CN106153538A - A kind of addressable sample panel and the application in unicellular micro-imaging thereof - Google Patents
A kind of addressable sample panel and the application in unicellular micro-imaging thereof Download PDFInfo
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- CN106153538A CN106153538A CN201510186211.3A CN201510186211A CN106153538A CN 106153538 A CN106153538 A CN 106153538A CN 201510186211 A CN201510186211 A CN 201510186211A CN 106153538 A CN106153538 A CN 106153538A
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Abstract
The invention belongs to micro-imaging field, disclose a kind of addressable sample panel and the application in unicellular micro-imaging of this sample panel.Being carved with the array of N number of n × m in the sample panel of the present invention, the cell size in described array is 100 μm2-4×104μm2.The invention also discloses the application in unicellular micro-imaging of the described sample panel and the method for unicellular micro-imaging.The method of described unicellular micro-imaging includes being fixed in described sample panel cell, is marked with fluorescent dye, and the cell in same cell carries out fluorescence imaging analysis and mass spectrum imaging analysis respectively.When the addressable sample panel of the present invention is applied to unicellular microscopic image analysis, be conducive to accurately clearly positioning same cell in wide area, when reducing the combination of multiple analytical tool, position single celled difficulty.Additionally, the sample panel of the present invention will not hinder the adherent of cell and growth, produce harmful effect thus without to analysis result.
Description
Technical field
The invention belongs to micro-imaging field, in particular it relates to a kind of addressable sample panel and this sample
The application in unicellular micro-imaging of the product plate.
Background technology
Up to now, people are the deepest to the study mechanism of metal antitumor drug suppression tumor cell
Enter.And the medicine of research can not react thin with the interaction mechanism of bio-target molecule on molecular level
The truth of intracellular.Therefore medicine understands antineoplastic agent in the research of intracellular spatial distribution to deep
The mechanism of action of thing, the MOLECULE DESIGN optimizing them further has very important significance, and this is also medicine
One of the most challenging problem that thing research and development field is faced.
At present, drugs has fluorescent labeling imaging method and mass spectrum imaging in the main method of intracellular distribution
Method.It is good that fluorescent marker method has specificity, and fluorescence efficiency is high, the advantages such as sample consumption is little.But simultaneously
Require that medicine itself is containing launching the group of fluorescence at a particular wavelength or needing to add on drug molecule
Add fluorophor；And pharmaceutical synthesis process so can be made to become more sophisticated, the fluorophor of interpolation is likely
The biological activity of medicine can be changed, in the complex environment of cell, be susceptible to fluorescent quenching etc..Secondary from
Sub-mass spectrography is highly sensitive, spatial resolution is high, and can obtain material in a data acquisition
Chemical information and space distribution information.But cellular environment is complicated, basal signal is strong, is sometimes difficult to find that
Specific mass-fragments.Cell carries out fluorescence pattern imaging analysis respectively (need not labeled drug divide
Son) and optical morphology imaging and chemical composition imaging can be realized for the mass spectrum imaging analysis of drug molecule
Directly perceived corresponding, more clearly understand medicine distribution in cell, will be able to be the work of further investigation medicine
Thering is provided effective experimental evidence by mechanism, the research to the metabolic process of intracellular complexity has critically important
Meaning.
But, when carrying out fluorescence pattern imaging and mass spectrum composition imaging combination analysis, typically it is difficulty with
To microcell same under nanoscale, the most same unicellular it is analyzed, causes morphology analysis and composition
Analyze result can not one_to_one corresponding, weaken the scientific value of combination analysis.
Summary of the invention
Exist when it is an object of the invention to overcome current pattern imaging and composition imaging combination analysis is slender
The technical barrier of born of the same parents' location difficulty, it is provided that one can find designated cell quickly and easily in large area
Addressable sample panel and the application in unicellular micro-imaging of this sample panel.
To achieve these goals, first aspect, the invention provides a kind of addressable sample panel, should
Being carved with the array of N number of n × m in sample panel, the size of the cell in described array is 100 μm2-
4×104μm2。
Second aspect, a kind of method that the invention provides unicellular micro-imaging, the method includes: will
Cell is fixed in the sample panel described in first aspect, is marked with fluorescent dye, by same cell
In cell carry out fluorescence imaging analysis and mass spectrum imaging analysis respectively.
The third aspect, the invention provides the sample panel described in first aspect in unicellular micro-imaging
Application.
When addressable sample panel of the present invention is applied to unicellular microscopic image analysis, be conducive to greatly
Accurately clearly positioning same cell in areal extent, when reducing the combination of multiple analytical tool, location is single
The difficulty of cell.Additionally, the sample panel of the present invention will not hinder the adherent of cell and growth, thus without
Analysis result is produced harmful effect.
Other features and advantages of the present invention will be described in detail in detailed description of the invention part subsequently.
Accompanying drawing explanation
Accompanying drawing is used to provide a further understanding of the present invention, and constitutes the part of description, with
Detailed description below is used for explaining the present invention together, but is not intended that limitation of the present invention.?
In accompanying drawing:
Fig. 1 is for being prepared Following the procedure of Example 1 addressable silicon chip sample plate laser scanning co-focusing
Picture (the figure of the video camera shooting on the picture (Figure 1A) of microscope photographing and ion microprobe
1B)；
Fig. 2 is ruthenium compound 1 (Fig. 2 A) used by embodiment and the chemistry of ruthenium compound 2 (Fig. 2 B)
Structure chart；
Fig. 3 is the ruthenium compound 2 that in embodiment 3, secondary ion mass spectrum (SIMS) imaging analysis detects
(m/z 598, C26H28ClFN5O2Ru+) standard sample and in the mass spectrum (Fig. 3 A) of surface of cell membrane
With ruthenium element in nucleus (m/z 102,102Ru+) mass spectrum (Fig. 3 B)；
Fig. 4 is that to be prepared Following the procedure of Example 2 the MCF-7 that in sample panel, compound 1 is hatched thin
Born of the same parents' cell membrane red fluorescence dyestuff DiI fluorescent staining figure, m/z 184 (C5H15NPO4 +, cell membrane component
Phosphatidylcholine molecules fragment) spatial distribution map, m/z 240 (C8H14N2Ru+) spatial distribution map and three
The stacking chart of person；Nucleus blue fluorescent dyes Hoechst33342 fluorescent staining figure, m/z 81
(C5H5O+, deoxyribose molecule fragment) spatial distribution map, m/z 102 (ruthenium element,102Ru+) empty
Between scattergram and the stacking chart of three；
Fig. 5 hatches MCF-7 cell for being prepared Following the procedure of Example 3 compound 2 in sample panel
Cell membrane red fluorescence dyestuff DiI fluorescent staining figure, m/z 184 (C5H15NPO4 +, cell membrane component phosphorus
Phosphatidylcholine molecule fragment) spatial distribution map, m/z 598 (C26H28ClFN5O2Ru+) spatial distribution map
Stacking chart with three；Nucleus blue fluorescent dyes Hoechst33342 fluorescent staining figure, m/z 81
(C5H5O+, deoxyribose molecule fragment) spatial distribution map, m/z 102 (ruthenium element,102Ru+) space
Scattergram and the stacking chart of three；
Fig. 6 is to be prepared Following the procedure of Example 5 in sample panel the HeLa cell that compound 1 is hatched
Microscope light field figure, m/z 184 (C5H15NPO4 +, cell membrane component phosphatidylcholine molecules fragment) and empty
Between scattergram and both stacking charts；Nucleus blue fluorescent dyes Hoechst33342 fluorescent staining figure,
m/z 81(C5H5O+, deoxyribose molecule fragment) and spatial distribution map and both stacking charts；
Fig. 7 is the sample panel schematic diagram according to one embodiment of the present invention.
Description of reference numerals
U cell
Detailed description of the invention
Hereinafter the detailed description of the invention of the present invention is described in detail.It should be appreciated that this place is retouched
The detailed description of the invention stated is merely to illustrate and explains the present invention, is not limited to the present invention.
As it is shown in fig. 7, be carved with the array of N number of n × m in addressable sample panel of present invention offer,
The size of the cell u in described array is 100 μm2-4×104μm2。
Preferably, N=1-10 and each array respective markers have different numbering (such as, A, B, C,
D ... or I, II, III, IV ...).Spacing between adjacent array is had no particular limits, Ke Yiwei
0.3-1mm。
In a preferred embodiment of the invention, n=4-20, m=4-20 and each cell u are to having
Different numberings (as it is shown in fig. 7, the numbering of cell can be arranged on the side of each array).n
Can be identical or different integer with m, but preferably n=m.
According to the preferred embodiment of the present invention, described cell u is a size of 10 × 10 μm2-
200×200μm2Square and/or the circle of a diameter of 10-200 μm, the most described square.
To the not particularly requirement of the spacing between described cell u, it can be 0-100 μm.
In the present invention, the size of described sample panel can select according to the analysis that reality is carried out, excellent
Elect 0.5 × 0.5cm as2-3×3cm2.The thickness of described sample panel is had no particular limits, as long as meeting
The requirement of microscope imaging, such as, for light transmissive material (such as glass), thickness is generally less than 170
μm；And for light-proof material (such as silicon chip), thickness not requirement.
In the present invention, in order to be further ensured that the most adherent of cell and growth, the quarter in described sample panel
The width of line is preferably 1-10 μm, and the degree of depth is preferably 1-10 μm.Width and the degree of depth can be identical or not
With.
In the present invention, described sample panel can use material commonly used in the art to prepare, preferable case
Under, the material of described sample panel is monocrystal silicon or glass.
A kind of particularly preferably embodiment, 1cm according to the present invention2Silicon chip sample plate on be carved with 4
The array of 4 × 4, wherein the size of cell is 200 × 200 μm2(square).
The sample panel of the present invention can write out figure in version by EBL electron beam lithography, corrodes chromium
Film makes chromium masterplate.Carry out uv-exposure with chromium masterplate again, make the photoresist mask of figure.Finally
Sample with a scale is obtained after substrate (such as monocrystal silicon or glass) being carried out ICP etching under the protection of mask
Product plate.The concrete operations of photoengraving and ICP etching are known to those skilled in the art, the most superfluous at this
State.
The method of the unicellular micro-imaging that the present invention provides includes: cell is fixed on above-mentioned sample panel
On, be marked with fluorescent dye, the cell in same cell is carried out respectively fluorescence imaging analysis and
Mass spectrum imaging is analyzed.Described fluorescence imaging analysis can be conventional fluorescence pattern imaging analysis, is preferably
Laser scanning co-focusing fluorescence imaging analysis.Described mass spectrum imaging analysis can be that conventional mass spectrum becomes to be divided into
As analyzing, preferably secondary ion mass spectrum imaging analysis.
According to the present invention, to the not particularly requirement of cell density in sample panel, it is preferable that cell
Density in described sample panel is preferably 104-105Individual cell/cm2。
According to the present invention, to the not particularly requirement of described cell, can be various types of normal cell
Or tumor cell (general diameter is less than 100 μm).Preferably, described cell is that diameter is in 20-80 μm
In the range of cell.
According to the present invention, the method for fixing cell can be: by by the cell after trypsinization and sample panel
Contact, quiescent culture a period of time can (temperature be 37 DEG C, 5%CO2, the time is 24h).
According to the present invention, described fluorescent dye is had no particular limits, can be can indicate that cell and
The fluorescent dye of subcellular organelle pattern.Preferably, described fluorescent dye is can specificity instruction subcellular fraction
Device or the fluorescent dye of intracellular matter.It is highly preferred that described fluorescent dye is cell membrane red fluorescence dye
Material DiI and nucleus blue fluorescent dyes Hoechst 33342.
According to the present invention, the selection to described secondary ion mass spectrum imaging analysis intermediate ion is the most particularly wanted
Ask, as long as subcellular organelle or intracellular matter can be indicated characteristically, under preferable case, described
Secondary ion is m/z 184 (C5H15NPO4 +, cell membrane component phosphatidylcholine molecules fragment) and m/z
81(C5H5O+, deoxyribose molecule fragment).
Operating for convenience, the sample after the method for the present invention can also include labelling carries out freezing dry
Dry.Liquid nitrogen flash freezer can be used, make distribution during intracellular matter holding living cells state, then pass through
Freezer dryer removes moisture removal.
In the present invention, described fluorescence imaging analysis and mass spectrum imaging analysis are (such as laser scanning co-focusing fluorescence
Imaging analysis and secondary ion mass spectrum imaging analysis) concrete operation step be those skilled in the art public affairs
Know, do not repeat them here.
Present invention also offers the application in unicellular micro-imaging of the sample panel of the invention described above.
Hereinafter will be described the present invention by embodiment.
In following example, human breast cancer cell (MCF-7) and human cervical carcinoma cell (HeLa) are purchased from
BJ Union Hospital's cellular resources center.Cultivate MCF-7 cell and the group of HeLa cell used medium
Cheng Shi: DMEM (high sugar) is added antibiotic (penicillin of 100U/mL and 100mg/mL's
Streptomycin) and the hyclone (FBS) of 10%, wherein, DMEM and FBS is purchased from HyClone
Company；NH4COOCH3Buffer 150mM (pH 7.4)；The pancreatin that peptic cell uses is (containing 0.25%
EDTA) purchased from Gibco company, its article No. is 15050-065；Silicon chip (1 × the 1cm of two-sided polishing2)
Purchased from KYKY Technology Co., Ltd., on silicon chip, figure masterplate is by EBL electron beam lithography
(Germany SUSS-MA6) photoetching forms, and ICP silicon etching uses the ICP180 of England Oxford company；Carefully
After birth red fluorescence dyestuff DiI is purchased from green skies biotechnology research institute；Nucleus blue fluorescent dyes
Hoechst33342 is purchased from Sigma company.
Embodiment 1
The present embodiment is used for illustrating preparation and the sign of addressable sample panel of the present invention.
(1) preparation of chromium masterplate
With EBL electron beam lithography write out figure on plate as shown in Figure 7 (A, B, C, D, 1,
2,3,4,4 × 4 arrays, the size of each cell is 50 × 50 μm2), corrosion chromium film makes chromium masterplate.
(2) preparation of sample panel
The masterplate obtained by above-mentioned steps (1) carries out uv-exposure, makes the photoresist mask of figure.
To 1 × 1cm under the protection of mask2The silicon chip of two-sided polishing carries out ICP silicon etching, required for etching
Width and the degree of depth are 3 μm.With 5mL acetone ultrasonic cleaning three times, nitrogen dries up and obtains sample panel.
(3) sign of sample panel
Observable sample panel under microscope (× 100), image is essentially identical with Fig. 7.
Embodiment 2
The present embodiment is used for illustrating that the sample panel that embodiment 1 prepares is combined imaging for SIMS-Confocal
The method that research ruthenium compound 1 (structural formula is as shown in Figure 2 A) is distributed at MCF-7 intercellular spaces.
(1) cultivation of MCF-7 cell and lyophilizing
Sample panel is placed in the Tissue Culture Dish bottom center of laser co-focusing, the MCF-7 digested
Cell is with 104Individual cell/cm2Concentration cultivates 24h in culture dish, adds containing 100 μMs of ruthenium compounds
Hatch 24h after the culture medium of 1 again, remove culture medium, with the Hoechst's 33342 containing 1 μ g/mL
Cultivate and hatch 10min based in 37 DEG C of incubators.Remove this culture medium, then with containing 5 μMs of cell membrane
The cultivation of red fluorescence dyestuff hatches 20min based in 37 DEG C of incubators.Remove culture medium, cell warp
Concentration is the NH of 150mM4COOCH3(pH 7.4) buffer solution three times, each 30s.By sample
Product plate is quickly put in liquid nitrogen freezing, is then transferred in freezer dryer (-60 DEG C) lyophilization 12h,
And it is gradually increased to room temperature, obtain measuring samples plate.
(2) laser scanning co-focusing fluorescence imaging analysis
The cell attachment of the measuring samples plate of preparation is faced down, is placed in laser co-focusing Tissue Culture Dish
In, this culture dish is moved on inverted fluorescence microscope (FV1000-IX81, OLYMPUS), use
The reflection light of mercury lamp or Hoechst 33342 exciting light 405nm find the microscale labelling in sample panel and
Unicellular in respective markers region.Switch to fluorescence imaging mode to selection area (C23) cell
In unicellular carry out laser scanning co-focusing fluorescence imaging analysis, obtain this cell at cell membrane and cell
The fluorescence imaging figure in core region.
(3) secondary ion mass spectrum imaging analysis
Measuring samples plate is placed in the sample room of ion microprobe, uses built-in taking the photograph on mass spectrometer
It is same unicellular, at following experiment condition that camera finds in this sample panel in specific region (C23)
The secondary ion mass spectrum information of lower this cell of collection: cell surface imaging, 30keV Bi3 +Primary ions bundle,
Emission current 0.8 μ A, sampling area is 60 × 60 μm2, cation imaging pattern, m/z 184
(C5H15NPO4 +, phosphatidylcholine fragment) as indicator cells film distribution mark fragment, m/z 240
(C8H14N2Ru+) it is the mark fragment of ruthenium compound 1.Mass spectrum is by CH+、CH3 +、C2H3 +、C3H5 +
And C5H15NPO4 +Correction.Total ion current is maintained at 1012Below individual ion, it is ensured that in a static mode
Collection surface secondary ion signal.In nucleus, material information is obtained by depth profiling, uses 1kV
(200nA) oxygen source is as the source of anatomy, often dissects 1s, gathers 1 secondary ion mass spectrum signal, time
It is spaced apart 1s.Analyzed area area is 60 × 60 μm2, dissecting region is 300 × 300 μm2。m/z 81
(C5H5O+, ribose fragments) and as the mark fragment of indicator cells film distribution, m/z 102 (102Ru+)
Indication signal for compound 1.
Result shows, can quickly find same according to the method for embodiment 2 in two kinds of analysis methods
Unicellular, the measuring samples plate that the obtains light in laser scanning co-focusing microscope and secondary ion mass spectrum
Learn image as shown in Figure 1A and 1B.
(4) micro-imaging is overlapping
The cell membrane that step (2) and step (3) are obtained and nuclear fluorescence imaging figure and secondary from
Sub-mass spectrum obtains cell membrane and nucleus special component image saves as jpeg format, at Image J
Software is adjusted to identical form and size, uses merge order to be superimposed by two pictures.
Result shows, the present embodiment utilizes addressable silicon chip sample plate can realize laser scanning co-focusing
Microscope single celled imaging analysis identical to the same area with secondary ion mass spectrum, as shown in Figure 4.Ruthenium
Compound 1 is mainly distributed in MCF-7 nucleus, and its action target is DNA.
From the present embodiment it can be seen that addressable silicon chip sample plate that the inventive method prepares can be accurate
And quickly locate same unicellular, it is achieved laser confocal microscope and the associating of secondary ion mass spectrum
Analyze.
Embodiment 3
The present embodiment is used for illustrating that the sample panel that embodiment 1 prepares is combined imaging for SIMS-Confocal
The method that research ruthenium compound 2 (structural formula is as shown in Figure 2 B) is distributed at MCF-7 intercellular spaces.
(1) cultivation of MCF-7 cell and lyophilizing
Sample panel is placed in the Tissue Culture Dish bottom center of laser co-focusing, the MCF-7 digested
Cell is with 104Individual cell/cm2Concentration cultivates 24h in culture dish, adds containing 100 μMs of ruthenium compounds
Hatch 24h after the culture medium of 2, remove culture medium, with the training of the Hoechst 33342 containing 1 μ g/mL
Support and hatch 10min based in 37 DEG C of incubators.Remove this culture medium, then with containing 5 μMs/L cell membrane
The cultivation of red fluorescence dyestuff hatches 20min based in 37 DEG C of incubators.Removing culture medium, cell is through dense
Degree is the NH of 150mM4COOCH3(pH 7.4) buffer solution three times, each 30s.By sample
Plate is quickly put in liquid nitrogen freezing, is then transferred in freezer dryer (-60 DEG C) lyophilization 12h, and
It is gradually increased to room temperature, obtains measuring samples.
(2) laser scanning co-focusing fluorescence imaging analysis
The cell attachment of the measuring samples plate of preparation is faced down, is placed in laser co-focusing Tissue Culture Dish
In, this culture dish is moved on inverted fluorescence microscope (FV1000-IX81, OLYMPUS), use
The reflection light of mercury lamp or Hoechst 33342 exciting light 405nm find the microscale labelling in sample panel and
Unicellular in respective markers region.Switch to fluorescence imaging mode to selection area (A43) cell
In unicellular carry out laser scanning co-focusing fluorescence imaging analysis, obtain this cell at cell membrane and cell
The fluorescence imaging figure in core region.
(3) secondary ion mass spectrum imaging analysis
Video camera built-in for measuring samples plate ion microprobe is found selected district in this sample panel
Same unicellular in territory (A43), gathers this cell secondary ion mass spectrum under following experiment condition
Information: cell surface imaging, 30keV Bi3 +Primary ions bundle, electric current 0.8 μ A, sampling area is 60 × 60
μm2, cation imaging pattern, m/z 184 (C5H15NPO4 +, phosphatidylcholine fragment) and as instruction
The mark fragment of cell membrane pattern, m/z 598 (C26H28ClFN5O2Ru+) as instruction ruthenium compound 2
The mass-fragments of distribution.Mass spectrum is by CH+、CH3 +、C2H3 +、C3H5 +And C5H15NPO4 +Correction.Always
Ion stream is maintained at 1012Below individual ion, it is ensured that collection surface secondary ion signal in a static mode.
In nucleus, material information is obtained by depth profiling, use 1kV (200nA) oxygen source as dissect source,
Often dissecting 1s, gather 1 secondary ion mass spectrum signal, time interval is 1s.Analyzed area area is
60×60μm2, dissecting region is 300 × 300 μm2。m/z 81(C5H5O+, deoxyribose molecule fragment)
As the mark fragment of indicator cells film pattern, m/z 102 (102Ru+) as instruction ruthenium compound 2
Ms fragment.Concrete outcome sees Fig. 3, and wherein, Fig. 3 A is that ruthenium compound 2 is at surface of cell membrane
Mass spectrum imaging figure, Fig. 3 B is that ruthenium element is at endonuclear mass spectrum imaging figure.
(4) micro-imaging is overlapping
The cell membrane that step (2) and step (3) are obtained and nuclear fluorescence imaging figure and secondary from
Sub-mass spectrum obtains the mass spectrum imaging figure of cell membrane and nuclear characteristics ion and saves as jpeg format,
Image J software is adjusted to identical form and size, uses merge order by two pictures superpositions
Together.
Data show, the present embodiment utilizes addressable silicon chip sample plate can realize laser scanning co-focusing
Microscope and secondary ion mass spectrum are to single celled imaging analysis same in the same area, as shown in Figure 5.
Ruthenium compound 2 (m/z 598) has gathering on cell membrane, and with cell membrane mark (m/z 184)
It is distributed identical, illustrates that cell membrane is the action site of compound 2.Compound 2 also has gathering at nucleus,
And it is distributed identical with nucleus marks thing (m/z 81), illustrates that compound 2 can simultaneously act on carefully
After birth and nucleus, have the potential of Mutiple Targets Comprehensive Treatment cancer.
From the present embodiment it can be seen that the inventive method prepare addressable silicon chip sample plate can accurately and
Quickly locate same unicellular, it is achieved laser confocal microscope and the connection of secondary ion mass spectrum imaging
With analysis.
Embodiment 4
The present embodiment is used for illustrating the preparation method of the sample panel of the present invention.
Preparing addressable sample panel according to the method for embodiment 1, except for the difference that, material used is 0.17
Mm is thick, area 22 × 22mm2Coverslip (glass), this sample panel has 84 × 4 arrays,
Wherein the cell in four arrays is square, and four is circular, the size of each square shaped cells lattice
It is 50 × 50 μm2, and a diameter of 50 μm of each circular cell lattice.
Embodiment 5
The present embodiment is used for illustrating that embodiment 4 prepares addressable glass sample plate for studying compound 1
The method of the spatial distribution in HeLa cell (D34).
According to the method for embodiment 3, the micro-imaging of research HeLa cell, except for the difference that, cell with
Compound 1 hatch after only by Hoechst 33342 fluorescent labeling, then with 4% paraformaldehyde (PBS)
After solution fixes 30min under the conditions of 4 DEG C, lyophilizing 48h is analyzed again.Glass sample plate cell is pasted
Wall upward, finds the microscale labelling on glass sample plate, unicellular in selected respective markers region
Directly carry out laser scanning co-focusing fluorescence imaging analysis and secondary ion mass spectrum imaging analysis.
Data show, addressable glass sample plate of embodiment 4 preparation can be applied to unicellular micro-
Imaging analysis (see Fig. 6), can be accurately positioned medicine distribution in cyton.
The preferred embodiment of the present invention described in detail above, but, the present invention is not limited to above-mentioned reality
Execute the detail in mode, in the technology concept of the present invention, can be to the technical side of the present invention
Case carries out multiple simple variant, and these simple variant belong to protection scope of the present invention.
It is further to note that each the concrete technology described in above-mentioned detailed description of the invention is special
Levy, in the case of reconcilable, can be combined by any suitable means, in order to avoid need not
The repetition wanted, various possible compound modes are illustrated by the present invention the most separately.
Additionally, combination in any can also be carried out between the various different embodiment of the present invention, as long as its
Without prejudice to the thought of the present invention, it should be considered as content disclosed in this invention equally.
Claims (10)
1. an addressable sample panel, it is characterised in that be carved with the battle array of N number of n × m in this sample panel
Row, the size of the cell (u) in described array is 100 μm2-4×104μm2。
Sample panel the most according to claim 1, wherein, N=1-10 and each array respective markers
There is different numberings.
Sample panel the most according to claim 1 and 2, wherein, n=4-20, m=4-20 and each
Cell (u) is to there being different numberings.
Sample panel the most according to claim 1 and 2, wherein, described cell (u) is size
It is 10 × 10 μm2-200×200μm2The square and/or circle of a diameter of 10-200 μm.
Sample panel the most according to claim 1, wherein, the size of described sample panel is 0.5 × 0.5
cm2-3×3cm2。
Sample panel the most according to claim 1, wherein, the width of the groove in described sample panel
For 1-10 μm, the degree of depth is 1-10 μm.
7., according to the sample panel described in claim 1,5 or 6, wherein, the material of described sample panel is
Monocrystal silicon or glass.
8. the method for a unicellular micro-imaging, it is characterised in that the method includes: cell is solid
It is scheduled in claim 1-7 in the sample panel described in any one, is marked with fluorescent dye, will be with
Cell in one cell carries out fluorescence imaging analysis and mass spectrum imaging analysis respectively, and described fluorescence imaging divides
Analysis is preferably laser scanning co-focusing fluorescence imaging analysis, and described mass spectrum imaging analysis is preferably secondary ion
Mass spectrum imaging is analyzed.
Method the most according to claim 8, wherein, cell density in described sample panel is
104-105Individual cell/cm2。
10. sample panel described in any one application in unicellular micro-imaging in claim 1-7.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2018189365A1 (en) * | 2017-04-13 | 2018-10-18 | European Molecular Biology Laboratory | Single-cell imaging mass spectrometry |
| CN110763752A (en) * | 2019-11-27 | 2020-02-07 | 清华大学 | Single cell extraction electrospray mass spectrometry system and method |
| CN114019010A (en) * | 2021-11-04 | 2022-02-08 | 上海交通大学 | Microorganism unicellular metabonomics analysis method |
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