CN101685051A - Open-tubular capillary column enriching phosphoeptide or phosphorylated protein and method - Google Patents
Open-tubular capillary column enriching phosphoeptide or phosphorylated protein and method Download PDFInfo
- Publication number
- CN101685051A CN101685051A CN200810211727A CN200810211727A CN101685051A CN 101685051 A CN101685051 A CN 101685051A CN 200810211727 A CN200810211727 A CN 200810211727A CN 200810211727 A CN200810211727 A CN 200810211727A CN 101685051 A CN101685051 A CN 101685051A
- Authority
- CN
- China
- Prior art keywords
- phosphoeptide
- phosphorylating protein
- capillary column
- enrichment
- protein
- 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
Landscapes
- Other Investigation Or Analysis Of Materials By Electrical Means (AREA)
Abstract
The invention relates to a method for enriching phosphoeptide or phosphorylated protein in an off-line way and an on-line way for a functionally modified open-tubular capillary column, which comprisesthe following steps: preparing an open-tubular capillary column, an inner wall of which can selectively adsorb the zirconium phosphate functional group of phosphoeptide or phosphorylated protein, enriching the phosphoeptide or phosphorylated protein after loading samples, and then, using a cleaning buffer solution to remove non-phosphoeptide or non-phosphorylated protein and salt; and after the cleaning process is finished, using eluent to elute the phosphoeptide or phosphorylated protein, wherein the eluted phosphoeptide or phosphorylated protein can be marked on the MALDI target to carry out normal MALDI-TOF-MS analysis, or the eluted phosphoeptide or phosphorylated protein can be inoculated on the liquid phase part of microliter or nanoliter LC-ESI-MS to be combined with ESI-MS/MS to automatically enrich and analyze the phosphoeptide or phosphorylated protein in the on-line way.
Description
Technical field
The invention belongs to the analytical chemistry field, be before utilizing mass-spectrometric technique Analysis and Identification phosphoeptide or phosphorylating protein, at first it is carried out a kind of method of selective enrichment, the promptly a kind of open pipe capillary column realization off-line of functional modification and method of on-line selection enrichment acid peptide or phosphorylating protein utilized.
Background technology
The phosphorylation modification of protein is one of covalent modification important in the biosome, and life processes such as the propagation of pair cell, differentiation, signal transduction, Apoptosis have important regulation.In recent years, mass spectrum (MS) had been widely used in the evaluation of phosphorylating protein.Because the stoichiometric number of phosphorylating protein is lower in the cell/tissue, and there is signal suppressing in non-phosphorylating peptide section to phosphoeptide during mass spectrophotometry, makes direct mass spectrophotometry phosphorylating protein or phosphoeptide have very big difficulty.Therefore developing the separation and concentration technology of high selectivity, is very necessary for the evaluation of phosphated peptide section, can promote the Mass Spectrometer Method of phosphoeptide.
At present, the method for the most frequently used enrichment acid peptide comprises solid metallic ion affinity chromatography (IMAC).In IMAC, utilize phosphate and the IMAC filling surface chelated metal ion such as the Fe of phosphoeptide
3+, Ga
3+Deng between specific suction-operated, make phosphoeptide be retained on the immobilised metal-chelating ion.Some metal oxides such as TiO
2, ZrO
2, Al
2O
3Deng the enrichment that also is used for phosphoeptide.In nearest research, people are with the surface of basic zirconium phosphate (ZrP) base group modification at polysilicon, the concentration and separation that is used for phosphorylated protein, and binding matrix auxiliary laser desorption ionization mass-spectrometric technique (Matrix-assisted laserdesorption/ionization mass spectrometry, MALDI-MS), obtained good concentration effect [Houjiang Zhou, Songyun Xu, Mingliang Ye, Shun Feng, Chensong Pan, Xiaogang Jiang, Xin Li, Guanghui Han, YuFu, and Hanfa Zou.Zirconium phosphonate-modified porous silicon for highly specific capture ofphosphopeptides and MALDI-TOF MS analysis (utilizing the polysilicon high-selectivity enrichment phosphoeptide of basic zirconium phosphate base group modification and ground substance assistant laser to resolve the ionization massspectrum analysis) .J.Proteome Res, 2006,5:2431-2437].Equally, modify the ZrP group in poly (glycidyl methacrylate) [poly (glycidyl methacrylate-co-ethylene dimethacrylate), GMA-EDMA] bead surface, the method that is used for the phosphorylated protein concentration and separation is also appeared in the newspapers.Because the strong coordination between the phosphate group of ZrP group and phosphoeptide makes concentration effect significantly improve.But these enrichment methods or operation steps are very loaded down with trivial details, handle a plurality of sample difficulties simultaneously.Because manual operations in centrifuge tube, transfer process causes sample loss, once can only handle several samples at most, can't satisfy that protein science is extensive, the requirement of high throughput analysis.These drawbacks limit its further application in extensive phosphorylating protein group analysis.
Summary of the invention
The open pipe capillary column that the purpose of this invention is to provide a kind of enrichment acid peptide or phosphorylating protein, and the method for selective enrichment phosphoeptide or phosphorylating protein.
The principle of this scheme is as follows:
The open pipe capillary column for preparing a kind of functionalization, its inwall have the functional group of alternative absorption phosphoeptide or phosphorylating protein.Can directly carry out enrichment behind the application of sample to phosphoeptide or phosphorylating protein, utilize cleaning buffer solution to remove non-phosphopeptide or non-phosphorylating protein and salt again, utilize eluent that phosphoeptide or phosphorylating protein are carried out wash-out after cleaning is finished, the phosphoeptide of wash-out or phosphorylating protein can be online or off-line carry out mass spectrophotometry.
The capillary column that invention is used is a kind of open pipe capillary column of functionalization, and its inwall has the functional group of alternative absorption phosphoeptide or phosphorylating protein.Specifically, be to utilize the silicon hydroxyl of kapillary open tubular column inside surface and phosphoeptide or phosphorylating protein sorbing material chemistry ways of connecting to make, this sorbing material is basic zirconium phosphate (ZrP) preferably.
For realizing the purpose of invention, the technical solution used in the present invention may further comprise the steps:
(1) the functional open pipe capillary column of preparation;
(2) sample is injected in the open pipe capillary column, hatches with selective adsorption phosphoeptide or phosphorylating protein;
(3) eluent is injected the open pipe capillary column, phosphoeptide or phosphorylating protein are carried out wash-out;
(4) phosphoeptide behind the wash-out or phosphorylating protein are carried out mass spectrophotometry.
When hatching absorption phosphoeptide or phosphorylating protein, can adopt continuous injection form or sample introduction after static state hatch.
For improving the accuracy of analyzing, when enrichment saliferous biological sample, between step (2) and (3), also should carry out the desalination operation.Specifically, be that cleaning fluid is injected the open pipe capillary column, remove non-phosphopeptide or non-phosphorylating protein and salt.
Can adopt off-line or online mode when carrying out mass spectrophotometry.Can add matrix in eluent when off-line enrichment acid peptide or phosphorylating protein carries out a target and analyzes on MALDI-TOF-MS; For ease of operation, can adopt the micro-sampling pump when off-line enrichment acid peptide or phosphorylating protein, and can guarantee the stability of sample solution, cleaning solution and elute soln flow velocity.When phosphoeptide or phosphorylating protein are analyzed in the automatic on-line enrichment, the open pipe capillary column can be connected on microlitre or receive the upgrading kapillary high pressure liquid chromatography-mass spectrum (LC-ESI-MS) liquid phase part on, directly carry out ESI-MS/MS and analyze, improve the utilization factor of automation of operation level and sample.
Technical scheme provided by the invention has the following advantages:
1, easy and simple to handle efficient, applied widely.The present invention is based on the strong coordination between the phosphate group of basic zirconium phosphate functional group and phosphoeptide or phosphorylating protein to the enrichment of phosphoeptide or phosphorylating protein, thereby has very high bioaccumulation efficiency.In the operation, both can utilize micro-sampling pump off-line enrichment acid peptide or phosphorylating protein, the whole operation process is very simple, convenient; Can be connected on microlitre again or receive on the liquid phase part of upgrading ESI-MS, realize that the automatic on-line enrichment analyzes phosphoeptide or phosphorylating protein, promote its analysis efficiency greatly.Aspect the coupling of mass analyzer, both can analyze in conjunction with MALDI-MS, can analyze in conjunction with ESI-MS/MS again, satisfy different sample analysis demands.
2, cost is low, is easy to reuse.The used open pipe capillary column preparation of the present invention is simple, cheap and easy to get, and can make the enriching column of dissimilar specifications, satisfies different analyze demands.To used open pipe capillary column, can carry out the function reparation by the mode that injection contains free zirconium ion solution, thereby prolong its serviceable life.
3, be specially adapted to the processing of large sample.The used open pipe capillary column of the present invention can a plurality of samples of batch processing.No matter adopt the mode of off-line enrichment, still adopt the mode of on-line preconcentration, different samples can load on respectively on the open pipe capillary column successively, hatch finish after by cleaning successively and wash-out, and then the phosphoeptide that elutes is carried out mass spectrophotometry.Especially when adopting the on-line preconcentration mode, can easily realize upward sample, cleaning, wash-out and the mass spectrophotometry automatically of sample in enormous quantities by the Control Software of robotization, its analysis efficiency is higher than phosphoeptide or phosphorylating protein enrichment method commonly used at present far away.
4, be specially adapted to the complex sample Treatment Analysis.When the present invention utilized phosphoeptide in the open pipe capillary column enrichment complex sample or phosphorylating protein, complex sample pre-treatment some components commonly used can not kept by pillar, can not disturb the enrichment of phosphoeptide or phosphorylating protein yet.Whole process does not need sample is carried out the desalination pre-treatment, has greatly reduced sample loss, be fit to handle the sample of trace, as peptide hydrolysis in the glue of two-dimensional gel electrophoresis etc.
Description of drawings
Fig. 1 is the process flow diagram of invention technical scheme.(a) be the open pipe capillary column of unmodified; (b) for modifying the open pipe capillary column of aminopropyl; (c) for modifying the open pipe capillary column of phosphate group; (d) for modifying the open pipe capillary column of basic zirconium phosphate group; (e) be the signal of the open pipe capillary column enrichment acid peptide of basic zirconium phosphate base group modification; (f) be the signal after by wash-out of the phosphoeptide of enrichment.1 is last sample operation, and 2 is cleaning operation, and 3 are the wash-out operation.
Fig. 2 is the MALDI-TOF-MS figure of alpha-casein matter enzymolysis product.(a) the direct MS of not enrichment of enzymolysis product analyzes; (b) MS analyzed after enzymolysis product utilized the open pipe capillary column enrichment of basic zirconium phosphate base group modification.
Fig. 3 is the MALDI-TOF-MS figure of β casein protein enzymolysis product.(a) the direct MS of not enrichment of enzymolysis product analyzes; (b) MS analyzed after enzymolysis product utilized the open pipe capillary column enrichment of basic zirconium phosphate base group modification.
Fig. 4 is the MALDI-TOF-MS figure after alpha-casein matter enzymolysis product utilizes the open pipe capillary column enrichment of basic zirconium phosphate base group modification.(a) do not contain urea, SDS and sodium chloride in the enzymolysis product solution; (b) contain 1M urea in the enzymolysis product solution; (c) contain 1%SDS in the enzymolysis product solution; (d) contain 1M sodium chloride in the enzymolysis product solution.
Fig. 5 is that the beta-caseins of different amounts are directly analyzed with bovine serum albumin(BSA) enzymolysis product potpourri and utilized the MALDI-TOF-MS after the open pipe capillary column enrichment of basic zirconium phosphate base group modification to scheme.(a) beta-casein: bovine serum albumin(BSA), 1: 1, directly analyze; (b) beta-casein: bovine serum albumin(BSA), 1: 1, enrichment post analysis; (c) beta-casein: bovine serum albumin(BSA), 1: 10, directly analyze; (d) beta-casein: bovine serum albumin(BSA), 1: 10, enrichment post analysis; (e) beta-casein: bovine serum albumin(BSA), 1: 100, directly analyze; (f) beta-casein: bovine serum albumin(BSA), 1: 100, enrichment post analysis.
Fig. 6 is that 30 μ L milk protein extract enzymes are cut the MALDI-TOF-MS figure that liquid utilizes the open pipe capillary column enrichment front and back of basic zirconium phosphate base group modification.(a) before the enrichment, (b) after the enrichment.
Fig. 7 is and receives upgrading LC-ESI-MS and combine and realize that the automatic on-line enrichment analyzes phosphoeptide.
Embodiment
0.1M HCl solution is pushed capillary column with the micro-sampling pump, with 1 μ L/min left and right sides flow velocity flushing 10min, seal end with rubber plug after then kapillary being filled with HCl solution, normal temperature leaves standstill 30min.HCl solution is released, water wash to effluent pH be about 7,0.1M NaOH solution is injected kapillary, with 1 μ L/min left and right sides flow velocity flushing 10min, seal end with rubber plug after then kapillary being filled with NaOH solution, normal temperature leaves standstill 2h.NaOH solution is released, water wash to effluent pH be about 7.
10% 3-aminopropyl triethoxysilane (APES) methanol solution (v/v) is pushed in the kapillary, and 8h will be heated in 70 ℃ of water-baths of kapillary immersion in sealed hair tubule two ends.Then APES solution is released, with 20 μ L methanol solutions flushing kapillary, nitrogen dries up.
With syringe fast with POCl
3Solution pushes in the capillary column, seals end, normal-temperature reaction 12h.With POCl
3Solution is released, and with 20 μ L methanol solutions flushing kapillary, nitrogen dries up.
Take by weighing the ZrOCl of 1mg
2Powder is dissolved in it in 1ml water, then ZrOCl
2Hydrolysis fast fully after the vibration, pushes in the quartz capillary with syringe, leaves standstill reaction 4h.Solution in the pipe is released, with the pure water rinsing capillary column to effluent pH be 7.
The enzyme of standard phosphorylating protein alpha-casein matter is cut liquid 50%ACN, the 0.1%TFA solution dilution is to 2pmol/ μ L, push with the flow velocity of 0.2 μ L/min in the open pipe capillary column (internal diameter 50 μ m, long 150cm) of basic zirconium phosphate base group modification with the micro-sampling pump, altogether sample introduction 20 μ L.With the 50%ACN of 50 μ L, 0.1TFA% solution and 50 μ L pure water are with the flow velocity of 1.0 μ L/min elder generation post-flush capillary column inwall, to remove impurity such as the non-phosphopeptide that sticks on the inwall and salt.Inject the capillary column wash-out by the phosphoeptide of special absorption with 0.1M ammonia spirit 10 μ L with 0.2 μ L/min flow velocity, the solution under the wash-out is put on the MALDI-TOF target, point is gone up DHB solution (50%ACN, 0.1%H behind the natural air drying
3PO
4), carry out mass spectrophotometry after to be crystallized.During not enrichment, have a lot of non-phosphopeptides to occur, only detect 7 phosphoeptides, as Fig. 2 a, and after the open pipe capillary column enrichment that utilizes the basic zirconium phosphate base group modification, most of non-phosphopeptide is removed, and has detected 16 phosphoeptides, shown in Fig. 2 b.Detected phosphoeptide has carried out mark in the drawings.
Utilize the open pipe capillary column load β casein protein enzymolysis product of basic zirconium phosphate base group modification, enrichment is carried out in the operation that repeats embodiment 2, MALDI-TOF-MS analyzes.During not enrichment, signal to noise ratio (S/N ratio) is very low, and after the enrichment, most of non-phosphopeptide disappears, and detects two phosphoeptides, becomes two peaks the strongest in the spectrogram, shown in Fig. 3 a-b.
Embodiment 4
The alpha-casein matter enzymolysis product 20 μ L (2pmol/ μ L) that contain 1M urea, 1%SDS and 1MNaCl are respectively pushed open pipe capillary column (the internal diameter 75 μ m of basic zirconium phosphate base group modification respectively, column length 50cm), repeat above-mentioned enriching step, carry out MALDI-TOF-MS and analyze.The alpha-casein matter enzyme that utilizes 20 μ L not contain urea, SDS and NaCl is simultaneously cut liquid and is contrasted.There is not significant difference between their concentration effects, shown in Fig. 4 a-d.Detected phosphoeptide has carried out mark in the drawings.
Embodiment 5
The enzyme of alpha-casein matter and bovine serum albumin white matter is cut peptide section solution respectively with amount of substance ratio 1: 1, and 1: 10,1: 100 ratio was mixed mutually, and bovine serum albumin white matter concentration keeps 2pmol/ μ L, and alpha-casein matter concentration reduces successively.The open pipe capillary column (internal diameter 50 μ m, column length 50cm) that utilizes the basic zirconium phosphate base group modification carries out enrichment according to above-mentioned steps to the potpourri of three kinds of ratios, and MALDI-TOF-MS analyzes.Sample to alpha-casein matter and bovine serum albumin white matter are mixed with 1: 100 ratio of amount of substance ratio still can enrichment detect 4 phosphoeptides, and control sample can only detect 1 phosphoeptide, shown in Fig. 5 a-f.
30 μ L milk are dissolved in the NH of 900 μ L 50mM
4HCO
3In the solution, vibration mixing, centrifugal 15 minutes at 16000r/min.Get clear liquid, repeated centrifugation 3 times.With clear liquid 37 ℃ of water-bath reduction 4h in the solution that contains urea (8M) and DTT (10mM), add IAA (10mM) dark place placement 0.5h and carry out alkylation, enzymolysis is hatched in 37 ℃ of water-baths.
30 μ L milk protein enzymes are cut liquid push the open pipe capillary column (internal diameter 50 μ m, long 150cm) of basic zirconium phosphate base group modification with the micro-sampling pump with the flow velocity of 0.3 μ L/min, carry out enrichment according to above-mentioned steps, MALDI-TOF-MS analyzes.Can enrichment detect 10 phosphoeptides, and control sample can only detect 4 phosphoeptides, shown in Fig. 6 a-b.
Embodiment 7
Open pipe capillary column (internal diameter 50 μ m with the basic zirconium phosphate base group modification, long 150cm) is connected on and receives before the pre-column of upgrading LC-ESI-MS liquid phase, utilize automatic sampler, Control Software by robotization, easily realize upward sample, cleaning, wash-out and the mass spectrophotometry automatically of sample in enormous quantities, as shown in Figure 7.
Claims (9)
1. an open pipe capillary column that is used for enrichment acid peptide or phosphorylating protein is characterized by the functional group that inwall has alternative absorption phosphoeptide or phosphorylating protein.
2. open pipe capillary column according to claim 1 is characterized by its inside surface silicon hydroxyl and is connected with phosphoeptide or phosphorylating protein sorbing material chemistry.
3. open pipe capillary column according to claim 2 is characterized in that the phosphoeptide or the phosphorylating protein sorbing material that connect are basic zirconium phosphate.
4. the method for enrichment acid peptide or phosphorylating protein is characterized in that may further comprise the steps:
(1) the described functional open pipe capillary column of preparation claim 1;
(2) sample is injected in the open pipe capillary column, hatches with selective adsorption phosphoeptide or phosphorylating protein;
(3) eluent is injected the open pipe capillary column, phosphoeptide or phosphorylating protein are carried out wash-out;
(4) phosphoeptide behind the wash-out or phosphorylating protein are carried out mass spectrophotometry.
5. the method for enrichment acid peptide according to claim 4 or phosphorylating protein, when it is characterized in that step (2) selective adsorption phosphoeptide or phosphorylating protein, can adopt continuous injection form or sample introduction after static state hatch.
6. the method for enrichment acid peptide according to claim 4 or phosphorylating protein, it is characterized in that step (2) is finished after, cleaning fluid is injected the open pipe capillary column, remove non-phosphopeptide or non-phosphorylating protein and salt, carry out step (3) again.
7. the method for enrichment acid peptide according to claim 4 or phosphorylating protein, it is characterized in that step (4) for the phosphoeptide behind the wash-out or phosphorylating protein sample spot on the MALDI target, add matrix and carry out MALDI-TOF-MS and analyze.
8. the method for enrichment acid peptide according to claim 4 or phosphorylating protein is characterized in that step (4) for to combine with electrospray ionization mass spectrum, and phosphoeptide or phosphorylating protein are analyzed in the automatic on-line enrichment.
9. the method for enrichment acid peptide according to claim 8 or phosphorylating protein is characterized in that: use the microlitre or the upgrading electrospray ionization mass spectrum of receiving when automatic on-line enrichment acid peptide or phosphorylating protein are analyzed.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN200810211727A CN101685051A (en) | 2008-09-23 | 2008-09-23 | Open-tubular capillary column enriching phosphoeptide or phosphorylated protein and method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN200810211727A CN101685051A (en) | 2008-09-23 | 2008-09-23 | Open-tubular capillary column enriching phosphoeptide or phosphorylated protein and method |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN101685051A true CN101685051A (en) | 2010-03-31 |
Family
ID=42048319
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN200810211727A Pending CN101685051A (en) | 2008-09-23 | 2008-09-23 | Open-tubular capillary column enriching phosphoeptide or phosphorylated protein and method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101685051A (en) |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101967181A (en) * | 2010-09-21 | 2011-02-09 | 南京农业大学 | Method for accumulating cotton phosphorylated protein |
| CN102909000A (en) * | 2012-09-06 | 2013-02-06 | 成都理工大学 | Method for preparing molecular imprinting solid-phase extraction film in separation capillary |
| CN105403638A (en) * | 2015-12-22 | 2016-03-16 | 复旦大学 | Liquid phase open tubular column with stationary phase of polymethyl methacrylate and titania and production method and application thereof |
| CN106546676A (en) * | 2016-11-02 | 2017-03-29 | 华南师范大学 | Fibrin modifies open tubular column and the application in monoclonal antibody isomer separation |
| CN107462651A (en) * | 2016-06-02 | 2017-12-12 | 复旦大学 | The enrichment method of phosphate cpd in biological sample based on titanium immobilization magnetic microsphere |
| CN113607868A (en) * | 2021-06-15 | 2021-11-05 | 广东省农业科学院农业生物基因研究中心 | Online automatic analysis device and method for phosphoproteomics |
| WO2022083799A1 (en) * | 2021-06-15 | 2022-04-28 | 广东省农业科学院农业生物基因研究中心 | Immobilized metal ion affinity chromatographic packing, chromatographic column, and preparation method therefor |
-
2008
- 2008-09-23 CN CN200810211727A patent/CN101685051A/en active Pending
Cited By (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101967181A (en) * | 2010-09-21 | 2011-02-09 | 南京农业大学 | Method for accumulating cotton phosphorylated protein |
| CN102909000A (en) * | 2012-09-06 | 2013-02-06 | 成都理工大学 | Method for preparing molecular imprinting solid-phase extraction film in separation capillary |
| CN102909000B (en) * | 2012-09-06 | 2014-10-08 | 成都理工大学 | Method for preparing molecular imprinting solid-phase extraction film in separation capillary |
| CN105403638A (en) * | 2015-12-22 | 2016-03-16 | 复旦大学 | Liquid phase open tubular column with stationary phase of polymethyl methacrylate and titania and production method and application thereof |
| CN105403638B (en) * | 2015-12-22 | 2018-02-27 | 复旦大学 | Polymethyl methacrylate titanium dioxide open tubular column and preparation method thereof and application |
| CN107462651A (en) * | 2016-06-02 | 2017-12-12 | 复旦大学 | The enrichment method of phosphate cpd in biological sample based on titanium immobilization magnetic microsphere |
| CN106546676A (en) * | 2016-11-02 | 2017-03-29 | 华南师范大学 | Fibrin modifies open tubular column and the application in monoclonal antibody isomer separation |
| CN106546676B (en) * | 2016-11-02 | 2019-06-18 | 华南师范大学 | Fibrin modifies open tubular column and the application in monoclonal antibody isomer separation |
| CN113607868A (en) * | 2021-06-15 | 2021-11-05 | 广东省农业科学院农业生物基因研究中心 | Online automatic analysis device and method for phosphoproteomics |
| CN113607868B (en) * | 2021-06-15 | 2022-03-15 | 广东省农业科学院农业生物基因研究中心 | Online automatic analysis device and method for phosphoproteomics |
| WO2022083799A1 (en) * | 2021-06-15 | 2022-04-28 | 广东省农业科学院农业生物基因研究中心 | Immobilized metal ion affinity chromatographic packing, chromatographic column, and preparation method therefor |
| WO2022117115A1 (en) * | 2021-06-15 | 2022-06-09 | 广东省农业科学院农业生物基因研究中心 | On-line automatic analysis device and analysis method for phosphoproteomics |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Ma et al. | Immobilized enzyme reactors in proteomics | |
| Haselberg et al. | Capillary electrophoresis–mass spectrometry for the analysis of intact proteins 2007–2010 | |
| CN101685051A (en) | Open-tubular capillary column enriching phosphoeptide or phosphorylated protein and method | |
| Rainer et al. | Analysis of protein phosphorylation by monolithic extraction columns based on poly (divinylbenzene) containing embedded titanium dioxide and zirconium dioxide nano‐powders | |
| Zhao et al. | Capillary electrophoresis‐mass spectrometry for analysis of complex samples | |
| Bladergroen et al. | Solid‐Phase Extraction Strategies to Surmount Body Fluid Sample Complexity in High‐Throughput Mass Spectrometry‐Based Proteomics | |
| Zhou et al. | Proteomic reactors and their applications in biology | |
| WO2003027682A2 (en) | Controlling isotope effects during fractionation of analytes | |
| CA2557997A1 (en) | Determination of analyte characteristics based upon binding properties | |
| AU2002341862A1 (en) | Controlling isotope effects during fractionation of analytes | |
| CN106770814B (en) | A kind of protein chromatography separation platform and its application | |
| CN107402269A (en) | Integrated proteomics sample pre-treatments platform and its application based on SCX/SAX mixed fillers | |
| US7407812B2 (en) | Method and kit for the isolation of phosphorylated peptides | |
| Jiang et al. | Technologies and methods for sample pretreatment in efficient proteome and peptidome analysis | |
| Olszowy et al. | Urine sample preparation for proteomic analysis | |
| Schaefer et al. | A peptide preconcentration approach for nano‐high‐performance liquid chromatography to diminish memory effects | |
| Hedrick et al. | Digestion, purification, and enrichment of protein samples for mass spectrometry | |
| Pero-Gascon et al. | Enrichment of histidine containing peptides by on-line immobilised metal affinity solid-phase extraction capillary electrophoresis-mass spectrometry | |
| CN111721939A (en) | Mass spectrum detection pretreatment method and treatment kit for protein gel sample | |
| CN101196527A (en) | Method for fast enriching and appraising phosphopeptide on MALDI-TOF-MS sample target | |
| CN102760543B (en) | Hydrophilic metal ion immobilization affinity magnetic bead and preparation and application thereof | |
| US20040053356A1 (en) | Enzyme/chemical reactor based protein processing method for proteomics analysis by mass spectrometry | |
| WO2017166897A1 (en) | Proteomic reactor, protein chromatographic separation platform and use thereof | |
| US20030153729A1 (en) | Enzyme/chemical reactor based protein processing method for proteomics analysis by mass spectrometry | |
| US20170261513A1 (en) | Monoclonal antibody based online phosphoprotein proteomics analysis method using microbore hollow fiber enzymatic reactor-tandem mass spectrometry |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Open date: 20100331 |