CN104483478B - A kind of detection method of Ancient Silk Textile - Google Patents
A kind of detection method of Ancient Silk Textile Download PDFInfo
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- CN104483478B CN104483478B CN201410846331.7A CN201410846331A CN104483478B CN 104483478 B CN104483478 B CN 104483478B CN 201410846331 A CN201410846331 A CN 201410846331A CN 104483478 B CN104483478 B CN 104483478B
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- 239000004753 textile Substances 0.000 title claims abstract description 11
- 238000001514 detection method Methods 0.000 title claims abstract description 8
- 241000283973 Oryctolagus cuniculus Species 0.000 claims abstract description 12
- 241000283707 Capra Species 0.000 claims abstract description 7
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 claims abstract description 7
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 30
- 239000000243 solution Substances 0.000 claims description 30
- 239000007853 buffer solution Substances 0.000 claims description 15
- 235000011187 glycerol Nutrition 0.000 claims description 15
- 238000004080 punching Methods 0.000 claims description 15
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims description 10
- BVKZGUZCCUSVTD-UHFFFAOYSA-L Carbonate Chemical compound [O-]C([O-])=O BVKZGUZCCUSVTD-UHFFFAOYSA-L 0.000 claims description 10
- 229940098773 bovine serum albumin Drugs 0.000 claims description 10
- 230000003139 buffering effect Effects 0.000 claims description 10
- 239000012153 distilled water Substances 0.000 claims description 10
- 238000002360 preparation method Methods 0.000 claims description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 10
- 238000011095 buffer preparation Methods 0.000 claims description 5
- 239000011888 foil Substances 0.000 claims description 5
- 230000000873 masking effect Effects 0.000 claims description 5
- 230000009870 specific binding Effects 0.000 claims description 5
- 238000010166 immunofluorescence Methods 0.000 abstract description 4
- 238000000034 method Methods 0.000 abstract description 4
- 108010022355 Fibroins Proteins 0.000 abstract description 2
- 238000004451 qualitative analysis Methods 0.000 abstract description 2
- 239000012895 dilution Substances 0.000 description 6
- 238000010790 dilution Methods 0.000 description 6
- 238000001237 Raman spectrum Methods 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 230000000007 visual effect Effects 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/536—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase
- G01N33/542—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase with steric inhibition or signal modification, e.g. fluorescent quenching
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
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- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
Abstract
The invention discloses a kind of detection method of Ancient Silk Textile, the method of immunofluorescence is adopted to detect Ancient Silk Textile, anti-for rabbit fibroin albumen antibody be impregnated in historical relic sample surface, after washes clean, add goat anti-rabbit igg (H+L) antibody of green fluorescein mark, form the anti-two anti-compounds of antigen-one, then washes clean is observed under historical relic sample is placed on laser confocal microscope.Whether can send green fluorescence per sample and carry out qualitative analysis.The present invention adopts the method for immunofluorescence to detect Ancient Silk Textile, on the one hand, highly sensitive, simple to operate.The interference from other albumen can be avoided on the other hand, high specificity.
Description
Technical field
The invention belongs to the detection field of Ancient Silk Textile, particularly relate to a kind of detection method of Ancient Silk Textile.
Background technology
Silk is one of Chinese history invention the earliest, is the preciousness culture of China National, but in recent years, about the origin of silk, but opinions vary, therefore how to adopt the means determination silk of science be China original become the matter of great urgency.At present the mainly technology such as Raman spectrum, X-diffraction is detected to silk goods, but due to the interference of impurity, later stage spectrum elucidation difficulty, can not judge intuitively to result.
Summary of the invention
The technical problem to be solved in the present invention is: for above-mentioned prior art Problems existing provide one intuitively, detection method efficiently.
Adopt following technical scheme: a kind of detection method of Ancient Silk Textile for this reason, it is characterized in that adopting following steps:
A) the PBS buffer preparation of pH7.4: KCl0.2g, KH
2pO
40.27g, NaCl8.0g, Na
2hPO
41.42g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 7.4; The carbonate buffer solution preparation of pH9.6: Na
2cO
30.15g, NaHCO
30.29g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 9.6; The preparation of buffering glycerine: the carbonate buffer solution of pH9.6 and glycerine are mixed according to volume ratio 1:1;
B) get the historical relic sample that three block sizes are consistent, be positioned in punching ware, be labeled as A respectively, B, C; Wash with the PBS buffer solution of pH7.4, ensure surface attachments washes clean; Respectively to adding the anti-fibroin albumen antibody of rabbit that doubly dilutes of bovine serum albumin solution 10-200 that 10-50 μ L mass concentration is 1% in A, in C, add the PBS solution of 10-50 μ LpH7.4; B does not deal with, and A, B, C is positioned over 12h in 4 DEG C of refrigerators respectively; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min, is washed three times, each three minutes;
C) respectively to add in A, B 10-50 μ L mass concentration be 1% bovine serum albumin solution 50-500 doubly dilute green fluorescein mark goat anti-rabbit IgG antibody, add the PBS solution of 10-50 μ LpH7.4 in C; Then respectively A, B, C being positioned over bottom is covered with in the box of wet towel, and sealed by box with masking foil, 37 DEG C of lucifuges hatch 0.5-2h; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min and is washed; Wash three times, each three minutes;
D) mounting is carried out respectively to the buffering glycerine adding 10 μ L in A, B, C;
E) by A, B, C observe under being positioned over laser confocal microscope;
If B, C do not send green fluorescence, and A have issued green fluorescence, illustrates that historical relic sample and the anti-fibroin albumen antibody of rabbit there occurs specific binding, proves that historical relic sample is silk goods.
Useful achievement of the present invention is: 1 the present invention adopts the method for immunofluorescence to detect Ancient Silk Textile, on the one hand, highly sensitive, simple to operate.The interference from other albumen can be avoided on the other hand, high specificity.2 compared with prior art, and cost of the present invention is low, visual result, is easy to analyze.
Embodiment
The present invention adopts the method for immunofluorescence to detect Ancient Silk Textile, anti-for rabbit fibroin albumen antibody be impregnated in historical relic sample surface, after washes clean, add the goat anti-rabbit IgG antibody of green fluorescein mark, form the anti-two anti-compounds of antigen-one, then washes clean is observed under historical relic sample is placed on laser confocal microscope.Whether can send green fluorescence per sample and carry out qualitative analysis.Below in conjunction with case study on implementation, the present invention is further described.
Embodiment 1 adopts following steps:
A) the PBS buffer preparation of pH7.4: KCl0.2g, KH
2pO
40.27g, NaCl8.0g, Na
2hPO
41.42g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 7.4; The carbonate buffer solution preparation of pH9.6: Na
2cO
30.15g, NaHCO
30.29g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 9.6; The preparation of buffering glycerine: the carbonate buffer solution of pH9.6 and glycerine are mixed according to volume ratio 1:1;
B) get the historical relic sample that three block sizes are consistent, be positioned in punching ware, be labeled as A respectively, B, C; Wash with the PBS buffer solution of pH7.4, ensure surface attachments washes clean; Respectively to adding the anti-fibroin albumen antibody of rabbit that 10 μ L mass concentrations are the bovine serum albumin solution 10 times dilution of 1% in A, in C, add the PBS solution of 10 μ LpH7.4; B does not deal with, and A, B, C is positioned over 12h in 4 DEG C of refrigerators respectively; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min, is washed three times, each three minutes;
C) respectively to adding the green fluorescein mark goat anti-rabbit IgG antibody that 10 μ L mass concentrations are the bovine serum albumin solution 50 times dilution of 1% in A, B, the PBS solution of 10 μ LpH7.4 in C, is added; Then respectively A, B, C being positioned over bottom is covered with in the box of wet towel, and sealed by box with masking foil, 37 DEG C of lucifuges hatch 0.5h; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min and is washed; Wash three times, each three minutes;
D) mounting is carried out respectively to the buffering glycerine adding 10 μ L in A, B, C;
E) by A, B, C observe under being positioned over laser confocal microscope;
If B, C do not send green fluorescence, and A have issued green fluorescence, illustrates that historical relic sample and the anti-fibroin albumen antibody of rabbit there occurs specific binding, proves that historical relic sample is silk goods.
Embodiment 2 adopts following steps:
A) the PBS buffer preparation of pH7.4: KCl0.2g, KH
2pO
40.27g, NaCl8.0g, Na
2hPO
41.42g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 7.4; The carbonate buffer solution preparation of pH9.6: Na
2cO
30.15g, NaHCO
30.29g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 9.6; The preparation of buffering glycerine: the carbonate buffer solution of pH9.6 and glycerine are mixed according to volume ratio 1:1;
B) get the historical relic sample that three block sizes are consistent, be positioned in punching ware, be labeled as A respectively, B, C; Wash with the PBS buffer solution of pH7.4, ensure surface attachments washes clean; Respectively to adding the anti-fibroin albumen antibody of rabbit that 30 μ L mass concentrations are the bovine serum albumin solution 100 times dilution of 1% in A, in C, add the PBS solution of 30 μ LpH7.4; B does not deal with, and A, B, C is positioned over 12h in 4 DEG C of refrigerators respectively; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min, is washed three times, each three minutes;
C) respectively to adding the green fluorescein mark goat anti-rabbit IgG antibody that 30 μ L mass concentrations are the bovine serum albumin solution 300 times dilution of 1% in A, B, the PBS solution of 30 μ LpH7.4 in C, is added; Then respectively A, B, C being positioned over bottom is covered with in the box of wet towel, and sealed by box with masking foil, 37 DEG C of lucifuges hatch 1h; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min and is washed; Wash three times, each three minutes;
D) mounting is carried out respectively to the buffering glycerine adding 10 μ L in A, B, C;
E) by A, B, C observe under being positioned over laser confocal microscope;
If B, C do not send green fluorescence, and A have issued green fluorescence, illustrates that historical relic sample and the anti-fibroin albumen antibody of rabbit there occurs specific binding, proves that historical relic sample is silk goods.
Embodiment 3 adopts following steps:
A) the PBS buffer preparation of pH7.4: KCl0.2g, KH
2pO
40.27g, NaCl8.0g, Na
2hPO
41.42g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 7.4; The carbonate buffer solution preparation of pH9.6: Na
2cO
30.15g, NaHCO
30.29g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 9.6; The preparation of buffering glycerine: the carbonate buffer solution of pH9.6 and glycerine are mixed according to volume ratio 1:1;
B) get the historical relic sample that three block sizes are consistent, be positioned in punching ware, be labeled as A respectively, B, C; Wash with the PBS buffer solution of pH7.4, ensure surface attachments washes clean; Respectively to adding the anti-fibroin albumen antibody of rabbit that 50 μ L mass concentrations are the bovine serum albumin solution 200 times dilution of 1% in A, in C, add the PBS solution of 50 μ LpH7.4; B does not deal with, and A, B, C is positioned over 12h in 4 DEG C of refrigerators respectively; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min, is washed three times, each three minutes;
C) respectively to adding the green fluorescein mark goat anti-rabbit IgG antibody that 50 μ L mass concentrations are the bovine serum albumin solution 500 times dilution of 1% in A, B, the PBS solution of 50 μ LpH7.4 in C, is added; Then respectively A, B, C being positioned over bottom is covered with in the box of wet towel, and sealed by box with masking foil, 37 DEG C of lucifuges hatch 2h; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min and is washed; Wash three times, each three minutes;
D) mounting is carried out respectively to the buffering glycerine adding 10 μ L in A, B, C;
E) by A, B, C observe under being positioned over laser confocal microscope;
If B, C do not send green fluorescence, and A have issued green fluorescence, illustrates that historical relic sample and the anti-fibroin albumen antibody of rabbit there occurs specific binding, proves that historical relic sample is silk goods.
Claims (1)
1. a detection method for Ancient Silk Textile, is characterized in that adopting following steps:
A) the PBS buffer preparation of pH7.4: KCl0.2g, KH
2pO
40.27g, NaCl8.0g, Na
2hPO
41.42g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 7.4; The carbonate buffer solution preparation of pH9.6: Na
2cO
30.15g, NaHCO
30.29g, dissolves with 800mL distilled water and is settled to 1000mL, regulates pH to 9.6; The preparation of buffering glycerine: the carbonate buffer solution of pH9.6 and glycerine are mixed according to volume ratio 1:1;
B) get the historical relic sample that three block sizes are consistent, be positioned in punching ware, be labeled as A respectively, B, C; Wash with the PBS buffer solution of pH7.4, ensure surface attachments washes clean; Respectively to adding the anti-fibroin albumen antibody of rabbit that doubly dilutes of bovine serum albumin solution 10-200 that 10-50 μ L mass concentration is 1% in A, in C, add the PBS solution of 10-50 μ LpH7.4; B does not deal with, and A, B, C is positioned over 12h in 4 DEG C of refrigerators respectively; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min, is washed three times, each three minutes;
C) respectively to add in A, B 10-50 μ L mass concentration be 1% bovine serum albumin solution 50-500 doubly dilute green fluorescein mark goat anti-rabbit IgG antibody, add the PBS solution of 10-50 μ LpH7.4 in C; Then respectively A, B, C being positioned over bottom is covered with in the box of wet towel, and sealed by box with masking foil, 37 DEG C of lucifuges hatch 0.5-2h; Then the PBS solution adding 2mLpH7.4 respectively, in punching ware, is soaked 5min and is washed; Wash three times, each three minutes;
D) mounting is carried out respectively to the buffering glycerine adding 10 μ L in A, B, C;
E) by A, B, C observe under being positioned over laser confocal microscope;
If B, C do not send green fluorescence, and A have issued green fluorescence, illustrates that historical relic sample and the anti-fibroin albumen antibody of rabbit there occurs specific binding, proves that historical relic sample is silk goods.
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| CN201410846331.7A CN104483478B (en) | 2014-12-31 | 2014-12-31 | A kind of detection method of Ancient Silk Textile |
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| CN104483478B true CN104483478B (en) | 2016-01-13 |
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Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105372420A (en) * | 2015-11-12 | 2016-03-02 | 浙江理工大学 | Method for detecting ancient wool fabric by microscope |
| CN105699642A (en) * | 2016-02-19 | 2016-06-22 | 浙江理工大学 | Method for detecting ancient wool fabrics by aid of fluoroimmunoassay |
| CN107057683B (en) * | 2016-12-28 | 2019-04-12 | 浙江大学 | A kind of preparation method of fibroin fluorescence probe |
| CN113030488B (en) * | 2021-04-01 | 2022-04-01 | 浙江理工大学 | Method for detecting silk fibroin by using immunomagnetic surface enhanced Raman substrate |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102590482A (en) * | 2011-12-27 | 2012-07-18 | 苏州大学 | Method for identifying male silk of mulberry silkworm |
| CN103509108A (en) * | 2013-08-07 | 2014-01-15 | 浙江大学 | Method of preparing bombyx mori silk fibroin specific antibody by utilizing characteristic dodecapeptide |
| CN103509107A (en) * | 2013-08-07 | 2014-01-15 | 浙江大学 | Method of preparing bombyx mori silk fibroin specific antibody by utilizing characteristic polypeptide |
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| JPH0572205A (en) * | 1991-09-14 | 1993-03-23 | Kanebo Ltd | Immunoassay and apparatus therefor |
| US9029168B2 (en) * | 2010-06-28 | 2015-05-12 | The Trustees Of Princeton University | Use and making of biosensors utilizing antimicrobial peptides for highly sensitive biological monitoring |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102590482A (en) * | 2011-12-27 | 2012-07-18 | 苏州大学 | Method for identifying male silk of mulberry silkworm |
| CN103509108A (en) * | 2013-08-07 | 2014-01-15 | 浙江大学 | Method of preparing bombyx mori silk fibroin specific antibody by utilizing characteristic dodecapeptide |
| CN103509107A (en) * | 2013-08-07 | 2014-01-15 | 浙江大学 | Method of preparing bombyx mori silk fibroin specific antibody by utilizing characteristic polypeptide |
Non-Patent Citations (1)
| Title |
|---|
| 利用丝素蛋白抗体鉴定古代丝织品;郑秦等;《蚕业科学》;20140615;第40卷(第3期);第520-526页 * |
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