KR100536461B1 - Micro-dissection system - Google Patents

Abstract

The disclosed microdissection system comprises a stage device having a collector stage capable of reciprocating a holder holder with a body disposed on a longitudinal side and a cap inserted into an insertion hole between holder holders; And a correction disposed between the dichroic mirror and the observation unit of the microscope device arranged to pass through the first optical axis of the microscope device and to deflect the second optical axis of the laser beam generated from the laser device to the objective lens side of the microscope device. It has an optical channel with a unit.

Description

Micro-dissection system

The present invention relates to a microscopic ablation system, and more particularly, to a microscopic ablation system for observing tissues such as cells with an optical microscope, collecting certain areas of the observed tissues by ablation using a laser and collecting them. .

In general, the microscopic ablation technique is to extract specific sites such as cells observed under an optical microscope for genetic changes or subsequent studies of DNA, RNA or proteins of specific cells or sites of biological tissues.

Prior art related to a conventional microdissection system is disclosed in US Patent Application No. 09 / 043,093, International Application Publication No. WO 99/39176, International Application Publication No. WO 00/34757, US Patent No. 6,010,888, and the like.

Such a conventional microdissection system is to excise tissue using a laser, and largely classified into two types according to the dissection method. One method uses the heat of the laser and the other method uses the laser as a knife.

Conventional microdissection systems collect a collection to collect the cells or regions to be resected, simply mount the slides, scan the tissue sections placed on the slides in a predetermined path, or horizontally slide the slides to excise the cell sample. There is a need for a stage device for fine movement. The collector is typically installed in a collector holder or tray that is closely associated with the stage device. The collector holder and the stage device may have various structures according to the ablation method or the structure of the collector.

By the way, in most conventional microdissection systems, the excised sample is collected in a cap rather than the body of the collector. Therefore, the conventional microablation system limits the slimming of the stage apparatus because a considerable amount of space is devoted to the collector body for mounting the collector body having a relatively longer length than the cap of the collector.

In addition, the conventional microablation system has a limitation in reducing the distance between the caps of the collector because it employs a plurality of collectors arranged in a row in one system. On the other hand, in order to alleviate these problems, one holder may be used to mount several holders to a stage device using one holder, but such systems are also a limitation on slimming the collector holder and / or the stage device. The long path from tissue sections to the corresponding collection bins can be lengthened to render the collection impossible or to increase collection inefficiency.

On the other hand, when the space between the caps of the collection bin is farther away, it is relatively time consuming for the horizontal movement of the stage device and / or the tray (or stage) and the vertical movement of the stage device during a plurality of collection operations. Problems arise with efficiency.

In addition, the structure of the conventional holder holder has a problem that the task of attaching and detaching the collector to the holder is inconvenient, the conventional stage devices had a problem in stability and accuracy in the movement of the slide, in particular, the arrangement with the holder holder There have been many problems with the interrelationship and space efficiency of.

In general, the microscopic ablation technique requires not only optically precise observation of a tissue sample and a delicate selection of the area to be excised, but also a fine ablation of the selected sample using a laser, thereby making it possible to use slides used in conventional microscopes. Slide structure different from the structure is adopted. That is, the slide structure used in the microscopic ablation technique is a sample that is dyed and dried in a state in which the sample is simply placed on the slide without covering the sample with the cover glass or treating with the encapsulant.

Conventional fine ablation systems use the slide structure described above to observe and ablate the sample, so that fine ablation is achieved with very low optical resolution of the sample. Therefore, lesions with fine changes, such as precancerous lesions, may not be distinguished or identified to a sufficient extent in the microscope field of view.

In addition, known microscopic ablation systems seek to increase the optical resolution described above by using a precision objective lens. However, such a method increases the resolution of the objective lens as well as increases the price of the objective lens.

On the other hand, in the known fine ablation systems, the focus for observation and the focus for ablation are often different from each other, so it is necessary to compensate for the difference between these focuses. In particular, the need for such compensation increases when the spot size and focus of the laser beam are controlled according to the change of the objective lens and the state of the samples. However, the optical channels employed in conventional microdissection systems lack such compensation devices.

In a conventional microdissection system, the stage device can only be operated by a keyboard or mouse connected with a computer. Accordingly, the necessity of a separate manipulator other than a keyboard or a mouse is increased for the convenience of the user in the operation of the microablation system.

In response to these demands, the proposed technique has been developed by Leica for manipulators that can operate X, Y and Z axes separately. The manipulator according to this technique has two rollers installed on one pillar to operate the X and Y axes of the stage apparatus, and a roller for operating the Z axis of the stage apparatus is installed on the other pillar.

Therefore, the Leica manipulator is expensive to manufacture the device, the volume of the device is not only large, there is a problem that the operation of the device is difficult.

SUMMARY OF THE INVENTION The present invention has been made to overcome the above problems, and an object thereof is to provide a microablation system in which a structure capable of compacting a stage device of a microscope device, an accuracy of an optical channel, and the like are easily improved.

The objects of the present invention are specifically listed as follows.

First, an object of the present invention is to provide a microablation system having a slimmer structure so that the space can be efficiently used in response to the structure of the collecting box, as well as to easily attach and detach the collecting box to the collecting holder.

Secondly, an object of the present invention is to provide a fine ablation system in which the collection efficiency and the working efficiency of the entire system are improved by minimizing the gap between the caps in a plurality of collectors arranged in a line.

Third, the present invention is to provide a fine ablation system that can realize the speed and accuracy of fine ablation work by making the stage device slim and compact.

Fourth, the present invention is to provide a microdissection system that can increase the collection rate (collection rate) for the region of cells or tissues excised by reducing the distance between collection from the region to be excised.

Fifthly, the present invention provides a microablation system capable of sufficiently improving the optical resolution of a specimen when observing a sample using the microablation system and compensating for the difference between the focus of the laser beam and the focal point of the microscope field of view. Its purpose is.

Sixth, an object of the present invention is to provide a fine ablation system having a structure that is low in manufacturing cost, easy to operate, and can minimize the volume of the device.

The fine ablation system according to a preferred embodiment of the present invention for achieving the above object is a collection capable of reciprocating between a holder insert and a collector holder having a main body disposed on the side in the longitudinal direction and a cap inserted into the insertion hole. A stage device having an enclosure stage; And a dichroic mirror arranged between the observation unit of the microscope device and the dichroic mirror arranged to be able to pass the first optical axis of the microscope device and to deflect the second optical axis of the laser beam generated from the laser device to the objective lens side of the microscope device. An optical channel with a calibrated unit.

Preferably, the stage device comprises: a base installed in the frame of the meridian device so as to be reciprocated in the Z-axis direction and having a first hole through which the first and second optical axes can pass; A Y stage installed at the base to enable reciprocating movement in the Y axis direction and having a second hole through which the optical axes can pass; And an X stage installed at the Y stage to enable the reciprocating movement in the X axis direction, the X stage having a third hole through which the optical axes can be passed, and having a slide installed therein.

Preferably, the holder insert is a space formed in at least one of the surface of the X stage and the surface of the Y stage with a predetermined width and a predetermined height.

Preferably, the X stage is coupled to an X ball screw operated by an X stepping motor installed in the Y stage and horizontally guided by X guide rails and X guide grooves formed on both sides of the contact surfaces of the X and Y stages. Is reciprocated.

Preferably, the Y stage is coupled to a Y ball screw operated by a Y stepping motor installed in the base, and is horizontally guided by a Y guide rail and a Y guide groove formed on both sides of the contact surface of the Y stage and the base. Is reciprocated.

Preferably the collector stage comprises: a rack coupled with a pinion operated by a collector stepping motor installed in the base, guided by a collector guide rail and a collector guide groove formed in the base and the collector stage. To move horizontally.

Preferably, the holder holder comprises: at least two or more body installation parts alternately provided at both sides of the plate at predetermined intervals so as to install the main body of the plurality of collectors in parallel to each other and both sides of the plate; At least two or more inlet portions each having a predetermined length inlet from each other in the width direction thereof from a side of the plate; At least two inserts formed in the plate to be connected to the inlet so that the cap of each collector may be installed; And a coupling portion coupled to one end of the collector stage.

Preferably, the X stage includes: a receiving groove into which one end of the slide can be inserted; And a slide retainer contacting the other end of the slide to fix the position of the slide.

Preferably, the correction unit includes a concave lens axially aligned with the first optical axis so as to correspond to the objective lens of the microscope device.

Preferably, the correction unit comprises: a plurality of correction lenses axially aligned on the first optical axis so as to correct the characteristics of each objective lens of the microscope device.

Preferably, the correction unit comprises: a correction tray in which the correction lenses are radially installed and rotatably installed at an angle.

Advantageously, the optical channel of said second optical axis comprises: a prism that refracts a laser beam generated from said laser device; Collimators for injecting a parallel beam to the dichroic mirror toward the laser beam emitted from the prism; And a shutter provided between the collimators.

Preferably, the optical channel further comprises a filter provided between the correction unit and the dichroic mirror.

Preferably, the microablation system of the present invention further comprises a manipulator electrically connected to the microscope device for converting the coordinates of the stage device.

Preferably, the manipulator comprises: a pad; A pole installed on the pad; Three rolls installed on the outer circumferential surface of the column to be independently rotatable; A sensing member installed on the pillar to correspond to the individual rolls so as to sense the rotation amount of the rolls; And a control member installed on the pad to control the movement of the stage device of the microablation system by the amount of rotation of the roll sensed by the sensing member.

Preferably, the sensing member includes: modulators each of which is inserted into the column and fixedly rotated on the inner circumferential surface of each of the rolls and each of the plurality of slit balls formed radially equidistantly about the center line of the column; A circuit board installed on an inner wall of the pillar to be connected to the control member; And a pair of photo interrupters installed on the circuit board so as to be disposed on both sides of the modulators.

Preferably, the manipulator further includes a cap installed at the end of the pillar.

Preferably, the manipulator is further provided with a separator installed at one end of the uppermost roll of the three rolls and a portion adjacent thereto, and the other end of the manipulator installed at the pad.

Preferably, the manipulator is a roll inside the separator is an X roll and a Y roll for moving the stage device in the X, Y direction, a single roll outside the separator is Z to move the stage device in the Z direction It's a roll.

In the accompanying detailed description, reference is made to the accompanying drawings, which form a part, and is shown in a manner that describes specific embodiments in which the invention may be practiced. These embodiments are described in sufficient detail to enable those skilled in the art to practice the invention, and it is to be understood that other embodiments may be utilized and structural changes may be made without departing from the scope of the present invention. The following detailed description, therefore, is not to be taken in a limiting sense, and the scope of the present invention is defined by the appended claims and their equivalents. In the description of the present invention, "vertical", "horizontal", "side", "up", "bottom", "rising", "falling", etc., are determined at the position of the microablation system in the context of the drawings and description. It should be regarded as a relative meaning to, and not as an absolute sense of.

The microdissection system according to the invention can in particular be used primarily for cancer research. In other words, the system can be used to determine the presence or absence of any specific gene at each stage of cancer development. In addition, the system is capable of roughly separating normal and tumor cells from the target tissue in order to identify certain tumor-specific genetic changes at the DNA level, thereby roughening the cells by chemical photolysis without heating the biological cells. It is a device that can be cut to a resolution of 1㎛.

Hereinafter, with reference to the accompanying drawings will be described in detail a microdissection system according to a preferred embodiment of the present invention.

1 is a schematic diagram of a microdissection system according to a preferred embodiment of the present invention.

Referring to FIG. 1, the microscopic ablation system 1000 according to a preferred embodiment of the present invention includes a first optical channel including a CCD camera 34 or the like for observing a sample (not shown) on a slide 40. 10) and the optical channel 100 including the second optical channel 20 including the laser device 22 for cutting the selected predetermined area of the sample, and the slide 40 containing the sample and Moving the stage device 200 for moving the slide 40 in the X, Y, and Z directions, and the holder holder 80 (FIG. 2) provided with the collecting box 70 relative to the stage device 200. The collector stage 250 for controlling the horizontal or vertical movement of the stage apparatus 200 or for the laser apparatus 22, the objective lens 18, the collimation lens 21, the microscope apparatus of the optical channel 100. By control unit 300, optical channel 10 for controlling lamps, lenses, etc. An operation including a display unit 400 including a monitor 402 or the like for displaying a predetermined area of the sample, and a keyboard 502, a mouse (not shown), a manipulator 504, etc. for operating the units. The unit 500 is provided.

The microdissection system includes an improved optical channel 100, an improved stage device 200, and a manipulator 504 that can operate the stage device 200 independently of a mouse or keyboard. The optical channel 100 includes the correction unit 60 having a function of increasing the resolution of the fine ablation system during observation and ablation of a sample by supplementing the characteristics of the objective lens 18, and the stage apparatus 200. Has a holder holder and a collector stage to make the system slim and compact.

Hereinafter, the features of the system will be described in detail.

2 is a perspective view illustrating main parts of the stage apparatus 200 illustrated in FIG. 1, FIG. 3 is a plan view of FIG. 2, FIG. 4 is a right side view of FIG. 2, and FIG. 5 is of FIG. 2. It is a front configuration diagram.

2 to 5, the stage device 200 includes a base 210 having a first hole 212 formed therein and movable in a Z direction with respect to the frame 92 of the microscope device of the microdissection system 1000. ), The Y stage 220 provided in the base 210 so that the second hole 222 is formed and reciprocated in the Y direction, and the Y stage so that the third hole 232 is formed and reciprocated in the X direction. Collector stage 250 for reciprocating the X stage 230 installed in the 220 and the holder holder 80 placed in the holder insert 240 provided between the X stage 230 and the Y stage 220. ).

The base 210 has a mounting part 214 installed in the micro ablation system 1000 for vertical reciprocating movement, and a bracket 216 on which the collecting stage 250 is installed. A condenser is installed below the base 210.

The mounting portion 214 is coupled to the frame 92 of the microscope device by a slider device (not shown) used to move a sample in a conventional microscope device. The bracket 216 is for installing a collector stepping motor assembly for reciprocating the collector stage 250 in the X direction.

In the mounting portion 214 of the base 210, a Z shaft (not shown) used for the base 210 of a general microscope device is installed therethrough, and one end of the Z shaft is a Z stepping motor 218 installed in the frame 92. (See Fig. 1). The other end of the Z shaft is provided with a Z knob 211 that can manually raise and lower the base 210. The Z knob is similar to that of a conventional microscope device. The Z stepping motor 218 is responsible for the focusing of the first optical channel 10 for sample observation of the microablation system 1000 and the laser focusing function of 20 of the second optical channel for ablation of the selected sample region. do.

The first hole 212 penetrates the base 210 so as to have a size that does not interfere with the optical axis of the microablation system 1000 even when the X and Y stages 210 and 220 are moved to the maximum and minimum. Is formed.

The Y stage 220 is for moving the slide 40 in the Y direction for observation and ablation of the cells on the slide 40.

The Y stage 220 may have a substantially rectangular shape having a second hole 222 penetrated in the center thereof. The Y stage 220 is coupled to a Y ball screw 226 operated by a Y stepping motor 224 installed in the Y stepping motor mounting portion of the base 210. In order for the Y stage 220 to reciprocate in the Y direction from the top of the base 210, one side of the Y stage 220 is coupled to the Y ball screw 226, the Y guide on both sides of the base 210 Rails 228 are installed, and Y guide grooves 221 coupled to the Y guide rails 228 are formed at both bottom surfaces of the Y stage 220. Of course, the Y guide rail 228 and the Y guide groove 221 may be provided in the Y stage 220 and the base 210, respectively.

The second hole 222 formed in the Y stage 220 may have a fine ablation system 1000 even if the Y stage 220 moves to the maximum or the X stage 230 moves to the maximum and minimum with respect to the base 210. It is desirable to have a size such that it does not cause interference to the optical axis of.

The upper surface of the Y stage 220 is preferably a portion of the holder inserting portion 240 into which the holder holder 80 of the collector stage 250 can be inserted. Of course, when the holder insertion unit 240 is formed at a sufficient height on the lower surface of the X stage 230 to be described later, the upper surface of the Y stage 220 may be kept flat.

One side of the Y stage 220 is preferably provided with an X stepping motor installation unit on which the X stepping motor 234 may be installed.

The X stage 230 basically stores the slide 40 and simultaneously moves the slide 40 in the X direction for observation and ablation of cells on the slide 40.

Preferably, the X stage 230 has a substantially rectangular shape in which a third hole 232 is formed at the center thereof. The X stage 230 is coupled to an X ball screw 236 operated by an X stepping motor 234 installed in the Y stage 220. In order for the X stage 230 to reciprocate in the X direction from the top of the Y stage 220, one side of the X stage 230 is coupled to the X ball screw 236, the upper surface of both sides of the Y stage 220 The X guide rails 238 are installed, and X guide grooves 231 coupled to the X guide rails 238 are formed at both bottom surfaces of the X stage 230. Of course, the X guide rail 238 and the X guide groove 231 may be provided in the X stage 230 and the Y stage 220, respectively.

The third hole 232 formed in the X stage 230 may move the maximum X stage 230 with respect to the Y stage 220 or the maximum maximum and minimum Y stage 220 with respect to the base 210. Even if the size of the fine ablation system 1000 does not interfere with the optical axis.

A portion of the holder inserting portion 240 into which the holder holder 80 of the collector stage 250 may be inserted is formed on the bottom surface of the X stage 230. Of course, when the holder insertion unit 240 is formed at a sufficient height on the upper surface of the above-described Y stage 220, the lower surface of the X stage 230 may be kept flat.

The X stage 230 is provided with a member for fixing the position of the slide 40 across the third hole 232. To this end, one end of the third hole 232 is provided with an accommodating groove 233 into which one end of the slide 40 can be inserted and received, and the other end of the X hole may be selectively contacted with the slide 40. A slide retainer 235 is installed to slide on the stage 230.

The slide retainer 235 includes a retainer plate 235a inserted into a retainer groove 237 formed at one side of the X stage 230, and a retainer knob 236b installed through the retainer plate 235a. The retainer plate 235a has a position fixing groove 235c that can be engaged with one side of the slide 40, preferably the corner of the slide 40.

The collector stage 250 is for reciprocating the collector holder 80 located in the holder inserter 240 in the X direction. The collector holder 80 is installed so that the plurality of collectors 70 are compact and slim, which will be described later.

The collector stage 250 is moved by a collector stepping motor 252 installed on the base 210. To this end, a pinion 254 is installed on the shaft of the collector stepping motor 252, which pinion 254 is coupled with a rack 256 installed in the body of the collector stage 250. Meanwhile, a collector guide rail 258 is installed at the base 210, and a collector guide groove 251 is formed at the collector stage 250 main body to be fitted to the collector guide rail 258. The end of the main body of the collector stage 250 is provided with a connector 253 to which the coupling portion 94 of the collector holder 80 can be coupled.

The X, Y, Z, and Collector Stepping Motors 214, 224, 234, 252 are each 2 mm lead / precision grade ground to have specifications of 25000 to 50000 step / rev (6 nm / step). A lead / precision grade ground ball-screw is used. In addition, the X, Y, Z, and collector box stepping motors 214, 224, 234, 252 are each managed by a pair of optical limit switches 94. The accuracy of the position control of the X, Y, Z, and collector box stepping motors 214, 224, 234, 252 is approximately ± 8 μm, and the repeatability is approximately ± 0.5 μm. The movement speeds and movement accelerations generated by the X, Y, Z, and collector box stepping motors 214, 224, 234 and 252 are 20000-40000 step / sec and 20000-96000 step / sec ^{2, respectively.} to be.

The travel intervals of the X, Y, and collector stages 214, 224, 234, 252 are preferably 60 mm, 40 mm, 75 mm, respectively.

6 is a plan view of the holder holder, Figure 7 is a perspective view showing a state in which the collector is coupled to the holder holder.

6 and 7, in the microdissection system 1000 according to a preferred embodiment of the present invention, the collector holder 80 has a tubular collector body having a predetermined length with one end closed and the other end open. 72, a collector cap 74 capable of closing the open end of the collector body 72, and a connecting ring 76 connecting the collector body 72 and the collector cap 74; It is for storing a plurality of collection boxes (70) compactly.

The holder holder 80 is a plate so as to communicate with the main body mounting portion 84 provided on both sides of the plate, an inlet portion 86 formed in the width direction center of the plate from the side of the plate, and the inlet portion 86 It is provided with an insertion portion 88 formed through.

The main body mounting portion 84 is disposed sequentially on one side of the plate to collect the first and second mounting portions (84a) (84b) and the other side of the plate, each having an elastic piece that can be elastically coupled to the main body The collector body 72 may be arranged in sequence so as to be elastically coupled, and the first and second installation units 84a and 84b may be provided with third and fourth installation units 84c and 84d disposed in a zigzag form. do.

The inlet 86 is preferably formed so as to penetrate in the width direction from the side of the plate so that the connecting ring 76 of the collecting box 70 does not interfere. The inlet 86 is for preventing the connecting ring 76 from interfering with the plate when the collection box 70 is mounted or detached to the collection holder 80. The inlet portions 86 are sequentially formed to correspond to the first to fourth installation portions 84a, 84b, 84c and 84d, and are arranged in a zigzag shape to form the first to fourth inlets 86a and 86b. 86c and 86d.

The inserting portion 88 has a shape in which the collecting cap 74 is elastically inserted to be fixed in position, and the first to fourth insertion holes 88a disposed in close contact with each other at minimum intervals in the longitudinal direction of the plate ( 88b) 88c and 88d.

Coupling portion 82 provided at the other end of the plate is not provided with the collecting box 70 is coupled to the connector 253 of the collecting stage 250 described above, the coupling guide 82a and the fixed plate A fixing knob 82b for fixing the position to the connector 253 is provided.

8 is a plan view of the holder holder 180 for the microdissection system according to another embodiment of the present invention, Figure 9 is a perspective view showing a state in which the collector is coupled to the holder holder 180. The same components as those described in FIGS. 6 and 7 are the same members with the same functions.

8 and 9, the holder holder 180 is for compactly storing a plurality of holders 70 in the same manner as the holder holder 80 described above.

The holder holder 180 is in communication with the main body mounting portion 184 provided on both sides of the plate, the inlet portion 186 formed in the width direction center of the plate from the side of the plate, and the inlet portion 186 The insertion part 188 is formed through.

The main body mounting portion 184 is the same as the main body mounting portion 184 of the above-described embodiment.

Unlike the inlet portion 86 of the above-described embodiment, the inlet portion 186 does not penetrate in the width direction of the plate, and an inlet groove is formed to have a predetermined depth of the plate. The retracting groove is provided with a connecting ring 76 of the collecting box 70 to prevent the connecting ring 76 from interfering with the other side of the plate.

The insertion part 188 has an insertion slit provided at the edge of the first through fourth insertion holes 188a, 188b, 188c and 188d so that the cap protrusion provided on the side of the collecting cap 74 is fitted. do. That is, the collection caps 74 are positioned in the insertion holes 188a-188d as a whole, and have a configuration in which the cap projection is fitted into the insertion slit.

10 is a schematic diagram showing an arrangement of optical channels employed in the microdissection system according to a preferred embodiment of the present invention.

Referring to FIG. 10, the optical channel 100 employed in the microablation system 1000 includes a first optical channel 10 and a second optical channel 20. The first optical channel 10 forms a first optical axis 12 of a microscope device (not shown), and the second optical channel 20 is a second optical axis of the laser beam generated from the laser device 22 ( 24).

The first optical channel 10 is an optical of a conventional microscope device capable of observing a sample (not shown) on the slide 40 using the eyepiece 32 or the CCD camera 34 of the observation unit 30. As a channel, it is formed by the combination of the eyepiece prism 14, the tube lens 16, the objective lens 18, the capacitor 11, and the lamp 13. The first optical axis 12 refers to an image observation path.

The second optical channel 20 is a movement path of the laser beam for cutting a predetermined region of the specimen, and the movement path is represented by the second optical axis 24. The second optical channel 20 has a prism 26, a pair of collimators 28, 21 and a shutter 23.

As shown in FIGS. 10 and 11, the second optical channel 20 has a substantially closed channel box 25 so that a predetermined optical axis can be stably formed even if the intensity of the laser beam is changed as necessary. It is preferably installed in.

The laser device 22 is designed to have a spot size of 1 μm or less, and the power of the laser beam generated therefrom is designed to be adjustable according to a user's request. The laser device 22 has a solid state laser with a wavelength of 355 nm, the specification having 20 uJ and a frequency of 50 to 10000 Hz.

One end of the channel box 25 is formed with a beam inlet 27 so that the laser beam generated from the laser device 22 can be incident. First and second ports 29a and 29b are provided at the other end of the channel box 25 so that the first optical axis 12 may be formed. The first port 29a is a portion connected to the observation unit 30, and the second port 29b is a portion connected to the objective lens 18.

10 and 11, the prism 26 is capable of refracting a laser beam generated from a laser device 22 such as an ultraviolet laser in the direction of the collimator 21, 28. ) Is installed inside.

The pair of collimators 21 and 28 has the function of helping the microablation system to achieve a predetermined spot size by ultimately forming the laser beams passing through the prism 26 in parallel. In other words, if the collimators 21 and 28 are incorrectly determined or arranged, the system may not only obtain a parallel laser beam, but also may result in improper focus even if it cannot or cannot be accurately focused. The collimator is divided into an eyepiece collimator 28 in the form of a concave lens and an objective collimator 21 in the form of a convex lens.

The collimators 21 and 28 are respectively installed at one end of the barrel having a predetermined length. The eyepiece collimator 28 has a concave lens shape in which both surfaces are substantially symmetrical. The object collimator 21 is arranged so that the concave lens 21a provided to face the shutter 23 and the convex lens 21b provided to face the dichroic mirror 50 to be described later are adjacent to each other, and the concave lens 21a is disposed. When the convex lens 21b is arranged as a whole, it is preferable to exhibit the characteristics of the convex lens.

The shutter 23 is installed between the eyepiece collimator 28 and the objective collimator 21, it is preferable to have a structure that is opened when cutting the sample, otherwise closed.

The dichroic mirror 50 is disposed on the first optical axis 10 in the vicinity where the second optical axis 20 is refracted. The dichroic mirror 50 passes the first optical axis 10 in a straight line and refracts the second optical axis 20. That is, the dichroic mirror 50 transmits the image of the sample observed through the objective lens 18 of the microscope device to the observation unit 30 side, and transmits the laser beam incident from the laser device 22 to the objective lens 18. In the direction of. The dichroic mirror 50 is a flat mirror coated with a transparent multilayer thin film. When the incident angle of light is 45 °, light of a certain wavelength range is reflected by the interference effect of light in the thin film, and the other is It has the property of permeation. The dichroic mirror 50 can have a flat plate shape and a prism shape, but in this embodiment, a flat plate shape is used. Of course, the thickness or the number of layers of the film can be adjusted, and part of the visible light can be freely selected and used depending on the material.

10 and 11, a correction unit 60 having a concave lens characteristic is disposed between the dichroic mirror 50 and the observation unit 30, and between the correction unit 60 and the dichroic mirror 50. The filter 70 is installed.

The correction unit 60 has a function of increasing the resolution of the fine ablation system during observation and ablation of the sample by supplementing the characteristics of the objective lens 18. In addition, the correction unit 60 is for correcting the difference between the focus of the sample recognized by the CCD camera 34 or the eyepiece 32 and the focus when the predetermined area of the sample is ablated using the laser beam. . By this correction unit 60, the spot size of the laser beam of the fine ablation system can be kept constant regardless of the power change of the laser beam. In particular, even when the laser beam power is increased to speed up the ablation operation, the spot size of the laser beam can be kept constant by the correction unit 60.

The filter 70 blocks the heat of reflection of the laser beam passing through the dichroic mirror 50 and ultimately prevents the observation unit 30, that is, the eyepiece 32 and the CCD camera 34 from being damaged. Has The filter 70 uses a flat mirror or plastic, and is disposed to be inclined at a predetermined angle with the first optical axis 10.

It is preferable that the correction unit 60 uses a concave correction lens having a characteristic capable of correcting the resolution in response to characteristics such as magnification of the objective lens 18 of the microscope device. The correcting lens 62 of the correcting unit 60 is axially aligned with the first optical axis 10. The correction lens 62 has a flat portion facing the dichroic mirror 50 and a portion facing the observation unit 30 has a concave shape.

13 and 14, in the optical channel, the correction unit 60 includes a plurality of correction lenses capable of correcting each objective lens according to the characteristics of the plurality of objective lenses installed in the microscope device. 62 is provided. Each of the correction lenses 62 needs to be aligned with the first optical axis 10 so as to correspond to the corresponding objective lens. To this end, the correction unit 60 includes a correction tray 64 in which a plurality of correction lenses 62 are radially installed and rotatably installed at a predetermined angle.

The correction tray 64 is installed on the rotation shaft 66 rotatably installed in the channel box 25. The gear 64a is formed in the outer peripheral surface of the correction tray 64, and this gear 64a meshes with the gear 68a provided in the rotating shaft of the motor 68. As shown in FIG. Of course, the correction tray may be configured to be directly connected to the rotating shaft of the motor. The motor 68 is controlled by a control unit, not shown, and is capable of forward and reverse rotation.

Six correction lenses 62 are arranged at the 60 degree angle in the correction tray 64. Therefore, in the fine ablation system, when the magnification change is generated by the change of the objective lenses, the correction lens 62 corresponding to the objective lens is rotated by the rotation of the correction tray 64 rotated by the motor 68. It can be located on the optical axis 10, so that the correction lens 62 can correct the objective lens 18 during operation of the system.

FIG. 15 is a perspective view schematically illustrating the manipulator shown in FIG. 1, and FIG. 16 is a partial cutaway view of FIG. 15.

As shown in FIGS. 15 and 16, the manipulator 504 includes a pad 510 that can be placed on a desk (not shown), a hollow pole 520 installed on the pad 510, and a pillar. Three rolls 530 installed on the outer circumferential surface of the 520 and the rotation of the sensing member 540 and the rolls 530 installed on the pillar 520 to detect the amount of rotation of the respective rolls 530. The control member 550 installed in the pad 510, the separator 560 separating the some rolls 530 from each other, and the cap 570 provided at the end of the pillar 520 so as to control the movement of the stage apparatus according to the quantity. ).

The pad 510 has a flat case structure in which a control member 550 connected to power applied from the outside is embedded. The pad 510 is provided on the upper surface of the pillar 520 installation hole 512 in which the pillar 520 may be installed. The top cover of the pad 510 is provided with lamps 514 for displaying the power on, off and various operating states, and buttons 516 for other operations of the stage apparatus. The lamps 514 and the buttons 516 are driven by the control member 550 installed inside the pad 510.

The pillar 520 is installed on the pad 510 and protrudes around the insertion portion 522 and the insertion portion 522 inserted into the installation hole 512 of the pad 510 so as to pad the pillar 520 to the pad 510. ) Is provided with a support portion 524. Screw support holes 526 are formed in the support part 524. The screw coupling hole 526 is for fixing the pillar 520 to the pad 510 by a bracket 528 positioned below the top cover of the pad 510.

The rolls 530 are provided with a Z-axis roll 530a, a Y-axis roll 530b, and an X-axis roll 530c in order from the top to the bottom of the drawing, respectively. Of course, the order may be modified as many as necessary.

The rolls 530 are installed to rotate around the column 520 by bearings 532, respectively. Each of the rolls 530 is rotated independently of each other by being positioned at the bottom of the bearing 532 and separated from each other by the support 534 fixed to the pillar 520. On the outer circumferential surface of each roll 530, slits of a predetermined pattern are formed to facilitate rotation of the roll 530. On the inner circumferential surface of each roll 530 the inner wall of the roll 530 so that the inlet 536 and the bearing 532 introduced into the inner wall of the roll 530 may be installed so that the modulator 542 to be described later is installed. And a bearing mounting portion 538 protruding therefrom. Reference numeral 531 in the drawing represents a spacer for the installation space of the separator 560.

The sensing member 540 includes a modulator 542 fixed to an inlet 536 provided on an inner wall of the corresponding roll 530, and a circuit board installed on an inner wall of the pillar 520 so as to be connected to the control member 550. 544 and a pair of photointerrupters 546 electrically connected to the circuit board 544 so as to be disposed on both sides of the modulator 542, respectively.

The sensing member 540 detects whether light is blocked by interference of the modulator 542 and generates a signal for controlling coordinates of the stage device according to the waveform.

As shown in FIG. 17, the modulator 542 is a type of encoder, and each modulator 542 is preferably annular. The outer circumferential surface of each modulator 542 is fixed to the inlet portion 536 of the corresponding roll 530, and a through hole 542b is formed at the center of the modulator 542, which is located close to the outer circumferential surface of the pillar 520. . The body of the modulator 542 is provided with a plurality of slit holes 542a formed at radial equal intervals with respect to the center line of the pillar 520. In the optical interference of a pair of photo interrupters 546 adjacent to each other, the slit holes 542a are preferably spaced apart from each other so as to maintain a phase difference of 90 degrees. In other words, the slit hole 542a of the modulator 542 rotates between the pair of photo interrupters 546 while the adjacent photo interrupters 546 alternately interfere with light while maintaining a phase difference of 90 degrees.

As shown in FIG. 18, each photointerrupter 546 may include a light emitting unit 546a such as a light diode capable of emitting light such as an infrared frequency and a photo transistor for receiving an infrared frequency. and a light receiving portion 546b such as a transistor). Reference numeral 546c in FIG. 16 denotes a connection portion electrically connected to the circuit board 544, and reference numeral 546d is a space in which the modulator 542 can be located.

When the light passes through the slit hole 542a, the photo interrupter 546 does not generate light interference. When the light does not pass through the plate between the slit holes 542a, light interference occurs, thereby causing a modulator ( According to the rotation operation of the 542, the waveform of the signal received by the light receiver 546b may be digitized. The pair of light emitting portions 546a of the photo interrupter 546 are arranged to have a phase difference of 90 degrees with each other.

On the other hand, the sensing member may be of any known technology similar to that can be employed to sense the rotation amount of the roll in addition to the optical interference technology such as the modulator and the photo interrupter.

15 and 16, the circuit board 544 is used to control the photo interrupter 546, and an amplifier for amplifying a current impulse value due to the rotation of the modulator 542 into which the light receiving unit 546b flows. (Not shown) and the like are arranged.

The control member 550 determines the sum of the current and its current impulse values and the type of the current by the rotation of the modulator 542, and transmits these values to the stage controller of the control unit (not shown) through the RS-232 interface. Send or control the stage device directly. That is, the current impulse increases or decreases by the rotation of the modulator 542, and the coordinates of the stage device can be moved when the value of the impulse is determined by the increase or decrease.

On the other hand, the control member 550 may set different coordinate movement displacements of the stage apparatus per unit rotation of the roll (modulator) according to the request of the consumer through a setting interface.

The separator 560 is for distinguishing the Z-axis roll 530a and the Y and X-axis rolls 530b and 53c, and one end thereof is installed between the Z-axis roll 530a and the Y-axis roll 530b. The other end is installed in the pad 510.

The separator 560 is bent from the inclined plate 562 to be arranged in parallel with the bottom surface of the inclined plate 562 and the Z-axis roll 530a provided at the position where the X, Y axis rolls 530b and 530c are positioned. A flat plate 564 is provided. The shape of the separator 560 is ergonomically designed to facilitate the user's finger movement and to distinguish the vertical plane from the X and Y planes when the coordinates of the stage device are converted.

The cap 570 is preferably installed at the end of the pillar 520 in the interference fit method. The diameter of the cap 570 is made substantially the same as that of the roll.

The microscopic ablation system according to an exemplary embodiment of the present invention may store not only a real-time image mode expressed in the display unit but also a pre-scanned image or, if necessary, immediately search for an image region of interest using a road map image, and search for the image region of interest. By selecting an ablation zone for the captured image and storing such data values, tele-consultation is possible through on-line or the like.

Referring to the operation of the microdissection system according to a preferred embodiment of the present invention.

First, in order to observe and ablate a cell region using the microdissection system 1000, the slide 40 is fixed to the storage groove of the X stage 230 of the stage apparatus for the microdissection system 1000, and the slide 40 is removed. Set to the initial position.

Next, the collector holder 80, in which the collectors 70 are installed, is connected to the collector stage 250 to prepare the collector stage 250 to move in the X direction.

The user then inputs inputs such as the maximum resolution of the CCD camera, the magnification of the objective lens, the size of the cell tissue, the selection of the scanning area and the ablation area, and the like.

Next, using the microdissection system 1000 to move the stage device 200 for observation of the cells on the slide 40 is mainly to move the X stage 230 and Y stage 220 in the X, Y direction, respectively. In operation, since the X, Y stages 210 and 220 are driven independently, the X and Y stages 210 and 220 can be operated simultaneously so that the X and Y stages 210 and 220 can move simultaneously.

The movement of the Y stage 220 relative to the base 210 causes the X stage 230 to also move in the Y direction, and as a result, the slide 40 moves. Then, the observation area displayed on the monitor through the optical channel 100 of the microablation system 1000 is also moved. However, the movement of the X stage 230 relative to the Y stage 220 only moves the X stage 230 itself.

In the above-described observation process, the control unit combines the image of the local area obtained by the CCD camera to form a mosaic-like overall image and expresses it on the display unit. The overall image is made such that the entire image is connected without interruption on a reduced scale. The reduced accumulation is automatically calculated so that the overall image is displayed properly on the monitor screen. Since the comprehensive image file for searching is stored in, for example, a jpg image file in the computer memory, it can be repeatedly read. That is, the user can open a search file and bring up a screen by using a mouse. At this time, the user may select only a part of the screen for detailed observation. The selected part is displayed as a navigation picture in the view window, which is automatically calculated according to the size of the computer monitor. A specific position of the view window, for example, an upper left corner point, may be set as a starting point for the search module for the overall image.

The search process for the general image uses a zoom function for detailed observation of the cell image. The picture file number of the image selected by the user is determined according to the view window coordinates indicated in the mosaic image construction process. If the zoom button is pressed in this state, the accumulation can be enlarged up to 8: 1.

Meanwhile, the image may be moved in a desired direction by dragging the image through a mouse operation, or the image may be moved in a desired direction by manipulating the manipulator. This moving operation is possible by the X and Y stages of the stage device.

In this process, adjacent video files (videos next to the selected part) are automatically stored in the RAM of the computer main body during the movement, and appear on the monitor screen with a predetermined selected amount. Cleared.

Next, in order to ablate such an ablation zone with the ablation zone determined through observation, the collector stage is moved to position the collector on which the cell region to be excised is collected on the optical axis. Then, the X, Y stages 210 and 220 are cut by the control unit while moving according to predetermined coordinates. In this case, when the thickness of the tissue is thick, it may be adjusted to cut the tissue cell at least twice.

The user can operate the mouse to select the required collection box. In addition, the user may draw the laser cutting path required for the image displayed on the monitor screen using a mouse point.

The control unit may automatically assign a color to indicate the cutting path according to the type of cell sample, upon selection of the collection. The collector stage 250 causes the collector stage 250 to reciprocate in the X direction to locate the collector where the cell region to be excised by the control unit is determined and will collect the excised cell portion.

On the other hand, the cutting path is displayed along with the cell image according to the accumulation selected by the user on the monitor screen, the coordinates of the cutting path may be stored in the computer memory.

The microdissection system according to the present invention has the following effects.

First, the plate of the collector holder can be made thin by ideally disposing the cap, the connecting ring and the body of the collector on the plate of the collector holder. Therefore, the space required can be minimized, and the collectors can be mounted stably in the collector holder.

Second, since the gap between the caps of the collectors installed in the collector holder through the above configuration can be minimized, the movement interval of the collector stage can be minimized, thereby increasing the workability of the microdissection system.

Third, according to the slimming of the holder holder and the stage device, the path from the ablation zone to the corresponding bin for the tissue sections is shortened, thereby increasing the collection efficiency of the tissue sections.

Fourth, the optical resolution of the system can be improved by a correction unit including a concave lens-type correcting lens capable of correcting the characteristics of the objective lens, so that microscopic observation of tissue samples and precise ablation of microscopic areas are possible.

Fifth, by introducing the correction unit, the difference between the focus of the laser beam required for ablation of the sample and the focus required for observation of the sample can be corrected, so that the spot size of the laser beam emitted from the laser device can be kept constant. .

Sixth, since the stage device can be moved separately from the keyboard or mouse, the microablation system can be conveniently operated.

Seventh, since the X, Y, Z-axis operation according to the coordinate movement of the stage device can be integrated and managed in one axis (pillar), the convenience of operation is increased.

Eighth, by installing three rolls adjacent to one pillar, it is possible to reduce the assembly cost of the device and at the same time reduce the size of the device.

It is to be understood that the above detailed description is intended to be illustrative, and not restrictive. Many other embodiments will be apparent to those skilled in the art upon reading or understanding the above detailed description. It should be noted that the embodiments discussed in other parts of the detailed description and discussed with reference to other figures may be combined to form additional embodiments of the present invention. Accordingly, the scope of the invention should be determined with reference to the appended claims, along with the full scope of equivalents to which such claims are entitled.

The following drawings attached to this specification are illustrative of preferred embodiments of the present invention, and together with the detailed description of the invention to serve to further understand the technical spirit of the present invention, the present invention is a matter described in such drawings It should not be construed as limited to.

1 is a schematic structural diagram of a microdissection system according to a preferred embodiment of the present invention.

Figure 2 is a perspective view showing the main portion of the stage apparatus shown in FIG.

3 is a plan view of FIG. 2.

4 is a configuration of the right side of FIG.

5 is a front configuration diagram of FIG. 2.

6 is a plan view of a collector holder for a microdissection system according to a preferred embodiment of the present invention.

Figure 7 is a perspective view showing a state in which the collector is coupled to the holder holder.

8 is a plan view of a collector holder for a microdissection system according to another embodiment of the present invention.

FIG. 9 is a perspective view illustrating a state in which a collector is coupled to the holder of FIG. 8; FIG.

10 is a schematic structural diagram of an optical channel of the fine ablation system shown in FIG.

FIG. 11 is a cross-sectional view of the channel box in which the optical channel and correction unit portion of FIG.

12 is a plan view of FIG.

FIG. 13 is an enlarged plan view of a portion of the correction unit of FIGS. 11 and 12.

14 is a cross-sectional view taken along the line A-A of FIG.

15 is a perspective view schematically showing the manipulator shown in FIG.

16 is a partial cutaway view of FIG. 15.

FIG. 17 is a plan view of the modulator shown in FIG. 15. FIG.

18 is a side configuration diagram of the photo interrupter shown in FIG. 15.

<Explanation of symbols for main parts of the drawings>

10 first optical channel 12 first optical axis 14 eyepiece prism

18 ... objective lens 20 ... second optical channel 22 ... laser unit

24 ... 2nd optical axis 25 ... Channel box 26 ... Prism

21, 28 ... collimator 30 ... observation unit 32 ... eyepiece

34 ... CCD camera 40 ... Slide 50 ... Differential mirror

60 ... calibration unit 62 ... compensation lens 64 ... compensation tray

66 ... rotation shaft 68 ... motor 70 ... collection box

72 ... collection body 74 ... collection cap 80 ... collection holder

82 Joining part 84 Main body mounting part 86

88 Insert ... 200 Stage unit 210 Base

220 ... Y stage 230 ... X stage 240 ... holder insert

250 Collector Stage 504 Manipulator 510 Pad

520 posts 530 rolls 540 sensing element

542 Modulator 544 Circuit Board 546 Photointerrupter

550 ... Control member 560 ... Separator 570 ... Cap

Claims (19)

A stage device having a collector stage capable of reciprocating between a holder insert and a collector holder having a main body disposed on the side in the longitudinal direction and a cap inserted into the insertion hole; And Disposed between the dichroic mirror and the observation unit of the microscope device arranged to be able to pass the first optical axis of the microscope device and to deflect the second optical axis of the laser beam generated from the laser device to the objective lens side of the microscope device A microablation system comprising an optical channel with a calibration unit. The method of claim 1, The stage device is: A base which is installed in the frame of the apparatus for reciprocating in the Z-axis direction and has a first hole through which the first and second optical axes can pass; A Y stage installed at the base to enable reciprocating movement in the Y axis direction and having a second hole through which the optical axes can pass; And an X stage installed at the Y stage to enable reciprocating movement in the X axis direction, and an X stage having a third hole through which the optical axes can pass and a slide installed therein. The method of claim 2, And the holder insertion portion is a space formed in at least one of a surface of the X stage and a surface of the Y stage at a predetermined width and a predetermined height. The method of claim 2, The X stage is: It is coupled to an X ball screw operated by an X stepping motor installed in the Y stage, and reciprocating horizontally while being guided by X guide rails and X guide grooves formed on both sides of the contact surfaces of the X and Y stages. Fine ablation system. The method of claim 2, The Y stage is: It is coupled to the Y ball screw operated by the Y stepping motor installed in the base, and is reciprocated horizontally while being guided by the Y guide rail and the Y guide groove formed on both sides of the contact surface of the Y stage and the base. Fine ablation system. The method of claim 2, The collector stage is: And a rack coupled to a pinion operated by a collector stepping motor installed in the base, and moved horizontally while being guided by a collector guide rail and a collector guide groove formed in the base and the collector stage. Fine ablation system. The method of claim 1, The collector holder is: At least two main body mounting portions alternately provided at both sides of the plate at predetermined intervals so that the main bodies of the plurality of collecting boxes are arranged side by side and alternately with each other on both sides of the plate; At least two or more inlet portions each having a predetermined length inlet from each other in the width direction thereof from a side of the plate; At least two inserts formed in the plate to be connected to the inlet so that the cap of each collector may be installed; And And a coupling portion capable of coupling to one end of the collector stage. The method of claim 2, The X stage is: An accommodation groove into which one end of the slide can be inserted and received; And And a slide retainer in contact with the other end of the slide to fix the position of the slide. The method of claim 1, The correction unit And a concave lens axially aligned with the first optical axis so as to correspond to the objective lens of the microscope device. The method of claim 1, The calibration unit is: And a plurality of correction lenses axially arranged on the first optical axis so as to correct characteristics of each objective lens of the microscope device. The method of claim 10, The calibration unit is: And a correction tray in which the correction lenses are radially installed and rotatably installed at a predetermined angle. The method of claim 1, The optical channel of the second optical axis is: A prism that refracts a laser beam generated from the laser device; Collimators for injecting a parallel beam to the dichroic mirror toward the laser beam emitted from the prism; And And a shutter installed between the collimators. The method of claim 1, And a filter provided between the correction unit and the dichroic mirror. The method of claim 1, And a manipulator electrically connected to the microscope device for converting coordinates of the stage device. The method of claim 14, The manipulator is: pad; A pole installed on the pad; Three rolls installed on the outer circumferential surface of the column to be independently rotatable; A sensing member installed on the pillar to correspond to the individual rolls so as to sense the rotation amount of the rolls; And And a control member provided on the pad to control the movement of the stage device of the fine ablation system by the amount of rotation of the roll sensed by the sensing member. The method of claim 15, The sensing member is: Modulators each of which is inserted into the pillar and is fixedly fixed to the inner circumferential surfaces of the rolls and has a plurality of slit balls radially equidistant with respect to the center line of the pillar; A circuit board installed on an inner wall of the pillar to be connected to the control member; And And a pair of photo interrupters mounted on the circuit board so as to be disposed on both sides of the modulators. The method according to claim 15 or 16, Fine cutting system characterized in that it further comprises a cap installed at the end of the pillar. The method according to claim 15 or 16, One of the three rolls of the three rolls and the adjacent portion is installed on one end and the other end of the fine ablation system, characterized in that further comprising a separator installed on the pad. The method of claim 18, The rolls inside the separator are X rolls and Y rolls for moving the stage device in the X and Y directions, and the single roll outside the separator is a Z roll for moving the stage device in the Z direction. Ablation system.