CN1609590A - Three-differential focasing micro-three-dimensional super-resolution imaging method - Google Patents

Abstract

The present invention belongs to the field of optical microscopic imaging and micro measurement technology, and is one 3D three-differential confocal microscopic imaging method with very high S/N ratio and 3D super resolution imaging capacity. The present invention fuses the three-differential confocal scanning method with high axial resolution and pupil filtering confocal scanning method with high transverse resolution to constitute the 3D pupil filtering three-differential confocal microscopic imaging method. The method may be used in measuring 3D surface appearance, 3D fine structure, micro step, micro channel, IC line width, etc.

Description

Three-differential focasing micro-three-dimensional super-resolution imaging method

Technical field

The invention belongs to optical microphotograph imaging and microscopic measurement technical field, relate to a kind of have high property make an uproar than with the three-differential focasing micro-three-dimensional super-resolution imaging method of three-dimensional super-resolution imaging ability, it can be used for 3 d surface topography, three-dimensional microstructure, little step, little groove, integrated circuit live width of measuring samples etc.

Background technology

The thought of confocal microscope is proposed in nineteen fifty-seven first by American scholar M.Minsky the earliest, and obtains the United States Patent (USP) mandate in 1961, and the patent No. is US3013467.Confocal microscope places the conjugate position that corresponds to each other with pointolite, some thing and point probe three, and the some illumination and the point that constitute unique chromatography ability in the optical microphotograph imaging are surveyed micro imaging system.The ultimate principle of general confocal microscope as shown in Figure 1, the light that light source 32 sends returns along former road after the testee surface is focused into hot spot through pin hole 33, object lens 5, to import from the flashlight of object by spectroscope 10 again and be placed in the pin hole 7 of detector 8 fronts, forming point at detector 8 places detects, the detector 8 main flashlights that receive from the object focal point place, the back light beyond the focus is blocked by pin hole 7.When object is positioned at focal plane A, the luminous energy maximum that detector 8 receives, when object departs from focal plane A, reflected light is focused a certain position before or after the pin hole, this moment, detector only received the sub-fraction luminous energy, that is to say that signal that object detects is than in the focal plane time when out of focus a little less than, so just can change and reflect the position of object with respect to the focal plane by the power that detector detects light intensity signal.When object when doing scanning motion perpendicular to the x-y plane of optical axis direction, confocal microscope according to optical axis z to defocus signal, x to y to the displacement size, can construct the three-D profile of testee.

Confocal microscope is widely used in carrying out imaging measurement in the fields such as microelectronics, material, the accurate detection of industry, biomedicine, life science because of it has three-dimensional chromatography imaging capability, but, restricted the further raising of its imaging resolution characteristic owing to be subjected to the restriction of diffraction phenomena.Although its imaging resolution characteristic can be improved with traditional methods such as reducing optical wavelength by increasing numerical aperture of objective value NA, its improvement degree still is subjected to the restriction of diffraction limit.For fundamentally breaking through diffraction limit, improve the imaging resolution characteristic of confocal microscope, existing recently numerous unconventional confocal microscopic imaging principles and ultra-resolution method are suggested.Aspect the research of confocal microscope, 4PI confocal microscope, θ confocal microscope, confocal interference microscope have appearred and based on the two-photon of optical nonlinearity behavior and multi-photon confocal microscope etc.; At the super-resolution imaging technical elements, method of having studied and technology can be classified as following a few class, one class is the Airy disk that reduces by the Rayleigh criterion decision, but do not increase the spatial-cut-off frequency of optical system, technology commonly used comprises: pupil filtering technology, Phase-Shift Masking Technique, based on super resolution technology of optical property nonlinearities change etc.; Second class is by increasing the optical system spatial-cut-off frequency, increase high frequency light proportion, reducing the Airy disk main lobe of optical system; The 3rd class is by changing optical system incident beam spatial frequency distribution, reach the purpose that reduces optical system Airy disk main lobe, generally can realize by light illumination technology such as off-axis illumination technology, deformation illumination technology, crossed polarized light lighting engineering, ring light lighting engineering and interfering beam space frequency displacement methods.

On the whole, above-mentioned novel confocal microscope and ultra-resolution method and technology, improved the imaging resolution characteristic of confocal microscope, solved the demand that numerous confocal microscope super-resolution micro-imagings are measured, but still there is following problem demanding prompt solution in they: the one, and existing various forms of confocal microscopes all are to utilize the light intensity signal that detects directly to carry out imaging processing at present, it is subject to the influence of factors such as light-intensity variation, bias light interference, environment temperature drift, and the confocal microscopy imaging system signal to noise ratio (S/N ratio) is low; The 2nd, the axial chromatography precision of confocal microscope is limited by the non-linear of axial strength response curve, and existing super resolution technology easily causes the increase of secondary lobe and the increase of axial response curve nonlinearity erron in the super-resolution imaging process.For improving confocal microscope tomography ability, the applicant has proposed to improve the three differential confocal microscopic imaging method of confocal microscope signal to noise ratio (S/N ratio) and azimuthal resolution, and applied for being entitled as the Chinese invention patent of " three differential confocal microscopic imaging method and device ", application number is 20041000736524 (inventors: Zhao Weiqian, Tan Jiubin, Qiu Lirong).But this three differential confocal microscopic imaging method is mainly used in the axial resolution of improving confocal microscope, fail to improve the horizontal resolution characteristic of confocal microscope, and existing super-resolution pupil filter is used for confocal microscope when carrying out three-dimensional super-resolution imaging, should carry out transverse super-resolution and take into account axial super resolution again, the three-dimensional super-resolution effect is not remarkable especially usually.

Summary of the invention

The objective of the invention is for overcoming the deficiency of above-mentioned prior art, merge optical ultra-discrimination and three differential confocal microscope characteristics separately, the confocal microscopic imaging method with three-dimensional super-resolution imaging ability of a kind of lateral optical super-resolution, axial three differential confocal super-resolution is provided, comes that micro-imaging is carried out in fields such as microelectronics, material, the accurate detection of industry, biomedicine, life science and detect.

Technical solution of the present invention is: a kind of three-differential focasing micro-three-dimensional super-resolution imaging method comprises the following steps:

(1) incident light is passed through iris filter (2), polarization spectroscope (3), measurement object lens (5) through the three differential confocal microscopic system carry out scanning imagery to sample, and detector (8), (15) and (19) record the intensity response I that reflection sample convex-concave changes respectively ₁(v, u, 0), I ₂(v, u ,-u _M) and I ₃(v, u ,+u _M);

(2) with I ₁(v, u, 0) subtracts I ₂(v, u ,-u _M) must I _A(v, u), I ₁(v, u, 0) subtracts I ₂(v, u ,-u _M) must I _B(v, u), I ₂(v, u ,-u _M) subtract I ₃(v, u ,+u _M) must I _C(v, u), then obtain the intensity I that corresponding sample convex-concave changes (v u) is:

And intensity curved surface;

(3) optimize iris filter parameters such as amplitude type wave filter, phase-type wave filter, amplitude position phase hybrid filter, make I ₁(v, u, 0), I ₂(v, u ,-u _M) and I ₃(v, u ,+u _M) transverse strength response satisfies G _T, M and S designing requirement, (v, main lobe u) improve the transverse resolution of three differential confocal microscope to sharpening three differential confocal microscope I;

(4) optimize pin hole (14) and pin hole (18) optics normalization coordinate u apart from its corresponding condenser focal position _M, make the improvement of confocal microscope azimuthal resolution reach optimum;

(5) (v, u) intensity curve light intensity magnitude reconstruct the microcosmic three-dimensional appearance and the micro-scale of sample according to I;

(6) utilize I _C(v, u) intensity curve carry out the absolute tracking measurement of bipolarity to sample three-dimensional appearance and microtexture.

The inventive method will improve the three differential confocal scan method of azimuthal resolution and the super-resolution pupil filtering cofocus scanning method of raising transverse resolution merges, and constitute pupil filtering formula three-dimensional super-resolution differential confocal imaging method.The iris filter of employing particular design carries out the mask correction to the pupil function of three differential confocal microscope, and then changes wavefront, and sharpening Airy disk main lobe finally improves differential confocal microscope transverse super-resolution power.Iris filter can be phase-type wave filter, an amplitude type wave filter and amplitude position hybrid filter mutually.The raising of azimuthal resolution can be undertaken by three differential confocal light path arrangement and differential detection, like this, just can reach three-dimensional super-resolution imaging ability and high property and make an uproar and detect than micro-imaging.

Detection method of the present invention has following characteristics and good result:

The present invention is owing to merged the axial super resolution characteristic of pupil filtering formula confocal microscopy transverse super-resolution characteristic and three differential confocal light path arrangement method, avoided existing three-dimensional super-resolution iris filter should improve transverse resolution, improve azimuthal resolution again, thereby reduce the shortcoming of three-dimensional super-resolution combination property, make confocal microscope can improve the three-dimensional super-resolution imaging ability (axially with horizontal) of confocal microscope, can significantly strengthen environment interference, linearity and out of focus characteristic etc. again, this is one of innovative point that is different from prior art.

Utilize three detectable signals to carry out data processing, make confocal microscope have the function of differential confocal measuring system, compare with common confocal microscope, three differential confocal microscope also is convenient to the high precision absolute measurement of three-dimensional surface profile and microtexture, it merges surface microscopic topographic mutually with dimensional measurement, this be different from prior art innovative point two.

Measuring method of the present invention has following characteristics:

1) make confocal microscope have the three-dimensional super-resolution imaging ability;

2) the light path arrangement method and the differential in twos detection method that subtracts each other of three detector signals of three differential confocal reception can suppress the common-mode noise that ambient condition difference, light source intensity fluctuation, the electric drift of detector etc. cause, significantly improve signal to noise ratio (S/N ratio), sensitivity and the linearity etc. of measuring system;

3) measuring system has absolute tracking zero point and bipolarity tracking characteristics, can realize absolute measurement;

4) improved the out of focus characteristic of confocal system.

Description of drawings

Fig. 1 confocal microscope schematic diagram.

Fig. 2 is for adopting the three-differential focasing micro-three-dimensional super-resolution imaging method synoptic diagram of pupil filtering technology.

Fig. 3 is a formation method of the present invention, works as u _M=5.21 o'clock three-dimensional intensity response emulation of confocal microscope surface charts.

Fig. 4 is a formation method of the present invention, works as u _M=5.21 o'clock three-dimensional intensity response normalization of confocal microscope emulation surface charts.

Fig. 5 is a phase-type pupil filtering transverse super-resolution light intensity response curve comparison diagram.

Fig. 6 is differential wave sensitivity simulation curve figure of the present invention.

Fig. 7 works as u for the present invention _M=5.21 o'clock three differential confocal microscope axial strength response simulation curves.

Fig. 8 works as u for the present invention _M=5.21 o'clock three differential confocal microscope axial strength response normalization simulation curves.

Fig. 9 is three differential confocal microscope transverse strength response simulation curve figure of the present invention.

Figure 10 is three differential confocal microscope transverse strength response normalization simulation curve figure of the present invention.

Wherein, 1 incident beam, 2 iris filters, 3 polarization spectroscopes, 4 λ/4 wave plates, 5 object lens, 6,13,17 condensers, 7,14,18 pin holes, 8,15,19 detectors, 9,11,16 dimmers, 10,12 spectroscopes, the differential normalized unit that subtracts each other of 20,21,22 focus signals, 23 computer processing systems, 24 testees, 25 three-dimensional working platforms, 26 three differential confocal microscopes, 27I _C(0, u) confocal microscope axial response curve, 28 axial strengths response I (0, u) curve, 29 axial strengths response I ₂(0, u ,-u _M) curve, 30 axial strengths response I ₃(0, u ,+u _M) curve, 31 axial strengths response I ₃(0, u, 0) curve, 32 LASER Light Source, 33 diaphragms, 34I (v, u) intensity response curve, 35 sensitivity k _A(0,0, u _M) simulation curve, 36 sensitivity k _B(0,0, u _M) simulation curve, 37 sensitivity k _C(0,0, u _M) simulation curve, 38 transverse strengths response I ₁(v, 0,0) curve, 39 transverse strengths response I (v, 0) curve.

Embodiment

Of the present invention three differential micro imaging methods are to adopt the three differential confocal microscopic imaging technology that the receiving light path of confocal microscope is arranged as Jiao far away, focal plane and nearly burnt three tunnel detection light paths, differential in twos the subtracting each other of detecting by three tunnel detection systems of three tunnel intensity response signals with coordination phase not reaches the purpose of improving azimuthal resolution and improving antijamming capability, improve the transverse resolution of confocal microscope by super-resolution pupil filtering type confocal microscopic imaging method, make confocal microscope final realize high property make an uproar than with the three-dimensional super-resolution micro-imaging.

As shown in Figure 2, empty frame part 26 is the micro-light path arrangement of three differential confocal, incident beam 1 becomes the p light that the polarization direction is parallel to paper after seeing through polarization spectroscope 3 (PBS) through iris filter 2, after this p light transmission λ/4 wave plates, 4 backs are focused on testee 24 surfaces by object lens 5, seen through λ/4 wave plates 4 once more by testee 24 backspaces and become the s light of polarization direction perpendicular to paper, polarization spectroscope 3 reflection s light are to spectroscope 10.Spectroscope 10 at first is divided into measuring beam two bundles, and the measuring beam that reflects through spectroscope 10 is focused on by condenser 6, enters the pin hole 7 that is positioned at condenser 6 focal positions, is detected device 8 and receives; Light through spectroscope 10 transmissions is divided into two bundles by spectroscope 12 once more, is focused on by condenser 13 through the measuring beam of spectroscope 12 reflection, enters that distance be the pin hole 14 of M position after condenser 13 focuses, after be detected device 15 receptions; Measuring beam through spectroscope 12 transmissions is focused on by condenser 17, and entering before condenser 17 focuses apart from focal length is the pin hole 18 of M, is received by the detector behind the pin hole 18 19.

When 25 pairs of samples of worktable 24 carry out axially (axially normalization optical displacement be made as u) and laterally (when horizontal normalization optical displacement was made as and v) scans, the signal that detector 8 detects was I ₁(v, u, 0), the signal that detector 15 detects are I ₂(v, u ,-u _M), the signal that detector 19 detects is I ₃(v, u ,+u _M), the differential disposal system 22 of subtracting each other is with I ₁(v, u, 0) subtracts I ₂(v, u ,-u _M) must I _A(v, u), the differential disposal system 21 of subtracting each other is with I ₁(v, u, 0) subtracts I ₃(v, u ,+u _M) must I _B(v, u), the differential disposal system 20 of subtracting each other is with I ₂(v, u ,-u _M) subtract I ₃(v, u ,+u _M) must I _C(v, u), with I _A(v, u), I _B(v, u) and I _C(v, u) input is handled behind the computer processing system 23:

(v, u) as shown in Figure 3, normalized intensity responds as shown in Figure 4 three differential confocal microscope intensity response I.Intensity I (v, u) corresponding sample convex-concave changes, and (v, the u) light intensity magnitude of intensity curve in measurement range reconstruct the 3 d surface topography and the micro-scale of sample, can realize that the three-dimensional super-resolution micro-imaging of confocal microscope is surveyed according to I.

Its concrete principle is as follows:

When sample 24 is near Jiao, near the maximal value of the corresponding curve 30 of the axial response signal that detector 19 detects, the A end of the differential response curve 28 of corresponding confocal microscope this moment; When sample 24 is in the focal position, the signal that detector 19 detects is in the descending branch of curve 30, and the signal that detector 15 detects is in the ascent stage of curve 29, and the signal that detector 8 detects is in the most significant end of detective curve 31 just; When sample 24 is in burnt position far away, the signal that detector 8 detects is in the descending branch of detective curve 31, the signal that detector 19 detects is in the descending branch of curve 30, and the signal that detector 15 detects is in the ascent stage of curve 29, at this moment the descending branch of corresponding curve 28; When sample moves in the AB zone, the variation of curve 28 corresponding measured object defocusing amount sizes can be finished the tomography function of confocal microscope.

When being used for object three-dimensional contour outline and microtexture and measuring, when sample 24 is in nearly Jiao Qu point A place, corresponding curve 30 maximal values of the signal that detector 19 detects, this moment, a of differential wave correspondence 27 curves held; Near sample 24 was in the focal position, and the signal that detector 19 detects is in the middle part of curve 30 descending brancies, the signal that detector 15 detects was near 29 ascent stages of the curve middle part, the actual zero point of corresponding 27 curves of differential confocal signal; Distinguish the B place when sample 24 is in nearly Jiao, the signal that detector 15 detects is near the maximal value of curve 30, and this moment, the b of corresponding 27 curves held; In near the ab zone of sample focus, move the ab section of corresponding 27 curves of differential confocal signal.

From 27 curves, as can be seen, compare with 31 hypotenuse section with single channel confocal microscopy family curve 29,30,27 characteristic hypotenuse section steepenings, sensitivity is improved, and azimuthal resolution improves.

The present invention is had high property to make an uproar and further specifies as follows than the three-differential focasing micro-three-dimensional super-resolution imaging method with the three-dimensional super-resolution imaging ability:

Still as shown in Figure 2, empty frame partly is that micro-three receiving light paths of three differential confocal arrange 26, and iris filter 2 is the ring-like iris filter of N district concentric circles.Under the monochromation illumination condition, have near the distribution of amplitudes of pupil function P (ρ) system focus to be:

$U(v,u)=2{\∫}_0^{1}p\left(\mathrm{\ρ}\right)\·\exp ({-\mathrm{ju\ρ}}^2/2)\·J_0\left(\mathrm{v\ρ}\right)\mathrm{\ρd\ρ}$

ρ-normalization radius, the radial coordinate r on the corresponding receiving plane of v-, the u-correspondence is that the axle of initial point is gone up coordinate z with the focus,

Wherein:

t _jTransmittance function, for the j district _jBe the phase differential in j district, after measuring object lens 5 focusing, the lateral amplitude of vibration response characteristic on the focal plane is:

To the amplitude type iris filter, in the formula (5), _j=C (C is a constant), t _jBe variable (j=1,2,3 ..., N) to pure phase bit-type iris filter, in the formula (5), t _j=C (C is a constant), _jBe variable (j=1,2,3 ..., N) to amplitude position phase hybrid filter, in the formula (5), t _j, _jBe variable (j=1,2,3 ..., N)

Analyze with pure phase bit-type iris filter at this, other roughly the same.Consider pure phase bit-type iris filter, then t _j=C (j=1,2,3 ..., N), make C=1.

The radius of supposing emergent pupil is R, and incident wavelength is λ, (R _j=a _jR, a ₀=0, a _N=1, ₁=0), for the symmetrical phase-type iris filter of N district circle, focal plane field amplitude expression formula is:

$I(v,0)={\left|U(v,0)\right|}^2$

(7)

Because $\frac{J_1\left(x\right)}{x}=\frac{1}{2}(1-\frac{x^2}{4\×2}+\frac{x^4}{2\×4^2\×6}+\mathrm{LL})---\left(8\right)$

It is approximate to get two-stage, promptly gets $\frac{J_1\left(x\right)}{x}\≈\frac{1}{2}(1-\frac{x^2}{8}),$ Corresponding light intensity is;

Order $I(v,0)=a-\frac{v^2}{4}b+\frac{v^2}{64}c---\left(10\right)$

Wherein

During no pupil wave filter, $I_0(v,0)=1-\frac{{\mathrm{<figure-callout\; id="2"\; label="v"\; filenames="A20041009077400132.png,A20041009077400142.png"\; state="\{\{state\}\}">v</figure-callout>}}^2}{4}+\frac{{\mathrm{<figure-callout\; id="4"\; label="v"\; filenames="A20041009077400141.png,A20041009077400154.png"\; state="\{\{state\}\}">v</figure-callout>}}^4}{64}---\left(12\right)$

$\frac{\&PartialD;I}{\&PartialD;v}=0\&DoubleRightArrow;v_1=0,v_{\mathrm{2,3}}=\&PlusMinus;2\sqrt{2}$

The central light strength extreme value is: I ₀(0,0)=1

(13)

When iris filter is arranged, $\frac{{\&PartialD;I}_F}{\&PartialD;v}=0,$ Then $-\frac{b\&CenterDot;\mathrm{<figure-callout\; id="2"\; label="v"\; filenames="A20041009077400132.png,A20041009077400142.png"\; state="\{\{state\}\}">v</figure-callout>}}{2}+\frac{{\mathrm{<figure-callout\; id="3"\; label="v"\; filenames="A20041009077400132.png,A20041009077400154.png"\; state="\{\{state\}\}">v</figure-callout>}}^3\&CenterDot;\mathrm{<figure-callout\; id="16"\; label="c"\; filenames="A20041009077400132.png"\; state="\{\{state\}\}">c</figure-callout>}}{16}=0,$ Solve an equation to such an extent that the extreme point coordinate is as follows:

(15)

The ratio G of the response curve halfwidth when iris filter and no pupil wave filter are arranged is:

$G_T=\sqrt{\frac{b}{c}}$

(16)

The ratio Streh1 of the focus maximum of intensity when iris filter and no pupil wave filter are arranged than S is:

(17)

Utilize optimal design method, under the condition of given G, S and ε, determine the N position phase-plate phasic difference in zone separately _jWith normalization radius a _jOptimal conditions is:

Objective function F ( _j, a _j)=G _T-0.75≤ε, ε=0.02, S 〉=0.22,0＜ _j＜2 π, 0＜a _j＜1 a _N=1;

Objective function F ( _j, a _j)=G _T-0.80≤ε, ε=0.02, S 〉=0.28,0＜ _j＜2 π, 0＜a _j＜1 a _N=1;

Objective function F ( _j, a _j)=G _T-0.85≤ε, ε=0.02, S 〉=0.30,0＜ _j＜2 π, 0＜a _j＜1 a _N=1;

Optimized Algorithm adopts Generic Algorithms algorithm.

Get N=4, promptly when the phase-type iris filter is 4 district wave filters, choose following three groups of G _TWith S as optimization aim, optimize the iris filter parameter that obtains three groups of correspondences after finding the solution:

1) works as G _T=0.7643, during S=0.25, four corresponding position facies pattern iris filter a ₁=0.1, a ₂=0.2, a ₃=0.5199, a ₄=1, ₁=0, ₂=2.8634, ₃=1.5222rad, ₄=5.8372rad;

2) work as G _T=0.8020, during S=0.3, four corresponding position facies pattern iris filter a ₁=0.2, a ₂=0.3058, a ₃=0.5332, a ₄=1, ₁=0, ₂=1.5777rad, ₃=3.0112rad, ₄=5.7177rad;

3) work as G _T=0.8512, during S=0.35, four corresponding position facies pattern iris filter a ₁=0.3, a ₂=0.4, a ₃=0.5804, a ₄=1, ₁=0, ₂=1.6834rad, ₃=3.6583rad, ₄=6.2829rad.

With the curve plotting of above-mentioned three kinds of phase-type iris filter transverse super-resolutions in Fig. 5, therefrom as can be seen: after adding the phase-type iris filter, the transverse response curve obtains sharpening, and G value is more little, and sharpening is obvious more, transverse resolution raising obvious more.Disadvantage is that secondary lobe strengthens, it is that the S value diminishes that optical energy loss also increases, but secondary lobe can the pin hole by confocal microscope system suppress (this just iris filter combine with confocal microscopy, realize the reason of super diffraction resolved detection truly), optical energy loss can solve by the enlargement factor that increases detection system.

The improvement of azimuthal resolution is by preferred u _MThe value size reaches, with differential wave I _A(0, u) differentiate gets sensitivity k to u _A(0, u, u _M):

k _A(0，u，u _M)＝sin?c[(u/2π)]·[(u/2)·cos(u/2)-sin(u/2)]/(u/2) ²-

(18)

sin?c[(2u-u _M)/4π][{(2u-u _M)/4}·cos{(2u-u _M)/4}-sin{(2u-u _M)/4}]/{(2u-u _M)/4} ²

Slope value k in linearity range _A(0, u, u _M) and k _A(0,0, u _M) equate therefore have:

k _A(0，0，u _M)＝sin?c[(u _M)/4π]·[(u _M/4)·cos(u _M/4)-sin(u _M/4)]/{(u _M)/4} ² (19)

With differential wave I _B(0, u) differentiate gets sensitivity k to u _B(0, u, u _M):

k _B(0，u，u _M)＝sin?c(u/2π)·[(u/2)·cos(u/2)-sin(u/2)]/(u/2) ² (20)

-sin?c[(2u+u _M)/4π]·[{(2u+u _M)/4}·cos{(2u+u _M)/4}-sin{(2u+u _M)/4}]/{(2u+u _M)/4} ²

Slope value k in linearity range _B(0, u, uM) and k _B(0,0, u _M) equate therefore have:

k _B(0，0，u _M)＝-sin?c[(u _M)/4π]·[(u _M/4)·cos(u _M/4)-sin(u _M/4)]/{(u _M)/4} ² (21)

With differential wave I _C(0, u) differentiate gets sensitivity k to u _C(0, u, u _M):

k _C(0，u，u _M)＝sin?c[(2u-u _M)/4π]·[{(2u-u _M)/4}·cos{(2u-u _M)/4}-sin{(2u-u _M)/4}]{(2u-u _M)4} ² (22)

-sin?c[(2u+u _M)/4π]·[{(2u+u _M)/4}·cos{(2u+u _M)/4}-sin{(2u+u _M)/4}]/{(2u+u _M)/4} ²

Slope value k in linearity range _C(0, u, u _M) and k _C(0,0, u _M) equate therefore have:

k _C(0，0，u _M)＝-2sin?c[(u _M)/4π]·[(u _M/4)·cos(u _M/4)-sin(u _M/4)]/(u _M/4) ² (23)

According to formula (19), (21) and (23), with I _A(0, u), I _B(0, u) and I _C(0, u) sensitivity curve of intensity response linearity range is plotted among Fig. 6, therefrom works as u as can be seen _M=± 5.21 o'clock, sensitivity k _A(0,0, u _M) curve 35, sensitivity k _B(0,0, u _M) curve 36 and sensitivity k _C(0,0, u _M) the absolute value maximum of curve 37 correspondences, at this moment, corresponding I _A(0, u), I _B(0, u) and I _C(0, u) the sensitivity absolute value maximum of curve linear section, I (0, azimuthal resolution optimum u).

Fig. 7 is for working as u _M=5.21 o'clock, I ₁(0, u, 0), I ₂(0, u ,-u _M), I ₃(0, u ,+u _M), I _C(0, u) and I (0, response curve u), Fig. 8 are its normalized response curve.During the confocal microscope tomography, often be operated in I (0, measuring section u) 〉=0, as can be seen from Figure 8, (0, u) halfwidth of curve compares I at this surveying work section I ₁The little twice of halfwidth of (0, u, 0) curve, promptly that the confocal microscope azimuthal resolution has been improved than confocal microscope is about more than 65% for three-differential focasing micro-three-dimensional super-resolution imaging method, and (0, u) linearity of two hypotenuse sections obviously is better than I to I ₁The linearity of (0, u, 0) two hypotenuse sections, (0, surveying work section near-in sidelobe u)＞0 is minimum to the influence of measuring at I simultaneously.

Fig. 9 works as u _M=5.21 o'clock, I ₁The transverse response curve of (v, 0,0) and I (v, 0), during the confocal microscope tomography, (v, measuring section u) 〉=0, Figure 10 are its normalized curve often to be operated in I.As can be seen from Figure 10, the little I that compares of halfwidth of I (v, 0) curve in this surveying work section ₁The halfwidth of (v, 0,0) curve is little, and promptly iris filter improves the transverse resolution of confocal microscope.

Claims (1)

1. a three-differential focasing micro-three-dimensional super-resolution imaging method is characterized in that comprising the following steps:

(1) incident light is passed through iris filter (2), polarization spectroscope (3), measurement object lens (5) through the three differential confocal microscopic system carry out scanning imagery to sample, and detector (8), (15) and (19) record the intensity response I that reflection sample convex-concave changes respectively ₁(v, u, 0), I ₂(v, u ,-u _M) and I ₃(v, u ,+u _M);

(2) with I ₁(v, u, 0) subtracts I ₂(v, u ,-u _M) must I _A(v, u), I ₁(v, u, 0) subtracts I ₂(v, u ,-u _M) must I _B(v, u), I ₂(v, u ,-u _M) subtract I ₃(v, u, u _M) must I _C(v, u), then obtain the intensity I that corresponding sample convex-concave changes (v u) is:

And intensity curved surface;

(3) optimize iris filter parameters such as amplitude type wave filter, phase-type wave filter, amplitude position phase hybrid filter, make I ₁(v, u, 0), I ₂(v, u ,-u _M) and I ₃(v, u ,+u _M) transverse strength response satisfies G _T, M and S designing requirement, (v, main lobe u) improve the transverse resolution of three differential confocal microscope to sharpening three differential confocal microscope I;

(4) optimize pin hole (14) and pin hole (18) optics normalization coordinate u apart from its corresponding condenser focal position _M, make the improvement of confocal microscope azimuthal resolution reach optimum;

(5) (v, u) intensity curve light intensity magnitude reconstruct the microcosmic three-dimensional appearance and the micro-scale of sample according to I;

(6) utilize I _C(v, u) intensity curve carry out the absolute tracking measurement of bipolarity to sample three-dimensional appearance and microtexture.

Images