution corrosion 20s, takes out the back with distilled water flushing three times; Fiber is placed 1.0mol/L NaOH alkali wash water soaking at room temperature 0.5h, take out the back with distilled water flushing three times; Place 1.0mol/L HCl pickle to soak 0.5h quartz fibre after the alkali cleaning, take out the back with distilled water flushing three times; Quartz fibre is put into culture dish place 150 ℃ of dehydrations of baking oven activation 1h.From baking oven, take out quartz fibre, put into the acetone soln of 10% trimethylol-propane trimethacrylate immediately, Silanization reaction 0.5h takes out the back with absolute ethyl alcohol flushing three times, and nitrogen dries up.
(2) in the ground conical flask, add 27.0mL acetone polymer solvent, 172mg acrylamide function monomer, 3.00mL trimethylol-propane trimethacrylate crosslinking agent and 45.0mg azodiisobutyronitrile initator successively; Fully shake up, make pre-polymer solution.Get a clean small test tube, add the 2.0mL pre-polymer solution, ultrasonic degas 5min.In test tube, put into a silanization quartz fibre, logical nitrogen deoxygenation 1min seals the test tube mouth with plug immediately; Test tube is placed thermostat water bath, and 60 ℃ of heated polymerizable 2h open plug; At the uniform velocity extract quartz fibre, be transferred in another clean tube, logical nitrogen deoxygenation; Seal with plug, place 65 ℃ of following heat ageing 12h of thermostatic drying chamber.
(3) from drying box, take out the above-mentioned test tube that is coated with a polymer quartz fibre of stain that is equipped with, place air to cool off, add above-mentioned fresh of 2.0mL and the pre-polymer solution after the 5min ultrasonic degas is handled; Logical nitrogen deoxygenation 1min seals the test tube mouth with plug immediately, and test tube is placed thermostat water bath; 60 ℃ of heated polymerizable 2h open plug, at the uniform velocity extract quartz fibre; Be transferred in another clean empty test tube; Logical nitrogen deoxygenation is sealed with plug, places 65 ℃ of following heat ageing 12h of thermostatic drying chamber.Repeat above-mentioned steps, steep 5 times porous polymer coating quartz fibre, place 150 ℃ of following heat ageings of thermostatic drying chamber to handle 2h at last until making to repeat to be coated with.
(4) get and be coated with 5 times porous polymer coating quartz fibre of stain, will scrape off from the polymer coating more than the fiber head 1cm with pocket knife, remaining porous polymer coated length is unified to be 1.0cm.Get a dried and clean test tube, in test tube, add 1.5mL 0.7mol/L carbonyl dimidazoles solution (dry acetonitrile dissolving), put into the above-mentioned porous polymer coating quartz fibre of handling well, take out behind the bonding 1.5h, with dry acetonitrile flushing 3 times, nitrogen dries up.
(5) (10mmol/L tris-HCl, 1mmol/L EDTA pH=7.5), put into the porous polymer coating quartz fibre after the carbonyl dimidazoles bonding is handled, and hatch 30min under the room temperature in the dried and clean sample bottle, to add the 1.5mLTE buffer solution.Fiber take out the back insert ATP aptamer standard liquid that 100 μ L6.25 μ g/mL are housed (dissolving of TE buffer solution, 10mmol/l tris-HCl, 1mmol/LEDTA, in glass-lined pipe pH=7.5), bonding reaction 24h under the room temperature.Take out fiber,, obtain ATP aptamer/porous polymer coating SPME extracting head with TE buffer solution flushing 3 times.
ATP aptamer/porous polymer coating SPME the extracting head of this case study on implementation preparation has the following advantages:
1. adopt the chemical bonding method; Free radicals copolymerization reaction through optimal conditions is in the surface-coated porous polymer coating of quartz fibre, and preparation method's good reproducibility prepares 20 porous polymer coated fibers simultaneously; The coating average thickness is 10.5 μ m, and precision is 4.9%.
2. the porous polymer coating for preparing is even, fine and close, and the surface is loose and porous structure (as shown in Figure 2), can significantly improve aptamers supported quantity in the adaptive bonding step of follow-up nucleic acid.This polymer coating has good chemistry and mechanical stability, reuse more than 200 time with direct extraction mode after, coating any cracking do not occur, peels off or wear phenomenon.
3. as shown in Figure 3; ATP aptamer/porous polymer coating SPME the extracting head of the present invention's preparation has very high selective extraction capacity (the ATP extraction quantity is 6 ~ 62 times of other analytic target) to ATP; ATP analogue such as ADP, AMP, GTP had selectivity preferably; Non-structural similarity thing such as theophylline, isopropyl methoxalamine etc. are not had selective extraction capacity, can be used for separation and the enrichment of trace ATP, ADP, AMP in the complex biological sample.Change two crucial recognition site bases (21 and No. 34 bases) in the ATP aptamer base sequence; The double alkali yl mispairing ATP aptamer that makes/porous polymer coating SPME extracting head significantly descends to the extraction quantity of ATP, ADP, AMP; And extracting head does not have selectivity to ATP, ADP, AMP after adopting out of order ATP aptamer, and this shows that the selectivity recognition capability of the New type of S PME extracting head that the present invention is prepared stems from the ATP aptamer of extracting head surface bond.
4. the prepared ATP aptamer/porous polymer coating SPME extracting head of the present invention has higher separation and concentration ability to ATP, ADP, AMP, and loading capacity is about 7.4,0.7,0.3ng, and is as shown in Figure 4.
Claims (9)
1. the preparation method of a solid-phase micro-extraction fibre extracting head, carry out according to the following steps:
(1) naked quartz fibre is carried out the alkali cleaning of hydrofluoric acid burn into, pickling and activation processing, adopt silylating reagent to carry out silanization then and handle with unsaturated double-bond;
(2) in polymer solvent, add acrylamide function monomer, crosslinking agent and initator, fully mix, obtain pre-polymer solution;
(3) get a certain amount of pre-polymer solution in test tube, put into the quartz fibre of gained behind step 1 silanization, logical nitrogen deoxygenation, sealing test tube mouth, heat causes copolyreaction in the water-bath, makes the surface-coated porous polymer coating of quartz fibre;
(4) behind the polymerization certain hour, extract quartz fibre, put into another clean tube, logical nitrogen deoxygenation, sealing test tube mouth places drying box aging under uniform temperature;
(5) (2) to (4) step several times more than repeating on the same quartz fibre repeat to be coated with stain several times porous polymer coating on the quartz fibre surface;
(6) the porous polymer coating quartz fibre that obtains at last in (5) step is placed bonding agent solution, bonding reaction certain hour under the room temperature takes out quartz fibre, cleans with distilled water, and nitrogen dries up;
(7) will place aptamer solution through the quartz fibre after (6) step handled, bonding reaction certain hour under the room temperature takes out quartz fibre, uses buffer solution for cleaning, and nitrogen dries up, and low temperature is preserved.
2. the preparation method of solid-phase micro-extraction fibre extracting head as claimed in claim 1 is characterized in that: said silylating reagent with unsaturated double-bond is a trimethylol-propane trimethacrylate.
3. the preparation method of solid-phase micro-extraction fibre extracting head as claimed in claim 1 is characterized in that: said polymer solvent is an acetone, and crosslinking agent is a trimethylol-propane trimethacrylate.
4. the preparation method of solid-phase micro-extraction fibre extracting head as claimed in claim 3 is characterized in that: acetone and trimethylol-propane trimethacrylate volume ratio are 9:1, and acrylamide and trimethylol-propane trimethacrylate mol ratio are 1:4.
5. the preparation method of solid-phase micro-extraction fibre extracting head as claimed in claim 3 is characterized in that: the porous polymer coating repeats to be coated with stain and reaches 2 ~ 5 layers on the quartz fibre surface.
6. the preparation method of solid-phase micro-extraction fibre extracting head as claimed in claim 5 is characterized in that: the bonding agent that is adopted is a carbonyl dimidazoles.
7. the preparation method of solid-phase micro-extraction fibre extracting head as claimed in claim 6 is characterized in that: said carbonyl dimidazoles concentration is 0.7mol/L, and the bonding time is 1.5 hours.
8. the preparation method of solid-phase micro-extraction fibre extracting head as claimed in claim 6 is characterized in that: the buffer solution of aptamer solution is the TE buffer solution, and the aptamers bonding time is 24 hours.
9. by the prepared solid-phase micro-extraction fibre extracting head of the arbitrary method of claim 1 ~ 8.
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| CN102989432A (en) * | 2012-12-28 | 2013-03-27 | 南开大学 | Preparation of solid-phase microextraction (SPME) fiber and extraction device assembled by same |
| CN103055830A (en) * | 2012-12-26 | 2013-04-24 | 华南师范大学 | Preparation method for solid-phase micro-extraction head based on single-stranded DNA aptamer modified graphene oxide coating |
| CN104258833A (en) * | 2014-09-24 | 2015-01-07 | 华南师范大学 | Preparation method of novel solid phase microextraction fiber based on nucleic acid aptamer/ nanogold/ porous polymer coating |
| CN106267898A (en) * | 2016-10-16 | 2017-01-04 | 刘晶 | Solid-phase micro extraction fiber extraction head and preparation method thereof |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103055830A (en) * | 2012-12-26 | 2013-04-24 | 华南师范大学 | Preparation method for solid-phase micro-extraction head based on single-stranded DNA aptamer modified graphene oxide coating |
| CN102989432A (en) * | 2012-12-28 | 2013-03-27 | 南开大学 | Preparation of solid-phase microextraction (SPME) fiber and extraction device assembled by same |
| CN104258833A (en) * | 2014-09-24 | 2015-01-07 | 华南师范大学 | Preparation method of novel solid phase microextraction fiber based on nucleic acid aptamer/ nanogold/ porous polymer coating |
| CN104258833B (en) * | 2014-09-24 | 2016-11-09 | 华南师范大学 | Preparation method based on aptamer/nanometer gold/porous polymer coating novel solid phase micro extraction fiber |
| CN106267898A (en) * | 2016-10-16 | 2017-01-04 | 刘晶 | Solid-phase micro extraction fiber extraction head and preparation method thereof |
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