CN103543139A - Method for detecting low-abundance protein in blood plasma through surface-enhanced Raman spectroscopy (SERS) - Google Patents
Method for detecting low-abundance protein in blood plasma through surface-enhanced Raman spectroscopy (SERS) Download PDFInfo
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Abstract
The invention relates to a method for detecting a low-abundance protein in blood plasma through surface-enhanced Raman spectroscopy (SERS). The method comprises the following steps: adding a standard blood plasma sample into a centrifugal column of a hydrophobic polymer, performing vibration in a shaking table, and performing centrifugation to obtain a low-abundance protein solution; mixing the low-abundance protein solution with a blotting immobilized matrix and the standard blood plasma sample, and performing incubation to obtain an enrichment blood plasma low-abundance protein blotting immobilized matrix; adding glacial acetic acid and SERS active metal nano-particles; performing incubation to obtain a low-abundance protein SERS enhanced matrix mixed solution; performing SERS detection on the mixed solution; through the operation, acquiring blood plasma low-abundance protein SERS data of a cancer patient with an unknown cancer type; and determining the unknown cancer type by comparing the data with Raman spectral databases of healthy people and known cancer types. The method has the advantages of short detection time, low required power and good economic and social benefits; the immobilized matrix is low in cost; an enrichment process is simple and easy to operate.
Description
Technical field
The present invention relates to protein analysis detection field, particularly a kind of method of utilizing Surface enhanced raman spectroscopy (SERS) to detect low-abundance protein in blood plasma.
Background technology
Raman spectrum is a kind of molecular vibration spectrum, is scattering spectrum analyze different from incident light frequency obtained to molecular vibration, rotation aspect information, and is applied to a kind of analytical approach of molecular structure research.Raman spectrum analysis process operation is simple and quick, highly sensitive, be a kind of Non-Destructive Testing, but raman spectral signal is faint, is subject to autofluorescence and disturbs, and therefore utilizes Raman spectroscopy directly to detect and has certain limitation.Surface enhanced raman spectroscopy (Surface-enhanced Raman spectroscopy, abbreviation SERS) technology is a kind of conventional Raman signal Enhancement Method, when molecular adsorption is when some textured metal (as Au, Ag, Cu and Pt etc.) is surperficial, to produce internal field's enhancing or electric charge and shift, make the Raman scattering strength increase 10 of these molecules
4～10
14doubly, SERS effect that Here it is.SERS technology energy cancellation biomolecule fluorescence, the signal of acquisition is desirable, detects power demand lower, little, highly sensitive to biological tissue samples damage, can realize Single Molecule Detection.
Human plasma albumen contains the albumen that derives from nearly all cell, tissue, organ, yet, the abundance difference of plasma proteins is huge, the albumen of 21 kinds of main high abundances and medium abundance in blood plasma, as albumin, IgG, alpha1 Anti-trypsin, alpha2 Macroglobulin, transferrins etc. account for the more than 99% of blood plasma total protein content, remaining 1% is comprised of the even extremely low-abundance protein of numerous low abundance.While there is pathology in human tissue organ, albumen from pathological tissues or cell can enter into hematological system, pathology, the physiological situation of these protein and body are closely related, therefore, it is to find one of most worthy sample with various diseases associated biomolecule mark that these protein are carried out to determination and analysis, these protein major parts are all low-abundance proteins, yet, low-abundance protein is difficult to detect, and how effectively determination and analysis low-abundance protein is the challenging difficult problem in analysis science field always.Difficult point mainly aspect two, the one, the content of low-abundance protein is lower than the detection limit of analytical instrument; On the other hand, in plasma proteins, the difference of high and low abundance protein content is huge, often can reach 10
12, the signal of low-abundance protein is easily covered by high-abundance proteins.
At present the detection method of low-abundance protein is mainly contained to following two kinds: first method is first to pass through the separated high and low abundance protein of two-dimensional gel electrophoresis, then with mass spectrum, detects analysis after the low-abundance protein enzymolysis that separation is obtained; Second method is that the peptide section after chromatographic resolution protein digestion, finally carries out Mass Spectrometer Method analysis to peptide section by after the first mixed enzymolysis of high and low abundance protein.The problems such as said method all exists required sample size large, and consumptive material is expensive, and step is complicated, length consuming time.Mass spectrophotometry is subject to the interference of the factors such as eluent in addition, the quality protein such as fubaritic differentiation, and also mass spectrometer is expensive, is difficult to extensive popularization.
Summary of the invention
The object of the invention is to propose a kind of method of the SERS of utilization technology for detection blood plasma low-abundance protein Surface enhanced raman spectroscopy.This method adopts hydrophobic polymer to remove after the high-abundance proteins in blood plasma, utilize trace immobilization matrix enrichment low-abundance protein, utilize SERS technology for detection to obtain blood plasma low-abundance protein Surface enhanced raman spectroscopy, set up kinds cancer blood plasma low-abundance protein Surface enhanced raman spectroscopy database, through multivariate statistical analysis, carry out cluster analysis, the differentiation scatter diagram that the people that secures good health is corresponding with cancer patient's blood plasma low-abundance protein Surface enhanced raman spectroscopy, realizes the differentiation of various cancers accordingly.The features such as it is quick, simple to operate, with low cost that the present invention has, can effectively detect the Surface enhanced raman spectroscopy of low-abundance protein in blood plasma, thereby overcome deficiency of the prior art.
For achieving the above object, the technical solution used in the present invention is as follows:
(1) remove blood plasma high-abundance proteins
Getting hydrophobic polymer has in the collection tube of centrifugal column in cover, adds eluent, fully mixes, and after the centrifugal 1～5min of 3000～6000rpm, the liquid in collection tube is outwelled, and obtains in triplicate the hydrophobic polymer through eluent wash-out.Get blood plasma standard model, add the eluent of 1～10 times of volume to dilute, join and contain in the centrifugal column of the hydrophobic polymer of eluent wash-out, utilize pipettor repeatedly to aspirate fully and mix, be placed in shaking table 200～350rpm concussion 10～15min, 2000～5000rpm is centrifugal, removes sediment, collects the low-abundance protein solution on upper strata.
(2) enrichment blood plasma low-abundance protein
Get trace immobilization matrix, according to trace immobilization Ji Zhi ︰ blood plasma standard model, it is the ratio of (1～2) g ︰ (10～20) μ l, the low-abundance protein solution of collecting with step (1) mixes, hatch 8～15min, then trace immobilization matrix is proceeded in rinsing liquid and soak 10～20min, taking-up at room temperature dries up standby.
(3) low-abundance protein SERS strengthens the preparation of matrix mixed liquor
The trace immobilization matrix that step (2) is dried up shreds and is collected in centrifuge tube, add glacial acetic acid to dissolve, fully stir until while being rendered as transparent colloid state completely, add SERS active metal nano particle, fully stir, 15～30min, stratification are hatched in 42～65 ℃ of water-baths, until solid phase impurity is separated out, make low-abundance protein SERS and strengthen matrix mixed liquor.
(4) blood plasma low-abundance protein SERS spectral detection
Get low-abundance protein SERS enhancing matrix mixed liquor 2～20 μ l and carry out SERS detection.
(5) cancer is differentiated
Get a kind of patient's blood plasma of unknown cancer types, adopt the operation of step (1) to (4), obtain the cancer patient's of described unknown cancer types blood plasma low-abundance protein Surface enhanced raman spectroscopy data; These data and the Healthy People of having set up and the blood plasma low-abundance protein Surface enhanced raman spectroscopy database of known cancer kind are compared, determine the kind of unknown cancer.
Described hydrophobic polymer refers to ion exchange resin or silica gel.
Described eluent refers to phosphate buffer, and the concentration range of phosphate radical is every liter of 25～60 mM (mmol/L), and pH value is 4.
Described hydrophobic polymer and the amount ratio of eluent are the ratio of (2～25) g ︰ (100～1000) μ l,
Described trace immobilization matrix refers to cellulose acetate membrane, nitrocellulose filter, nylon membrane or PVDF membrane.
Described SERS active metal nano particle refers to Nano silver grain or golden nanometer particle.
The amount ratio of the use amount of described hydrophobic polymer and blood plasma standard model is (2～25) g:(10～100) μ l.
Described trace immobilization matrix and the amount ratio of glacial acetic acid are (0.2～2) g ︰ (150～1000) μ l.
The amount ratio of described SERS active metal nano particle and trace immobilization matrix is (5～50) mM: (0.2～2) g.
Described rinsing liquid refers to the potpourri of glacial acetic acid, 95% ethanol, distilled water, and it consists of glacial acetic acid: 95% ethanol: the volume ratio of distilled water is (1～3): (4～9): (8～12);
Described SERS detects, and the laser power of use is 0.1～20mW, and laser excitation wavelength is 450～800nm, and the measurement range of spectrum is 400～3500cm
-1;
The preparation method of described Nano silver grain is:
Adopt oxammonium hydrochloride reduction preparation of silver nano particle, 4.5ml sodium hydroxide solution (0.1M) is mixed with 5ml oxammonium hydrochloride solution (0.06M), join 90ml AgNO
3solution (1.1 * 10
-3m) in, vortex stirs 1min, can obtain newborn grey Nano silver grain, and particle diameter is approximately 40～60nm.Above-mentioned Nano silver grain, with the centrifugal 10min～15min of 4000～5000rpm, is abandoned to supernatant, the concentrated Nano silver grain of lower floor at room temperature lucifuge seal up for safekeeping standby.
The preparation method of described golden nanometer particle is:
Get containing 500 milliliters of the aqueous solution of chloraurate of 1mmol/L, be heated in rapid stirring, add after boiling the sodium citrate solution of 50 milliliters of 38.8mmol/L.Continue heating 10 minutes, wait the color of solution from faint yellow variation darkviolet, to remove coverture agitating heating 15 minutes again, obtain golden nanometer particle.Particle diameter is approximately 35～45nm.Above-mentioned golden nanometer particle, with the centrifugal 10min～15min of 5000～8000rpm, is abandoned supernatant, the concentrated golden nanometer particle of lower floor at room temperature lucifuge seal up for safekeeping standby.
Compared with prior art, the present invention has following beneficial effect:
1, the present invention adds glacial acetic acid when SERS active nano material mixes mutually with low-abundance protein, make environment keep faintly acid, because the SERS active nano materials itself such as silver or golden nanometer particle are electronegative, and the isoelectric point of blood plasma low abundance proteins is all below PH7.5, under sour environment, blood plasma low abundance proteins protein is all positively charged, low-abundance protein sample to be measured like this strengthens stromal surface by static adsorbed close at SERS, and promotion colloid aggregation, significantly improves detection sensitivity.The present invention uses SERS technology for detection low-abundance protein, and only need 1～10s detection time, detects required laser power and is low to moderate 0.1mW, and therefore, the present invention has fast, can't harm, sensitive detection advantage.
2, the present invention selects the immobilization matrix such as cellulose acetate film as the material of enrichment blood plasma low-abundance protein, material itself is cheap, enrichment process is simple, greatly reduce cost, and the SERS spectrum of the low-abundance protein detect obtaining comprises rich in protein structural information, can effectively make up the deficiency that mass spectroscopy is identified low abundance proteins.
3, the further differentiation of other relevant diseases of application of method of the present invention, has good economic and social benefit.
Accompanying drawing explanation
Fig. 1 is the liver cancer patient blood plasma low-abundance protein Surface enhanced raman spectroscopy figure that the present invention records.
Fig. 2 is that the present invention analyzes scatter diagram to the Surface enhanced raman spectroscopy PCA of Healthy People and liver cancer patient blood plasma low-abundance protein.
Fig. 3 is the nasopharyngeal carcinoma cancer patient blood plasma low-abundance protein Surface enhanced raman spectroscopy figure that the present invention records.
Fig. 4 is that the present invention analyzes scatter diagram to the Surface enhanced raman spectroscopy PCA of Healthy People and nasopharyngeal carcinoma cancer patient blood plasma low-abundance protein.
Fig. 5 is the Plasma of Patient With Gastric Cancer low-abundance protein Surface enhanced raman spectroscopy figure that the present invention records.
Fig. 6 is that the present invention analyzes scatter diagram to the Surface enhanced raman spectroscopy PCA of Healthy People and Plasma of Patient With Gastric Cancer low-abundance protein.
Embodiment
The intensity that in Fig. 1, Fig. 3 and Fig. 5, ordinate is spectral line, unit is the peak position that arbitrary unit (a.u) horizontal ordinate is each characteristic spectral line, with wave number (cm
-1) represent.
1, based on Nano silver grain, strengthen the liver cancer patient blood plasma low-abundance protein SERS spectral detection of substrate
(1) remove the high-abundance proteins in liver cancer patient blood plasma
Getting 2g ion exchange resin has in the collection tube of centrifugal column in cover, adds 100 μ l eluents, mixes, and after the centrifugal 5min of 3000rpm, the liquid in collection tube is outwelled, in triplicate.Get in the eluent that 10 μ l liver cancer patient blood plasma standard models are diluted in 100 μ l and mix, join in the centrifugal column that contains ion exchange resin, utilize pipettor repeatedly to aspirate fully and mix, be placed in shaking table 200rpm concussion 15min, the centrifugal collection liver cancer patient of 2000rpm blood plasma low-abundance protein solution.
(2) enrichment liver cancer patient blood plasma low-abundance protein
Get the liver cancer patient blood plasma low-abundance protein solution that 0.2g cellulose acetate film and step (1) collect and jointly hatch 15min, then cellulose acetate film is proceeded in rinsing liquid and soak 20min, take out at room temperature dry up standby.
(3) low-abundance protein SERS strengthens the preparation of matrix mixed liquor
The cellulose acetate film that step (2) is dried up shreds and is collected in 1.5ml centrifuge tube, add glacial acetic acid 150 μ l to dissolve cellulose acetate film, fully stir until while being rendered as transparent colloid state completely, add Nano silver grain, fully stir, 30min, stratification are hatched in 42 ℃ of water-baths, until solid phase impurity is separated out, make low-abundance protein SERS and strengthen matrix mixed liquor
(4) liver cancer patient blood plasma low-abundance protein SERS spectral detection
Get liver cancer patient blood plasma low-abundance protein SERS enhancing matrix mixed liquor 2 μ l and carry out SERS detection.The SERS spectrum of liver cancer patient blood plasma low-abundance protein detected as shown in Figure 1.
(5) liver cancer patient is differentiated
Repeating step (1), to the operation of (4), obtains the blood plasma low-abundance protein Surface enhanced raman spectroscopy data of a plurality of liver cancer patients and Healthy People; By data list input SPSS software corresponding to described SERS spectrum, data transposition is set up sample matrix, PCA method in application SPSS software analysis tool hurdle is calculated, and sets up corresponding Healthy People-liver cancer patient and analyzes scatter diagram as shown in Figure 2, thereby realize liver cancer patient, differentiates.
Based on golden nanometer particle, strengthen the plasma of patients with nasopharyngeal carcinoma low-abundance protein SERS spectral detection of substrate
(1) remove the high-abundance proteins in plasma of patients with nasopharyngeal carcinoma
Getting 25g ion exchange resin has in the collection tube of centrifugal column in cover, adds 1000 μ l eluents and hydrophobic polymer fully to mix, and after the centrifugal 1min of 6000rpm, the liquid in collection tube is outwelled, in triplicate.Get in the eluent that 100 μ l plasma of patients with nasopharyngeal carcinoma standard models are diluted in 1000 μ l and mix, join in the centrifugal column that contains ion exchange resin, utilize pipettor repeatedly to aspirate fully and mix, be placed in shaking table 350rpm concussion 10min, the centrifugal collection plasma of patients with nasopharyngeal carcinoma of 5000rpm low-abundance protein solution.
(2) enrichment plasma of patients with nasopharyngeal carcinoma low-abundance protein
Get the plasma of patients with nasopharyngeal carcinoma low-abundance protein solution that 2g PVDF membrane and step (2) collect and jointly hatch 8min, then PVDF membrane is proceeded in rinsing liquid and soak 10min, take out at room temperature dry up standby.
(3) plasma of patients with nasopharyngeal carcinoma low-abundance protein SERS spectral detection
The PVDF membrane that step (2) is dried up shreds and is collected in 7ml centrifuge tube, add glacial acetic acid 1000 μ l to dissolve PVDF membrane, fully stir until while being rendered as transparent colloid state completely, add golden nanometer particle, fully stir, 15min, stratification are hatched in 65 ℃ of water-baths, until solid phase impurity is separated out, make low-abundance protein SERS and strengthen matrix mixed liquor.
(4) plasma of patients with nasopharyngeal carcinoma low-abundance protein SERS spectral detection
Get plasma of patients with nasopharyngeal carcinoma low-abundance protein SERS enhancing matrix mixed liquor 20 μ l and carry out SERS detection.The SERS spectrum of plasma of patients with nasopharyngeal carcinoma low-abundance protein detected as shown in Figure 3.
(5) Nasopharyngeal Carcinoma Patients is differentiated
Repeating step (1), to the operation of (4), obtains the blood plasma low-abundance protein Surface enhanced raman spectroscopy data of a plurality of Nasopharyngeal Carcinoma Patients and Healthy People; By data list input SPSS software corresponding to described SERS spectrum, data transposition is set up sample matrix, PCA method in application SPSS software analysis tool hurdle is calculated, and sets up corresponding Healthy People-Nasopharyngeal Carcinoma Patients and analyzes scatter diagram as shown in Figure 4, thereby realize Nasopharyngeal Carcinoma Patients, differentiates.
1, based on Nano silver grain, strengthen the Plasma of Patient With Gastric Cancer low-abundance protein SERS spectral detection of substrate
(1) remove the high-abundance proteins in Plasma of Patient With Gastric Cancer
Getting 2g silica gel has in the collection tube of centrifugal column in cover, adds 150 μ l eluents, mixes, and after the centrifugal 5min of 3000rpm, the liquid in collection tube is outwelled, in triplicate.Get in the eluent that 10 μ l Plasma of Patient With Gastric Cancer standard models are diluted in 100 μ l and mix, join in the centrifugal column that contains silica gel, utilize pipettor repeatedly to aspirate fully and mix, be placed in shaking table 200rpm concussion 15min, the centrifugal collection liver cancer patient of 2000rpm blood plasma low-abundance protein solution.
(2) enrichment Plasma of Patient With Gastric Cancer low-abundance protein
Get the Plasma of Patient With Gastric Cancer low-abundance protein solution that 2g nitrocellulose filter and step (1) collect and jointly hatch 15min, then nitrocellulose membrane is proceeded in rinsing liquid and soak 20min, take out at room temperature dry up standby.
(3) low-abundance protein SERS strengthens the preparation of matrix mixed liquor
The nitrocellulose membrane that step (2) is dried up shreds and is collected in 1.5ml centrifuge tube, add glacial acetic acid 1000 μ l to dissolve nitrocellulose membrane, fully stir until while being rendered as transparent colloid state completely, add Nano silver grain, fully stir, 18min, stratification are hatched in 52 ℃ of water-baths, until solid phase impurity is separated out, make low-abundance protein SERS and strengthen matrix mixed liquor.
(4) blood plasma low-abundance protein SERS spectral detection
Get Plasma of Patient With Gastric Cancer low-abundance protein SERS enhancing matrix mixed liquor 2 μ l and carry out SERS detection.The SERS spectrum of Plasma of Patient With Gastric Cancer low-abundance protein detected as shown in Figure 5.
(5) patients with gastric cancer is differentiated
Repeating step (1), to the operation of (4), obtains the blood plasma low-abundance protein Surface enhanced raman spectroscopy data of a plurality of patients with gastric cancer and Healthy People; By data list input SPSS software corresponding to described SERS spectrum, data transposition is set up sample matrix, PCA method in application SPSS software analysis tool hurdle is calculated, and sets up corresponding Healthy People-patients with gastric cancer and analyzes scatter diagram as shown in Figure 6, thereby realize patients with gastric cancer, differentiates.
Certainly, the process of the present embodiment is only for illustrating technical scheme of the present invention, those skilled in the art are according to enlightenment of the present invention, also can be easy to expect other the multiple low-abundance proteins to except blood plasma low-abundance protein, as organize low-abundance protein, saliva low-abundance protein, seminal fluid low-abundance protein and urine low-abundance protein etc., with similar method, distinguish and differentiate.But all those skilled in the art are because of technology involved in the present invention enlightenment, and adopt to be equal to, replace or technical scheme that equivalent deformation mode forms all drops in protection scope of the present invention.
Claims (11)
1. a detection method for blood plasma low-abundance protein Surface enhanced raman spectroscopy, is characterized in that:
(1) remove blood plasma high-abundance proteins
Getting hydrophobic polymer has in the collection tube of centrifugal column in cover, adds eluent, fully mixes, and after centrifugal 1～5 min of 3000～6000rpm, the liquid in collection tube is outwelled, and obtains in triplicate the hydrophobic polymer through eluent wash-out;
Get blood plasma standard model, add the eluent of 1～10 times of volume to dilute, join and contain in the centrifugal column of the hydrophobic polymer of eluent wash-out, utilize pipettor repeatedly to aspirate fully and mix, be placed in shaking table 200～350 rpm concussion 10～15 min, 2000～5000 rpm are centrifugal, remove sediment, collect the low-abundance protein solution on upper strata;
(2) enrichment blood plasma low-abundance protein
Get trace immobilization matrix, the low-abundance protein solution of collecting with step (1) mixes, and hatches 8～15 min, then trace immobilization matrix is proceeded in rinsing liquid and soaks 10～20 min, and taking-up at room temperature dries up standby;
(3) low-abundance protein SERS strengthens the preparation of matrix mixed liquor
The trace immobilization matrix that step (2) is dried up shreds and is collected in centrifuge tube, add glacial acetic acid to dissolve, fully stir until while being rendered as transparent colloid state completely, add SERS active metal nano particle, fully stir, 15～30 min, stratification are hatched in 42～65 ℃ of water-baths, until solid phase impurity is separated out, make low-abundance protein SERS and strengthen matrix mixed liquor;
(4) blood plasma low-abundance protein SERS spectral detection
Get low-abundance protein SERS enhancing matrix mixed liquor 2～20 μ l and carry out SERS detection;
(5) cancer is differentiated
Get a kind of patient's blood plasma of unknown cancer types, adopt the operation of step (1) to (4), obtain the cancer patient's of described unknown cancer types blood plasma low-abundance protein Surface enhanced raman spectroscopy data; These data and the Healthy People of having set up and the blood plasma low-abundance protein Surface enhanced raman spectroscopy database of known cancer kind are compared, determine the kind of unknown cancer.
2. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, while it is characterized in that described trace immobilization matrix is mixed with blood plasma standard model, its ratio is (0.2～2) g ︰ (10～100) μ l.
3. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, it is characterized in that described hydrophobic polymer refers to ion exchange resin or silica gel.
4. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, it is characterized in that described eluent refers to phosphate buffer, the concentration range of phosphate radical is every liter of 25～60 mM, and pH value is 4.
5. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, it is characterized in that described hydrophobic polymer and the amount ratio of eluent are (2～25) g ︰ (100～1000) μ l.
6. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, it is characterized in that described trace immobilization matrix refers to cellulose acetate membrane, nitrocellulose filter, nylon membrane or PVDF membrane.
7. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, it is characterized in that described SERS active metal nano particle refers to Nano silver grain or golden nanometer particle.
8. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, it is characterized in that the use amount of described hydrophobic polymer and the amount ratio of blood plasma standard model are for (2～25) g:(10～100) μ l.
9. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, it is characterized in that described trace immobilization matrix and the amount ratio of glacial acetic acid are (0.2～2) g ︰ (150～1000) μ l.
10. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, it is characterized in that the amount ratio of described SERS active metal nano particle and trace immobilization matrix is (5～50) mM: (0.2 ~ 2) g.
11. according to the detection method of the described blood plasma low-abundance protein Surface enhanced raman spectroscopy of claim 1, it is characterized in that described SERS detects, the laser power of using is 0.1～20 mW, and laser excitation wavelength is 450～800 nm, and the measurement range of spectrum is 400～3500 cm
-1.
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| CN104122247A (en) * | 2014-06-10 | 2014-10-29 | 南京大学 | Glycoprotein detection method based on molecular imprinting technique and Raman spectrum and application |
| CN106436027A (en) * | 2016-09-09 | 2017-02-22 | 中国科学院合肥物质科学研究院 | Silver nanometer square-cellulose acetate composite microballoon membrane and preparation method and purpose thereof |
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| CN110646401A (en) * | 2019-10-12 | 2020-01-03 | 福建师范大学 | SERS detection method based on hydroxyapatite nanoparticle adsorbed protein |
| CN110646401B (en) * | 2019-10-12 | 2022-04-12 | 福建师范大学 | SERS detection method based on hydroxyapatite nanoparticle adsorbed protein |
| CN112331270A (en) * | 2021-01-04 | 2021-02-05 | 中国工程物理研究院激光聚变研究中心 | Construction method of novel coronavirus Raman spectrum data center |
| CN112331270B (en) * | 2021-01-04 | 2021-03-23 | 中国工程物理研究院激光聚变研究中心 | Construction method of novel coronavirus Raman spectrum data center |
| CN114034641A (en) * | 2021-12-08 | 2022-02-11 | 重庆大学 | Narrow-band filter-based Raman on-chip detection system and method |
| CN114705794A (en) * | 2022-04-15 | 2022-07-05 | 西湖大学 | Proteomics analysis method for biological sample |
| CN114705794B (en) * | 2022-04-15 | 2022-12-02 | 西湖大学 | Proteomics analysis method for biological sample |
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