CN105572046B - Fluoroscopic examination sample cell and preparation method thereof - Google Patents
Fluoroscopic examination sample cell and preparation method thereof Download PDFInfo
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- CN105572046B CN105572046B CN201410548940.4A CN201410548940A CN105572046B CN 105572046 B CN105572046 B CN 105572046B CN 201410548940 A CN201410548940 A CN 201410548940A CN 105572046 B CN105572046 B CN 105572046B
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- transparent substrates
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- fluoroscopic examination
- graphene oxide
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Abstract
The present invention provides a kind of fluoroscopic examination sample cell and preparation method thereof, which includes the first transparent substrates；Second transparent substrates, there is metallic film and graphene oxide successively in second transparent substrates, second transparent substrates and the first transparent substrates are bonded together and define the accommodation space for accommodating sample, and the metallic film and graphene oxide are between first transparent substrates and the second transparent substrates；First transparent substrates or the second transparent substrates have the sample holes being connected with the accommodation space, and first transparent substrates or the second transparent substrates have the sample outlet hole being connected with the accommodation space.The fluoroscopic examination sample cell of the present invention can use illumination path and imaging optical path in conventional microscope, be not limited to lambda1-wavelength and fluorescent marker type, at low cost, and longitudinal resolution is nanoscale.
Description
Technical field
The present invention relates to fluorescence detection devices, and in particular to a kind of fluoroscopic examination sample cell and preparation method thereof.
Background technology
Detection technique of fluorescence is one of current most widely used Measurement for Biotechnique.Detection technique of fluorescence is to utilize fluorescence
Determinand is marked indicia means, and quantity variation, movement locus and the light intensity variation of real-time tracing record fluorescence signal
Etc. information.Single molecule fluorescence detection is a kind of overdelicate detection technique developed rapidly nearly ten years, compared to tradition
Analysis and detection technology, since it can study monomolecular dynamic variation, to chemical analysis, nano material analysis and
The development in the fields such as live cell assays produces and is generating far-reaching influence.In order to accurately disclose monomolecular movement
Information and rule, the longitudinal resolution of fluoroscopic examination need to reach nanoscale (within 10nm).
Mainstream detection technique of fluorescence is using fluorescence microscopy determinand, due to the longitudinal resolution of microtechnic at present
Rate is limited by optical diffraction limit, and longitudinal resolution can only achieve 100 rans.In order to improve fluoroscopic examination
Longitudinal resolution, researcher has developed super-resolution fluorescence technology, but its longitudinal resolution can only achieve 10-20nm.Then, it grinds
Study carefully personnel and further have developed fluorescence resonance energy transfer technology, fluorescence signal can be detected and believed in nanoscale distance change
Breath, but the current lambda1-wavelength that can be used in fluorescence resonance energy transfer technology and fluorescent marker type are very limited, it is glimmering
Photoresonance energy transfer technique can not be applied to certain samples with special nature.Therefore, need one kind right at present
All samples carry out the longitudinal resolution of fluoroscopic examination, fluoroscopic examination as nanoscale and can utilize the illumination of conventional microscope
The fluoroscopic examination sample cell of light path and imaging optical path.
Invention content
For above-mentioned technical problem, An embodiment provides a kind of fluoroscopic examination sample cell, including：
First transparent substrates；
Second transparent substrates, have metallic film and graphene oxide in second transparent substrates successively, and described second
Transparent substrates and the first transparent substrates are bonded together and define the accommodation space for accommodating sample, the metallic film and
Graphene oxide is between first transparent substrates and the second transparent substrates；
First transparent substrates or the second transparent substrates have a sample holes being connected with the accommodation space, and described the
One transparent substrates or the second transparent substrates have the sample outlet hole being connected with the accommodation space.
Preferably, the metallic film is gold thin film or Ag films of the thickness no more than 50 nanometers.
Preferably, the fluoroscopic examination sample cell further includes the phospholipid molecule layer on the graphene oxide.
Preferably, the number of plies of the graphene oxide is 1.
Preferably, the edge of first transparent substrates and the edge of the second transparent substrates are bonded together.
Preferably, first transparent substrates are slide or quartz plate, and second transparent substrates are slide or quartz plate.
Preferably, the thickness of first transparent substrates or the second transparent substrates is 0.1~1 millimeter.
One embodiment of the present of invention additionally provides a kind of preparation method for being used to prepare above-mentioned fluoroscopic examination sample cell, packet
Include the following steps：
1) sample holes are formed in first transparent substrates or the second transparent substrates, in first transparent substrates or
Sample outlet hole is formed in two transparent substrates；
2) metallic film and graphene oxide are grown successively in second transparent substrates；
3) first transparent substrates and the second transparent substrates are bonded together so that first transparent substrates and
Two transparent substrates limit the accommodation space for accommodating sample, and it is transparent that the metallic film and graphene oxide are located at described first
Between substrate and the second transparent substrates, the sample holes and sample outlet hole are all connected with the accommodation space.
Preferably, also include growing phospholipid molecule layer on the graphene oxide in the step 2).
Preferably, in the step 2), the growth metallic film is to utilize vacuum evaporation or electron beam evaporation technique
Gold thin film or Ag films of the growth thickness no more than 50 nanometers in second transparent substrates, the growth graphene oxide are
Single-layer graphene oxide is grown using LB membrane technologies.
The fluoroscopic examination sample cell of the present invention can use illumination path and imaging optical path in conventional microscope, be not limited to
Lambda1-wavelength and fluorescent marker type, it is at low cost, and longitudinal resolution is nanoscale.In addition fluoroscopic examination using the present invention
Sample cell can monitor protein transmembrane process.
Description of the drawings
Embodiments of the present invention is further illustrated referring to the drawings, wherein：
Fig. 1 is the sectional view of the fluoroscopic examination sample cell of first embodiment of the invention.
Fig. 2 is the schematic diagram of the fluorescence detection device of present pre-ferred embodiments.
Fig. 3 is the sectional view of the fluoroscopic examination sample cell of second embodiment of the invention.
Fig. 4 is the sectional view of the fluoroscopic examination sample cell of third embodiment of the invention.
Fig. 5 is the sectional view of the fluoroscopic examination sample cell of four embodiment of the invention.
Specific embodiment
In order to make the purpose , technical scheme and advantage of the present invention be clearer, pass through below in conjunction with attached drawing specific real
Applying example, the present invention is described in more detail.It should be appreciated that the specific embodiments described herein are merely illustrative of the present invention,
It is not intended to limit the present invention.
Fig. 1 is the sectional view of the fluoroscopic examination sample cell 10 of first embodiment of the invention.As shown in Figure 1, sample cell 10
Including being in flat diascope 11 and diascope 12, there is 20 nanometers of gold thin film 16 and individual layer oxygen successively on slide 12
Graphite alkene 13 and the phospholipid molecule layer 27 in single-layer graphene oxide 13.Slide 11 and slide 12 are oppositely arranged, and
The edge of slide 11 and slide 12 is bonded together by binder 14 (such as double faced adhesive tape and/or silica gel), 14 thickness of binder
About 100 microns, so as to form one between slide 11 and slide 12 for accommodating the accommodation space 15 of sample to be tested, gold is thin
Film 16 and single-layer graphene oxide 13 are between slide 11 and slide 12.Slide 12 has what is be connected with accommodation space 15 to lead to
Hole 121 and through-hole 122, one of sample holes as sample, another is as sample outlet hole.
Gold thin film 16 and single-layer graphene oxide 13 are generated on slide 12, can cause fluorescent molecular feature be quenched away from
It is 4nm or so from (fluorescent intensity is reduced with a distance from corresponding to half), when the distance between fluorescent molecular and graphene oxide 13
When changing 1nm, significant changes can also occur for fluorescent intensity, therefore longitudinal resolution can reach 1nm.
Fig. 2 is the schematic diagram of the fluorescence detection device of present pre-ferred embodiments.Fluorescence detection device 50 is included shown in Fig. 1
Sample cell 10, illumination path 51 and imaging optical path 52.Wherein illumination path 51 is used to provide total internal reflection illumination field to sample,
Imaging optical path 52 is used to acquire fluorescence and the imaging of sample.Illumination path 51 and imaging optical path 52 are of the prior art complete interior anti-
It penetrates in fluorescence microscope (such as Olympus utilizing total internal reflection fluorescence microscope, Lycra utilizing total internal reflection fluorescence microscope, etc.)
Illumination path and imaging optical path, details are not described herein for specific light path.
Below in conjunction with the application method of Fig. 2 summary fluorescence detection devices 50.First, it will graft or be modified with fluorescent molecular
Protein molecular (sample) be injected into accommodation space 15 by through-hole 121 or 122, open illumination path 51 provided to sample it is complete
Internal reflection illuminated field, fluorescent molecular be stimulated after at once de excitation send out and send out fluorescence, imaging optical path 52 acquires the fluorescence that sends out, and
Measure fluorescent intensity.Efficiency is quenched to fluorescence based on graphene oxide：When between fluorescent molecular and single-layer graphene oxide
When distance changes, fluorescent intensity can also change, and fluorescent molecular is nearer apart from single-layer graphene oxide, and fluorescent intensity is got over
Weak, fluorescent molecular is more remote apart from single-layer graphene oxide, and fluorescent intensity is stronger.Therefore the reacting condition of fluorescent intensity goes out fluorescence point
The variation of distance between son and single-layer graphene oxide.Since the longitudinal resolution of sample cell 10 is 1nm, as protein molecular exists
During phospholipid molecule layer 27 (thickness is about 4nm), the fluorescent intensity of measurement changes, and can be according to glimmering
The variation of light light intensity reflects lengthwise position of the protein molecular in phospholipid molecule 27, that is, realizes chasing after in real time for protein transmembrane
Track, so as to solve the technical barrier that people want monitoring protein transmembrane process in real time always.In addition fluoroscopic examination of the invention
Sample cell can use illumination path and imaging optical path in conventional microscope, be not limited to lambda1-wavelength and fluorescent marker kind
Class, it is at low cost.
The preparation method of fluoroscopic examination sample cell 10 shown in FIG. 1 is summarized below, 121 He of through-hole is formed on slide 12
Through-hole 122 is thin in gold using the gold thin film 16 of one layer 20 nanometers of vacuum evaporation or electron beam evaporation technique growth on slide 12
Single-layer graphene oxide 13 is grown using langmuir-blodgett technologies (LB membrane technologies) on film 16, later in graphite oxide
Phospholipid molecule layer 27 is grown on alkene 13, is finally sticked the edge at the edge of slide 11 and slide 12 using double faced adhesive tape and/or silica gel
It ties together, so that slide 11 and slide 12 define the accommodation space 15 for accommodating sample.Those skilled in the art
Member understands to grow using gold thin film of the existing technological parameter growth thickness no more than 50 nanometers, using LB membrane technologies mono-layer oxidized
Graphene simultaneously grows Lipid monolayer in single-layer graphene oxide, and details are not described herein for specific process parameter.
Fig. 3 is the sectional view of the fluoroscopic examination sample cell 20 of second embodiment of the invention.It is essentially identical with Fig. 1, area
It is not do not have phospholipid molecule layer 27 in single-layer graphene oxide 13.
Fig. 4 is the sectional view of the fluoroscopic examination sample cell 30 of third embodiment of the invention.It is essentially identical with Fig. 3, area
It is not that the thickness of the slide 32 of sample cell 30 is suitable with slide 11, the thickness of slide 31 is suitable with the thickness of slide 12, through-hole
311st, it 312 is arranged on slide 31.
Fig. 5 is the sectional view of the fluoroscopic examination sample cell 40 of four embodiment of the invention.It is essentially identical with Fig. 4, area
It is not that in single-layer graphene oxide 13 also there is phospholipid molecule layer 47.
In other embodiments of the invention, the through-hole 121 in Fig. 1 and Fig. 3 and/or through-hole 122 can be arranged on slide
On 11.
In other embodiments of the invention, the through-hole 311 in Fig. 4 and Fig. 5 and/or through-hole 312 can be arranged on slide
On 32.
In other embodiments of the invention, quartz plate may be used or transparent organic glass is replaced in above-described embodiment
Slide 11, slide 12, slide 31 and slide 32.
In other embodiments of the invention, the thickness of gold thin film can be no more than any number of 50nm.In this hair
In bright other embodiment, Ag films of the thickness no more than 50nm may be used instead of the gold thin film 16 in above-described embodiment.
In other embodiments of the invention, the number of plies of graphene oxide can be 2 or 3 layers.
For the ease of the acquisition fluorescence (shown in Figure 2) of imaging optical path 52, the thickness of slide 11,32 is preferably smaller than 1 millimeter,
More preferably 0.1~0.2 millimeter.
Although the present invention has been described by means of preferred embodiments, the present invention is not limited to described here
Embodiment, further include made various changes and variation without departing from the present invention.
Claims (10)
1. a kind of fluoroscopic examination sample cell, which is characterized in that including：
First transparent substrates；
Second transparent substrates have metallic film and graphene oxide in second transparent substrates successively, and described second is transparent
Substrate and the first transparent substrates are bonded together and define the accommodation space for accommodating sample, the metallic film and oxidation
Graphene is between first transparent substrates and the second transparent substrates；
First transparent substrates have the sample holes being connected with the accommodation space and sample outlet hole or the second transparent lining
Bottom has the sample holes being connected with the accommodation space and sample outlet hole.
2. fluoroscopic examination sample cell according to claim 1, which is characterized in that the metallic film is that thickness is not more than 50
The gold thin film or Ag films of nanometer.
3. fluoroscopic examination sample cell according to claim 1 or 2, which is characterized in that the fluoroscopic examination sample cell also wraps
Include the phospholipid molecule layer on the graphene oxide.
4. fluoroscopic examination sample cell according to claim 1 or 2, which is characterized in that the number of plies of the graphene oxide is
1。
5. fluoroscopic examination sample cell according to claim 1 or 2, which is characterized in that the edge of first transparent substrates
It is bonded together with the edge of the second transparent substrates.
6. fluoroscopic examination sample cell according to claim 1 or 2, which is characterized in that first transparent substrates are slide
Or quartz plate, second transparent substrates are slide or quartz plate.
7. fluoroscopic examination sample cell according to claim 6, which is characterized in that first transparent substrates are second transparent
The thickness of substrate is 0.1~1 millimeter.
8. a kind of preparation method of fluoroscopic examination sample cell being used to prepare described in any one of claim 1 to 7, feature exist
In including the following steps：
1) sample holes and sample outlet hole are formed in first transparent substrates or forms sample holes in second transparent substrates
And sample outlet hole；
2) metallic film and graphene oxide are grown successively in second transparent substrates；
3) first transparent substrates and the second transparent substrates are bonded together so that first transparent substrates and second are thoroughly
Bright substrate limits the accommodation space for accommodating sample, and the metallic film and graphene oxide are located at first transparent substrates
And second between transparent substrates, the sample holes and sample outlet hole are all connected with the accommodation space.
9. preparation method according to claim 8, which is characterized in that the oxidation stone is additionally included in the step 2)
Phospholipid molecule layer is grown on black alkene.
10. preparation method according to claim 8 or claim 9, which is characterized in that in the step 2), the growth metal
Using vacuum evaporation or electron beam evaporation technique, the growth thickness in second transparent substrates is not more than 50 nanometers to film
Gold thin film or Ag films, the growth graphene oxide is to grow single-layer graphene oxide using LB membrane technologies.
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| CN108562544B (en) * | 2018-04-17 | 2020-11-03 | 东南大学 | Stretchable flexible liquid pool and manufacturing method thereof |
| CN109900667B (en) * | 2019-03-15 | 2021-08-06 | 电子科技大学 | All-fiber laser type selective hypersensitivity biochemical sensor |
| CN112051252A (en) * | 2020-09-16 | 2020-12-08 | 中国科学院长春应用化学研究所 | Sample cell and preparation method and application thereof |
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| CN101042333A (en) * | 2006-03-20 | 2007-09-26 | 宜兴市晶科光学仪器有限公司 | Colorimetric ware and preparation method thereof |
| CN101251470A (en) * | 2007-12-25 | 2008-08-27 | 中国科学院力学研究所 | Sample pool for optical microscope to view protein crystal etch |
| CN102692392A (en) * | 2011-03-25 | 2012-09-26 | 上海光刻电子科技有限公司 | Device for measuring gas and liquid refractive indexes |
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