CN104090098A - Ultrasensitive immune detection technology based on silica nanoparticle enhancement - Google Patents
Ultrasensitive immune detection technology based on silica nanoparticle enhancement Download PDFInfo
- Publication number
- CN104090098A CN104090098A CN201410261349.0A CN201410261349A CN104090098A CN 104090098 A CN104090098 A CN 104090098A CN 201410261349 A CN201410261349 A CN 201410261349A CN 104090098 A CN104090098 A CN 104090098A
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- China
- Prior art keywords
- silicon dioxide
- nano particles
- flat based
- phase carrier
- tubes
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Classifications
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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
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
- G01N33/54346—Nanoparticles
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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
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/551—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being inorganic
- G01N33/552—Glass or silica
Abstract
The invention discloses a biological molecule detection method based on silica nanoparticles as a solid-phase carrier, and the method is characterized in that: by hydrophilic adsorption or chemical action, a flat tube can uniformly absorb silica nanoparticles on the bottom of the flat tube, so that the bottom of the flat tube shows color for distinguishing different detection objects, after treatment of the silica nanoparticles as the solid-phase carrier, the adsorption capacity is enhanced, the surface is uneven, the specific surface area is increased for connecting more probes, and the detection sensitivity is significantly improved, and the detection time is saved.
Description
Technical field
What the present invention relates to is a kind of water wettability flat based tubes based on Nano particles of silicon dioxide.After certain processing, strengthen adsorptive power, and its surface irregularity, can increase specific surface area, connect more probe, and then significantly improve detection sensitivity, can be widely used in the field such as clinical detection, inspection and quarantine, environmental monitoring, food safety detection, drug screening, microbial identification and nucleic acid and protein function analysis.
Background technology
Sweden scholar Engvail in 1971 and Perlmann, Holland scholar Van Weerman and Schuurs report respectively immunological technique are developed into the solid-phase immunoassay method that detects micro substance in body fluid, be enzyme-linked immunosorbent assay (Enzyme-Linked ImmunoSorbent Assay, ELISA).ELISA has become the advanced subject in present analysis chemical field now, it is a kind of special reagent analysis method, is a kind of novel immunoassay growing up on the basis of immunoenzyme technics (immunoenzymatic techniques).At present, the ultimate principle of Enzyme-linked Immunosorbent Assay reaction is to make antigen or antibody be attached to certain surface of solid phase carriers, and keeps its immunocompetence.Make antigen or antibody and certain enzyme connect into enzyme-labelled antigen or antibody, this enzyme-labelled antigen or antibody had both retained its immunocompetence, retained again the activity of enzyme.In the time measuring, reacted by the antigen of different steps and surface of solid phase carriers or antibody examining sample (measuring antibody or antigen wherein) and enzyme-labelled antigen or antibody.With washing method the antigen antibody complex forming on solid phase carrier is separated with other materials, the enzyme amount being finally combined on solid phase carrier becomes certain ratio with the amount of tested substance in sample.Add after the substrate of enzyme reaction, substrate is become coloured product by enzymatic, and the amount of product is directly related with the amount of tested substance in sample, therefore can carry out qualitative or quantitative test according to the depth of color reaction.Because the catalysis frequency of enzyme is very high, thus iodine effect greatly, thus make assay method reach very high susceptibility.Its principle of the present invention is to adhere to layer of silicon dioxide nano particle in flat based tubes, connect antigen or antibody, taking immunological response as basis, the very high experimental technique of a kind of susceptibility that the specific reaction of antigen, antibody and enzyme are combined together the efficient catalytic effect of substrate.Taking Nano particles of silicon dioxide as the medium as absorption, after certain processing, strengthen adsorptive power, and its surface irregularity, can increase specific surface area, greatly improve the antigen of absorption or the amount of antibody, and then significantly improve detection sensitivity.Hydrophilic flat based tubes taking Nano particles of silicon dioxide as solid phase carrier himself with color, be beneficial to the differentiation of object! This flat based tubes inside can embed a mesh device, for dividing different regions, adheres to test tube bottom with different Nano particles of silicon dioxide, shows different colors, reaches the object that simultaneously detects multiple objects.
Summary of the invention
technical matters:
In order further to improve the sensitivity of test experience on ELISA experiment basis, cost-saving, improve experimental program.The invention provides the hydrophilic flat based tubes of having layer of silicon dioxide nano particle in a kind of bottom for Biological Detection.Nano particles of silicon dioxide is solid phase carrier, after certain processing, strengthens adsorptive power, and its surface irregularity, can increase specific surface area, connects more probe, and then significantly improves detection sensitivity, saves detection time.
technical scheme:
The present invention is a kind of hydrophilic flat based tubes, its bottom layer of silicon dioxide nano particle that tiled, as shown in Figure 2.After treatment, its surface irregularity, can increase specific surface area to Nano particles of silicon dioxide, connects more probe, and then significantly improves detection sensitivity, saves detection time.
beneficial effect:according to the present invention, the flat based tubes bottom layer of silicon dioxide nano particle that tiled is had the following advantages for biological detection:
(1) strengthen adsorptive power: test tube Bottomattached Nano particles of silicon dioxide, uneven, can increase specific surface area, connect more probe, and then significantly improve detection sensitivity.And seem more flexible with respect to its single self-existent feature of common 96 orifice plates, convenient.(2) this flat based tubes inside can embed a mesh device, for dividing different regions, adheres to test tube bottom with different Nano particles of silicon dioxide, shows different colors, reaches the object that simultaneously detects multiple objects.(3) save reagent: while washing plate, the present invention only needs 30 μ L solution, thereby save reagent, reach the object of controlling cost.(4) be beneficial to and distinguish different target thing: this flat based tubes self can be with color, according to the size of the Nano particles of silicon dioxide of tiling need not, this flat based tubes can show different colors, can be beneficial to its color carrying and distinguish object.(5) collocation flexibly: this flat based tubes different colours represents different detection objects, and tester can, according to different testing requirements, select different colors, flexibly collocation.
Brief description of the drawings
Fig. 1 is flat based tubes schematic diagram.
Fig. 2 is the tiled flat based tubes schematic diagram of Nano particles of silicon dioxide of bottom.
embodiment
The present invention is a kind of hydrophilic flat based tubes, its bottom layer of silicon dioxide nano particle that tiled, as shown in Figure 2.Nano particles of silicon dioxide after treatment, strengthens adsorptive power, and its surface irregularity, can increase specific surface area, connects more probe, and then significantly improves detection sensitivity, saves detection time.
Embodiment mono-:
Taking indirect immunofluorescence as example, use the present invention to detect tumor marker alpha-fetoprotein (AFP) and hepatitis B simultaneously:
1, dilute respectively tumor marker alpha-fetoprotein (AFP) antigen and antigen of hepatitis B virus with coated damping fluid, add bottom with in the flat based tubes of Nano particles of silicon dioxide, control group is set simultaneously, and control group can be selected the serum that does not contain tumor marker alpha-fetoprotein (AFP) antigen and antigen of hepatitis B virus.2,37 DEG C of water bath with thermostatic control 2h, remove in vitro liquid, and test tube is tipped upside down on thieving paper, to remove as much as possible in vitro original liquid; 3, TBST washes three times, and each 10 minutes, after last cleaning, fall dry invisible spectro solution, add confining liquid, 37 DEG C of water bath with thermostatic control 2h, or 4 DEG C spent the night; 4, remove in vitro liquid, and test tube is tipped upside down on thieving paper, to remove as much as possible in vitro original liquid; TBST washes three times, and each 10 minutes, after last cleaning, fall to do invisible spectro solution, test serum is diluted with sample diluting liquid, add respectively in flat based tubes.Cover lid, hatches 1 hour for 37 DEG C, or 4 DEG C are spent the night.5, TBST washes three times, and each 10 minutes, after last cleaning, fall to do invisible spectro solution, add respectively two to resist, hatch 1 hour for 37 DEG C, or 4 DEG C are spent the night.6, TBST washes three times, each 10 minutes, after last cleaning, fall to do invisible spectro solution, pat dry, add respectively substrate nitrite ion, color development at room temperature 10-15 minute, adds stop buffer, reacts 10-30 minutes, under 450nm wavelength, measure absorbance by microplate reader, also can show result by mark fluorescent.
Embodiment bis-:
Taking indirect immunofluorescence as example, use the present invention to detect the adjuvant melamine in milk:
1, with coated damping fluid dilution testing sample, add bottom with in the flat based tubes of Nano particles of silicon dioxide, control group is set simultaneously, control group can be selected the blank dilution that does not contain melamine.2,37 DEG C of water bath with thermostatic control 2h, remove in vitro liquid, and test tube is tipped upside down on thieving paper, to remove as much as possible in vitro original liquid; 3, TBST washes three times, and each 10 minutes, after last cleaning, fall dry invisible spectro solution, add confining liquid, 37 DEG C of water bath with thermostatic control 2h, or 4 DEG C spent the night; 4, remove in vitro liquid, and test tube is tipped upside down on thieving paper, to remove as much as possible in vitro original liquid; TBST washes three times, and each 10 minutes, after last cleaning, fall to do invisible spectro solution, by melamine antibody diluted, add respectively in flat based tubes.Cover lid, hatches 1 hour for 37 DEG C, or 4 DEG C are spent the night.5, TBST washes three times, and each 10 minutes, after last cleaning, fall to do invisible spectro solution, add respectively two to resist, hatch 1 hour for 37 DEG C, or 4 DEG C are spent the night.6, TBST washes three times, each 10 minutes, after last cleaning, fall to do invisible spectro solution, pat dry, add respectively substrate nitrite ion, color development at room temperature 10-15 minute, adds stop buffer, reacts 10-30 minutes, under 450nm wavelength, measure absorbance by microplate reader, also can show result by mark fluorescent.
Claims (9)
1. one kind is the method that solid phase carrier carries out biomolecule detection based on Nano particles of silicon dioxide.
2. it is characterized in that: flat based tubes can be attached to Nano particles of silicon dioxide uniformly by hydrophilic suction-operated, physical action or chemical bond the bottom of flat based tubes.
According to claim 1 a kind of be the water wettability flat based tubes that solid phase carrier carries out biomolecule detection based on Nano particles of silicon dioxide, it is characterized in that: layer of silicon dioxide nano particle has been enclosed in the bottom of flat based tubes.
4. this nano particle makes flat based tubes bottom present colour, can be used for distinguishing different detection objects.
According to claim 1 a kind of be the water wettability flat based tubes that solid phase carrier carries out biomolecule detection based on Nano particles of silicon dioxide, it is characterized in that: Nano particles of silicon dioxide is solid phase carrier, after certain processing, strengthen adsorptive power, and its surface irregularity, can increase specific surface area, connects more probe, and then significantly improve detection sensitivity, save detection time.
According to claim 1 a kind of be the water wettability flat based tubes that solid phase carrier carries out biomolecule detection based on Nano particles of silicon dioxide, it is characterized in that: the Nano particles of silicon dioxide surface being mounted with in flat based tubes can fixed nucleic acid or protein, for Biological Detection.
According to claim 1 a kind of be the water wettability flat based tubes that solid phase carrier carries out biomolecule detection based on Nano particles of silicon dioxide, it is characterized in that: this flat based tubes only needs 30 μ L reagent, thereby saving reagent, reaches the object of controlling cost.
According to claim 1 a kind of be the water wettability flat based tubes that solid phase carrier carries out biomolecule detection based on Nano particles of silicon dioxide, it is characterized in that: this flat based tubes different colours represents different detection objects, tester can be according to different testing requirements, select different colors, flexibly collocation.
According to claim 1 a kind of be the water wettability flat based tubes that solid phase carrier carries out biomolecule detection based on Nano particles of silicon dioxide, it is characterized in that: this flat based tubes inside can embed a mesh device, for dividing different regions, adhere to test tube bottom with different Nano particles of silicon dioxide, show different colors, reach the order ground that simultaneously detects multiple objects.
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| CN201410261349.0A CN104090098A (en) | 2014-06-13 | 2014-06-13 | Ultrasensitive immune detection technology based on silica nanoparticle enhancement |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105527431A (en) * | 2015-04-04 | 2016-04-27 | 吉林双正医疗科技有限公司 | Group B hemolytic streptococcus IgG antibody rapid-detection equipment and preparation method thereof |
-
2014
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105527431A (en) * | 2015-04-04 | 2016-04-27 | 吉林双正医疗科技有限公司 | Group B hemolytic streptococcus IgG antibody rapid-detection equipment and preparation method thereof |
| CN105548538A (en) * | 2015-04-04 | 2016-05-04 | 吉林双正医疗科技有限公司 | IgM antibody combined detection device of Coxsackie virus A16 and enterovirus 71 and making method thereof |
| CN105548534A (en) * | 2015-04-04 | 2016-05-04 | 吉林双正医疗科技有限公司 | Cardiac marker combined detection device and preparation method thereof |
| CN105588939A (en) * | 2015-04-04 | 2016-05-18 | 吉林双正医疗科技有限公司 | MPO, H-FABP and cTnT combined detection device and making method |
| CN105588939B (en) * | 2015-04-04 | 2017-07-07 | 吉林双正医疗科技有限公司 | MPO, H FABP and cTnT joint-detections device and preparation method |
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Application publication date: 20141008 |
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