Summary of the invention
The objective of the invention is to improve for the weak point that present various confocal optical scanners exist, as slow in single-point confocal laser scanning speed of scanner, sensitivity is low; Rotating disk confocal scanning instrument has bright dark alternate sweep trace interference, illumination intensity heterogeneity; Slide block confocal scanning instrument sweep velocity waits problem slowly, and a kind of new spininess hole confocal optical scanner is provided, and coordinates fluorescent microscope and face battle array detecting device to form confocal imaging system, thereby obtains at high speed high-quality Confocal Images.
The objective of the invention is to realize by following technical scheme:
As shown in Figure 1, the invention provides a kind of spininess hole confocal optical scanner, mainly comprise: light source 1; Excite color filter 2; Illuminating lens group 3; Microlens array 4 has been arranged lenticule 4a on it; Dichroic beam splitter 5; Pinhole array 6 has been arranged pin hole 6a on it; Delay lens group 7; Scanning galvanometer 8 has the first reflecting surface 8a and the second reflecting surface 8b; Cut-off diaphragm 9 is divided into light transmission part 9a and lightproof part 9b; Total reflective mirror group 10; Imaging lens group 11; Emission color filter 12; Controller 13; It is characterized in that:
Any one in described light source 1 use laser, light emitting diode, mercury lamp, xenon lamp and metal halide lamp or multiple as light source;
Describedly excite the effect of color filter 2 to be, see through the part wavelength of described light source 1 emission light, reflect the light of other wavelength, obtain the exciting light for the illumination sample;
The effect of described illuminating lens group 3 is, described light source 1 is launched and seen through the described exciting light of color filter 2 that excites to form parallel beam;
Described microlens array 4 is positioned at described illuminating lens group 3, and perpendicular to described parallel beam, has arranged described lenticule 4a a plurality of printing opacities, that have the phase parfocal on it, and remainder is light tight; Described lenticule 4a is circle or polygon, as triangle, square, rectangle, quadrilateral, pentagon and hexagon etc.; Described lenticule is one or more in Fresnel lens, curved reflector, miniature lenticular lens type lenticule, and has equal focal length;
Described dichroic beam splitter 5 has 45 degree angles with described parallel beam between described microlens array 4 and described pinhole array 6;
After described pinhole array 6 is placed on described dichroic beam splitter 5, perpendicular to described parallel beam, and be positioned on the focal plane of described dichroic beam splitter 5 one sides of described lenticule 4a;
Arranged the described pin hole 6a of a plurality of printing opacities on described pinhole array 6, remainder is light tight; Described pin hole 6a is circle or polygon, as triangle, square, rectangle, quadrilateral, pentagon and hexagon etc.;
When described pin hole 6a was circular, the big or small r of described pin hole 6a was defined as the radius of described circle; When described pin hole 6a was polygon, the big or small r of described pin hole 6a was defined as described polygonal inscribe radius of a circle;
The big or small r of described pin hole 6a is determined by the enlargement ratio M2 of numerical aperture NA, the enlargement ratio M1 of the object lens 15 of microscope 19, described delay lens group 7 and the wavelength X of described sample 16 emitting fluorescences:
r=0.61×λ÷NA×M1×M2
As shown in Figure 8, the described pin hole 6a on described pinhole array 6 is according to following regularly arranged: when described scanning galvanometer 8 rotation, described lighting point mobile direction on described sample 16 is defined as the direction of scanning; The arrangement that will be parallel to the described pin hole 6a of described direction of scanning is defined as row; To be defined as row perpendicular to the arrangement of the described pin hole 6a of described direction of scanning; The quantity of the described pin hole 6a of every delegation is equal, and the quantity of the described pin hole 6a of each row equates, and each walks to a rare described pin hole 6a; Pass the center of described pin hole 6a and draw the straight line that is parallel to described direction of scanning, the distance of two adjacent straight lines is the line space of the adjacent two described pin hole 6a of row, is called I; Pass the center picture of described pin hole 6a perpendicular to the straight line of direction of scanning, the distance of two adjacent straight lines is the column pitch of the adjacent two described pin hole 6a of row, is called J; Centre distance with adjacent two described pin hole 6a of delegation is defined as K; The centre distance of adjacent two described pin hole 6a of same row is defined as L; The arrangement of described pin hole 6a makes the big or small r of I, J, K and L and described pin hole 6a that following equation relation be arranged:
0<I≤2r
L=n×I,
K
2=n
2×L
2-I
2
J
2=L
2-I
2
N is the coefficient of described pin hole 6a spacing for a change, 1<n≤20;
As shown in Figure 9, described lenticule 4a and described pin hole 6a have identical arrangement, form one-to-one relationships through described dichroic beam splitter 5, namely described parallel beam by described lenticule 4a beam splitting, converge, the focus that converges is positioned at the center of the described pin hole 6a after described dichroic beam splitter 5; Described parallel beam beam splitting forms a plurality of pointolites after seeing through described pin hole 6a;
7 pairs of described pinhole array 6 imagings of described delay lens group as the picture planes overlapping of plane and 15 pairs of described sample 16 imagings of described object lens, make described pointolite form the lighting point one to one with described pin hole 6a on described sample 16;
Described scanning galvanometer 8 has two reflectings surface around the fixed rotating shaft reciprocating rotary, i.e. the first reflecting surface 8a and the second reflecting surface 8b; The first reflecting surface 8a and the second reflecting surface 8b are plane reflection or concave reflection; When described the first reflecting surface 8a and the second reflecting surface 8b are concave reflection, the focal length that equates or do not wait is arranged;
Described cut-off diaphragm 9 is positioned at described sample 16 through the picture plane of described object lens 15 imagings, is divided into described light transmission part 9a and described lightproof part 9b;
Described total reflective mirror group 10 is plane reflection or concave reflection;
11 pairs of described pinhole array 6 imagings of described imaging lens group are positioned at as the plane on the sensitive chip 14 of face battle array detecting device 20;
The effect of described emission color filter 12 is, sees through the fluorescence of the specific wavelength of described sample 16 emissions, reflects the light of other wavelength;
The effect of described controller 13 is: control position of rotation, the swing circle of described scanning galvanometer 8, and carry out signal and communication with described battle array detecting device 20, realize the rotation of described scanning galvanometer 8 and the exposure synchro control of described battle array detecting device 20.
Advantage of the present invention is as follows:
1, the present invention by the scanning galvanometer driven sweep, is used in conjunction with the high sensitivity face battle array detecting devices such as CCD, EMCCD, CMOS and sCOMS, can realize high speed, the burnt optical imagery of highly sensitive copolymerization.
2, the present invention is single pinhole array, and exciting light and fluorescence are installed and regulated simply through same pinhole array.
3, the present invention uses microlens array and the two array structures of pinhole array, and lenticule and pin hole are corresponding one by one, and the light source utilization factor is high, without the parasitic light ground unrest.
4, the scanning of scanning galvanometer of the present invention is synchronizeed with the exposure of face battle array detecting device, avoids scanning the visual bright dark inhomogeneous problem that not exclusively causes.
5, the present invention is fully equal to the illumination intensity of sample, visual field illumination intensity homogeneous.
Embodiment
Further describe the present invention below in conjunction with drawings and Examples.
Embodiment 1
Fig. 2 a and 2b are the schematic diagram of the first confocal optical scanner related to the present invention, and wherein, the first reflecting surface 8a of scanning galvanometer 8, the second reflecting surface 8b and total reflective mirror group 10 are all plane reflections.
As Fig. 2 a, in the present embodiment, before face battle array detecting device 20 (not shown)s began exposure, controller 13 (not shown) gated sweep galvanometers 8 rested on maximum spin angular position forward or backwards; Light is launched from light source 1, forms a plurality of shot point light sources through exciting the pin hole 6a on lenticule 4a, dichroic beam splitter 5 and the pinhole array 6 on color filter 2, illuminating lens group 3, microlens array 4; Delayed lens combination 7 imagings of shot point light source and the first reflecting surface 8a that is scanned galvanometer 8 are reflected in the lightproof part 9b of cut-off diaphragm 9, can not enter microscope 19 (not shown)s, form lighting point on the sample 16 that is positioned at object lens 15 focal planes.
As Fig. 2 b, battle array detecting device 20 (not shown)s begin exposure face to face, controller 13 (not shown) gated sweep galvanometers 8 begin clockwise or are rotated counterclockwise to the reverse or maximum spin angular position of forward from maximum spin angular position forward or backwards, make the shot point light source be entered microscope 19 (not shown)s through the light transmission part 9a of cut-off diaphragm, form lighting point on the sample 16 that is positioned at object lens 15 focal planes.Along with the rotation of scanning galvanometer 8, lighting point moves on sample 16.During battle array detecting device 20 (not shown) end exposure, scanning galvanometer 8 just rotate to reverse or the maximum spin angular position of forward face to face, and sample 16 is once illuminated.
The arrangement of all lighting points on sample 16 is identical with the arrangement of pin hole 6a, is all that isometric matrix is arranged, and moves on sample 16 with identical speed.So in 20 (not shown)s whens exposure of face battle array detecting device, sample 16 is by illumination is once intactly, equably; Perhaps, swing circle by controller 13 (not shown) gated sweep galvanometers 8, make time shutter of face battle array detecting device 20 (not shown)s equal the integral multiple of the swing circle of scanning galvanometer 8, sample 16 when the 20 (not shown)s exposure of face battle array detecting device by repeatedly intactly, illumination equably.
The fluorescence of lighting point excited sample 16 emissions is reflected by dichroic beam splitter 5 through the pin hole 6a of the first reflecting surface 8a, delay lens group 7 and the pinhole array 6 of object lens 15, the light transmission part 9a that ends diaphragm 9, scanning galvanometer 8, then is formed into picture point through the second reflecting surface 8b and the emission color filter 12 of total reflective mirror group 10, imaging lens group 11, scanning galvanometer 8 at sensitive chip 14.Imaging point moves along with scanning galvanometer 8 is rotated on sensitive chip 14, when face battle array detecting device 20 (not shown) end exposure, sensitive chip 14 obtains the complete fluorescence information of sample 16, and is processed, is shown as image on graphoscope 18 (not shown)s by computing machine 17 (not shown)s.The fluorescence of launching due to the sample 16 of the focal plane that only is positioned at object lens 15 could pass through the pin hole 6a of pinhole array 6, reflected by dichroic beam splitter 5 and see through emission color filter 12 arrival sensitive chips 14, so the image that shows on graphoscope 18 (not shown)s is the Confocal Images of sample 16.
Embodiment 2
Fig. 3 is the structural representation of the second confocal optical scanner related to the present invention, specific as follows with the difference of embodiment 1: the first reflecting surface 8a of scanning galvanometer 8, the second reflecting surface 8b, and first total reflective mirror 10 between reflecting surface 8a and pinhole array 6 be all concave reflection, can distinguish delay lens group 7 and 11 pairs of pinhole array 6 imagings of imaging lens group of alternate embodiment 1, the imaging multiplying power of the total reflective mirror 10 between the first reflecting surface 8a and the first reflecting surface 8a and pinhole array 6 is M2.Because reduced delay lens group 7 and imaging lens group 11, have optical interface still less in the light path of the present embodiment, can obtain higher optical efficiency.
Embodiment 3
Fig. 4 is the structural representation of the third confocal optical scanner related to the present invention, and is specific as follows with the difference of embodiment 2: the first reflecting surface 8a of scanning galvanometer 8 and total reflective mirror group 10 are plane reflection for concave reflection, the second reflecting surface 8b.
Embodiment 4
Fig. 5 is the structural representation of the 4th kind of confocal optical scanner related to the present invention, and specific as follows with the difference of embodiment 3: the first reflecting surface 8a and the second reflecting surface 8b, the total reflective mirror group 10 of scanning galvanometer 8 are all concave reflections.
Embodiment 5
Fig. 6 is the structural representation of the 5th kind of confocal optical scanner related to the present invention, specific as follows with the difference of embodiment 4: the first reflecting surface 8a and the second reflecting surface 8b are respectively on two identical scanning galvanometers 8, by controller 13 synchro control rotations, reduced a slice total reflective mirror 10.
Embodiment 6
Fig. 7 is the structural representation of the 6th kind of confocal optical scanner related to the present invention, specific as follows with the difference of embodiment 1: as to have increased the quantity of imaging lens group 11, made the image of sensitive chip 14 records and the imaging of 15 pairs of samples 16 of object lens can realize amplifying in strict 1: 1.