Classifications

• • G—PHYSICS
  • G02—OPTICS
  • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
  • G02B21/00—Microscopes
  • G02B21/36—Microscopes arranged for photographic purposes or projection purposes or digital imaging or video purposes including associated control and data processing arrangements
  • G02B21/365—Control or image processing arrangements for digital or video microscopes
• • G—PHYSICS
  • G02—OPTICS
  • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
  • G02B21/00—Microscopes
  • G02B21/0004—Microscopes specially adapted for specific applications
  • G02B21/002—Scanning microscopes
• • G—PHYSICS
  • G02—OPTICS
  • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
  • G02B21/00—Microscopes
  • G02B21/24—Base structure
  • G02B21/26—Stages; Adjusting means therefor
• • G—PHYSICS
  • G02—OPTICS
  • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
  • G02B21/00—Microscopes
  • G02B21/34—Microscope slides, e.g. mounting specimens on microscope slides

Definitions

• the present invention relates to instrumentation in the field of microscopy. More particularly, the present invention relates to the production of a microscope intended to be used as an observation and measurement instrument.
• the microscope is used in many applications both for observation and for measurement by analysis of microscopic images in biology and medicine. Since its conception, the microscope has essentially evolved in terms of the quality of the optics, allowing an improvement in the depth of field, the size of the field observed and the correction of lens errors. However, it remains dedicated to direct observation by an observer.
• Such a localized and sedentary nature of microscopy is further accentuated by an initial step of detecting the location of the objects to be observed within the sample. This preliminary step requires the exploration of the entire available sample and cannot be done remotely.
• the images obtained are images preselected by an observer as part of a specific therapeutic or diagnostic approach. Carrying out a new approach often requires a new analysis of the sample.
• the present invention therefore aims to provide a microscope which overcomes the problems described above.
• the present invention aims to propose such a microscope which provides images of optimal and defined quality.
• the present invention also aims to propose such a microscope which provides reproducible images.
• the present invention also aims to propose such a microscope which provides complete and / or partial images of a preparation.
• the present invention also aims to propose such a microscope which can be used both as an observation device and as a measuring instrument.
• the present invention also aims to propose such a microscope, the adjustments of which are carried out either autonomously or on command of the observer.
• the present invention also aims to propose such a microscope suitable for being used remotely independently or controlled.
• the present invention provides a microscope comprising at least one optical device for magnifying a preparation slide placed on a rectangular slot so that the entire length of the slide at least partially covers the length of the slot, the width of the slot being less than that of the blade, further comprising: means for moving the optical device in the direction of the length of the slot over this entire length; a blade holder for moving the blade across the width of the slot over the entire width of the blade; a digital camera associated with the optical device; and a means of reconstructing a partial or complete image of the blade, from the succession of rows and columns filmed by the camera during the movements of the device and the blade holder.
• the at least one optical device comprises first and second integral parts, suitable for passing, respectively, under and above the rectangular slot during movement of the device, the first part comprising at least one lighting device and the second part comprising at least one objective placed on the optical axis defined by the lighting device and the slot.
• the device for lighting the first part comprises at least one light source.
• At least one light source is mounted on the optical device.
• at least one light source is external to the device and an optical fiber brings in the first part of the optical magnification device the beam emitted by the source.
• At least one light source is a continuous source.
• At least one light source is a pulsed source.
• the lighting device further comprises a converging lens, fixed in the first part of the optical magnification device, whereby it results that a light beam coming from the light source is condensed towards the slit.
• the lighting device further comprises a diaphragm interposed between the lens and the slot.
• the second part of the optical magnification device comprises a plurality of objectives and a selection means suitable for placing a single objective on the optical axis.
• the selection means comprises a controllable stepping motor capable of moving perpendicularly to the optical axis a drawer containing the plurality of objectives.
• the second part of the optical device also comprises an auto-focusing means for moving the objective on the optical axis.
• the auto-focusing means comprises a laser diode placed on the optical axis above the objective, an associated piezoelectric element, to the objective and controlled by the means of reconstruction.
• the means for moving the optical magnification device comprises at least one servo element actuated by a controllable stepping motor.
• the microscope comprises two optical magnification devices, a first high magnification device and a second low magnification device, each device being associated with its own means of movement.
• the blade holder is rectangular and has a U shape with a length at least equal to that of a blade.
• a servo element connects an abutment part of a blade in the blade holder to a stepping motor controllable by the reconstruction means.
• the blade holder further comprises a fixed part supporting a target.
• the means for moving the optical device and the blade holder for moving the blade provide their position in real time by means of reconstruction.
• the digital camera is a CMOS camera.
• the digital camera associated with the optical magnification device is placed in the second part of the latter.
• the second part further comprises means for transmitting to the digital camera the light beam leaving the lens.
• the transmission means is an i-transparent blade.
• a diaphragm is interposed between the semi-transparent plate and the digital camera.
• FIG. 1 schematically illustrates a microscope according to the present invention
• Figure 2 illustrates, in top view, a support element of a microscopic preparation according to the present invention
• FIG. 3 schematically illustrates a read head of a microscope according to the present invention
• Figure 4 schematically and partially illustrates an embodiment of the present invention.
• a microscope according to the invention essentially comprises an element suitable for receiving a slide (not shown) or “slide holder”, at least one optical magnification device or “read head”, and a reconstruction means. The structure and cooperation of these three elements will emerge from the following description of FIGS. 1 to 4.
• Figure 1 is a schematic and partial perspective view of the internal structure of a microscope according to the invention.
• a rectangular blade holder 1 rests on a substantially horizontal plate or plate 2.
• the plate 2 has a rectangular slot 3.
• the length of the slot 3 is at least equal to that of the blade holder 1.
• the width of the slot 3 is on the other hand less than that of a blade.
• the axis defined by the width of the slot 3 will be designated by X below and the axis defined by the length of this slot will be designated by Y.
• Z will denote the axis perpendicular to the XY plane.
• the blade holder 1 is placed parallel to the slot 3, above the latter, so that their lengths overlap. Plate 2 is fixed, as shown in the figure
• the blade holder 1 by making it integral with a fixed base 4.
• the blade holder 1 is movable relative to the plate 2 along the axis X.
• the mobility of the blade holder 1 relative to the plate 2 is symbolized in FIG. 1 by castors 5.
• the movement of the blade holder 1 above the slot 3 takes place over at least the entire width of a blade. In this way, all the points of a blade can be placed above the slot 3.
• the successive positions of the blade holder 1 are provided by means of reconstruction (not shown). Preferably, the latter controls the movement of the blade holder 1.
• the microscope according to the present invention also comprises an optical magnification device or read head 6.
• the complete structure of the head 6 will be detailed later in relation. with FIG. 3.
• the head 6 has at least one device illumination light 7 in a lower part and at least one objective 8 in an upper part.
• the lighting device 7 and the objective 8 are placed on the same vertical.
• the output of the objective 8 is associated with a digital camera 9 placed in the upper part of the head 6.
• the camera 9 is associated with the reconstruction means (not shown) so as to provide it with any filmed image and. receive any appropriate order.
• the upper and lower parts are joined in the following manner.
• the read head 6 is movable relative to the base 4. This mobility of the head 6 is symbolized in FIG. 1 by rollers 10.
• the read head 6 is movable in the horizontal plane XY only along the axis Y, c that is to say in the direction of the length of the slot 3.
• the head 6 has a C shape and is arranged so that, during its movement, the lighting device 7 passes under the plate 2 and goal 8 above. More particularly, the structure of the head 6 and its position relative to the plate 2 are such that the vertical passing through the lighting device 7 and the lens 8 crosses the slot 3.
• the vertical coming from the lighting device - Rement 7 and crossing the slot 3 will be called hereinafter "optical axis".
• the optical axis follows the length of the slot 3.
• the head 6 can be moved over at least the length of a blade.
• the head 6 is movable over the entire length of the slot 3.
• the successive positions of the head 6 are provided by means of reconstruction. Preferably, the latter itself controls the movement of the head 6.
• the blade holder 1 and the read head 6 are associated with a reconstruction means (not shown) whose role is as follows. After introduction of a blade into the blade holder 1, the latter and the head 6 are moved in the ways described above. Thus, the camera 9 scans the entire surface of the blade. From the succession of rows and columns provided by the camera 9 and from the position information provided by the blade holder 1 and the head 6, the means of reconstruction reconstructs an image of the blade. This image is either a complete image if the head 6 comprises a low magnification objective 8, or a complete or partial image in the case of a reading head 6 comprising a high magnification objective 8.
• the reconstruction means also stores all the data relating to the conditions of the acquisition of the images such as, for example, the type of the lighting device 7, the type of objective 8, or information relating to the stability of the system, some of which will be detailed below in connection with FIGS. 3 and 4.
• FIG. 2 illustrates, in schematic and partial top view, the part of the plate 2 of Figure 1 comprising the slot 3 and the blade holder 1.
• the blade holder 1 has an open shape in horizontal U (or stirrup) ) to allow the insertion and removal of a blade.
• the dimensions of the blade holder 1 are at least equal to those of a blade. Stabilization of the blade in the blade holder 1 is ensured in a reproducible manner as follows.
• a receiving and ejecting device similar to that of a computer diskette allows the insertion or removal of a blade. This device includes an ejection claw
• the operations of inserting or removing a blade are automated by adding to the device a blade loader.
• the blade holder 1 comprises an element integral with the end portion 28 adapted to cooperate with a displacement means.
• the end portion 28 will be notched so as to cooperate with a worm screw system controlled by a stepping motor (not shown).
• the blade holder 1 also includes, beyond the terminal part 28 for abutting a blade, a fixed part 29.
• the slot 3 is then designed so as to also pass under this fixed part 29.
• the movement of the blade holder 1 is then ensured either by an extreme end part, beyond the fixed part 29, or, preferably, by the middle part between the part carrying a blade and the fixed part 29.
• the fixed part 29 includes a test element 30 or "test pattern" consisting of a plurality of transparent, semi-transparent, or opaque regions as well as various engraved or colored elements. The role of such a test pattern 30 will be detailed below in relation to FIG. 4.
• FIG. 3 illustrates, in partial and schematic sectional view, an embodiment of a read head 6 of a microscope according to the present invention such as that of FIG. 1.
• the lower part of the head 6, suitable for passing under the slot 3 (FIG. 1) comprises an illumination device 7 comprising a converging lens 70 and a field diaphragm 71.
• the diaphragm 71 is interposed between the exit of the lens. tille 70 and the slot 3.
• the lens 70 condenses a light beam from a light source (not shown), the characteristics of which will be detailed later.
• the head 6 comprises at least one objective, for example three objectives 81, 82 and 83.
• the different objectives 81, 82 and .83 are distributed so to be able to selectively place one, for example 81, on the optical axis 7-3.
• the active objective 81 can be replaced at any time by any of the other objectives 82 and 83. The selection of the active objective will be explained below.
• such a transmission is not direct but takes place via a semi-transparent plate 11 which reflects the beam leaving the objective 81 towards the camera 9.
• a diaphragm The field 12 is interposed between the semi-transparent plate 11 and the camera 9.
• the camera 9 is connected, as has been explained above, to the reconstruction means 50.
• the characteristics of the light source depend on the desired enlargement, that is to say on the active objective and on the desired acquisition speed. Indeed, it is known that the higher the magnification sought, the higher the light intensity must be.
• the lighting device can be mounted in the lower part, directly under the lens 70. Beyond a certain magnification , generally 10x, the light intensity used requires light sources with a duration of limited life and causing significant heating. It will then be preferable to use such a source external to the head 6, the light beam necessary for observation being brought to the level of the lens 70 by a device of optical fibers. This facilitates handling, limits the size of the head and preserves the various elements of the overall device 1 heat.
• the light source is preferably a continuous white source, for example, a halogen lamp of 100.
• the source will preferably be a pulsed laser diode.
• the selection of the source used is carried out either automatically by the reconstruction means 50, or by a command from an observer.
• the reconstruction means 50 records the lighting conditions during the acquisition of the image.
• the selection of the objective 81 placed on the optical axis 7-3 is carried out as follows.
• the objectives 81, 82 and 83 are placed in a housing or drawer 84 one next to the other, linearly, for example along the axis X of the width of the slot 3.
• the drawer 84 is movable according to the distribution axis of the objectives 81, 82 and 83, for example the X axis.
• the drawer 84 is connected to an actuator 85.
• the actuator 85 is connected to the reconstruction means 50 to receive an instruction for selecting a objective. Such an instruction is issued by the reconstruction means 50 either automatically or at the request of an observer.
• the active objective 81 is movable on the optical axis.
• the selection of the position of the active objective 81 on the optical axis 7-3 is carried out as follows.
• a displacement element, preferably a piezoelectric element 86, is associated with the objective 81.
• the piezoelectric element 86 is controlled by the reconstruction means 50.
• the reconstruction means 50 provides such a command either on instruction of the observer, either automatically. In the latter case, the position of the active objective 81 will be determined by means of automatic focusing.
• the automatic focusing means comprises a laser diode 13 placed on the optical axis 7-3, on the other side of the semi-transparent blade 11 relative to the output of the active objective 81.
• a diaphragm 14 is interposed between the laser diode 13 and the semi-transparent plate 11.
• the laser diode 13 emits a light beam on the optical axis 7-3 .
• This beam passes through the semi-transparent blade 11, then the active objective 81 and reaches a blade (not shown) placed on the slot 3.
• the beam then successively crosses the upper surface of the strip, the lower surface of the strip , the object to be observed, the upper surface of the glass slide and the lower surface of the glass slide.
• the reconstruction means 50 controls the piezoelectric element 86 so as to vary the position of the objective 81 between the two positions corresponding to the reflections on the surfaces. lower of the coverslip and upper of the slide, that is to say so as to optimize the contrast in the area corresponding to the object to be observed.
• the digital camera 9 is a camera with CMOS technology.
• a low magnification objective can be provided in the drawer 85, it is advantageous according to the present invention to dissociate a head of reading allowing a high magnification (high resolution) of a reading head allowing a low magnification (low resolution).
• high magnification high resolution
• low magnification low resolution
• a low magnification head according to the present invention is therefore of a simplified design. Thus, it will not need a displacement device (piezoelectric element 86) on the optical axis of the active objective, nor consequently of automatic focusing means.
• a low magnification device is, as standard, a single objective of magnification between lx and 4x.
• the drawer 84 and the actuator 85 are then no longer necessary.
• a low resolution head according to the present invention will therefore preferably comprise a single fixed objective. Dissociating a low magnification head from a high magnification head as described above in relation to FIG. 2 has various advantages which will be detailed later in relation to FIG. 4. It will be noted now that the use of two heads allows reduce the size and complexity of producing a single multi-purpose multi-purpose head.
• FIG. 4 illustrates, in partial and schematic top view, an embodiment of a microscope according to the invention with two read heads 41 and 42.
• the first head 41 is a high magnification head comprising several objectives, for example three, for example 10 ⁇ , 20 ⁇ and 40 ⁇ , of a structure such as, for example, that described previously in relation to FIG. 3.
• the second head 42 is a low magnification head comprising a single fixed objective, for example a 4x objective, of a simplified structure with respect to the first head 41 as described above in relation to FIG. 3.
• the head 41 is movable at least one timing guide 431 and integral with a servo element 441 controlled by a stepping motor 45 controlled by the reconstruction means 50.
• a motor 46 controlled by the reconstruction means 50 makes it possible to move the head 42 on at least one timing guide 432, by means of a servo element 442.
• the head 42 has been associated with a single timing and servo guide in FIG. 4
• wedging guides 43, 47 as necessary to ensure the stability of the heads 41, 42.
• the head of FIG. 3 is associated with two crossing wedging guides, one at the bottom under the device illumination 7 and one in the upper part, above and to the right of the drawer 84 of FIG. 3.
• the control system will for example be a worm screw associated with the motor 45, 46 passing through, for example, the lower part under the camera 9
• the speed of movement of each head 41, 42 is controllable by the reconstruction means 50.
• one of the timing and / or servo guides 43, 47 of each read head is a ruler provided with optical marks. Such marks allow the reconstruction means 50 to place the read head in question with an accuracy of 0.1 ⁇ m.
• FIG. 4 also illustrates the plate 2 and the slot 3, a blade 46 being in position in the blade holder 1.
• the blade 46 has a label part 47 and a "useful" part 48.
• FIG. 4 also shows a stepping motor 60 associated with an endless screw 61, cooperating with the middle part
• a blade is introduced, either manually or using a blade loader, as described previously with reference to FIG. 2.
• the low magnification (low resolution) device 42 then performs a first scan of this slide. This scanning begins with the label part 47 in which are registered identification references of the preparation being viewed. Then, by a combination described above of displacements of the blade holder 1 and of the head 42, a complete and relatively little enlarged image of the preparation is reconstructed by the means 50. This image will be designated hereinafter "navigation image”. This navigation image is either archived, presented (displayed) to an observer, or both.
• the microscope according to the invention returns the low resolution read head 42 (low magnification) to its garage position, which is located outside the useful part 48 , preferably above the label part 47.
• the system then goes into high magnification mode and validates the head 41.
• the movement of the head 41 avoids the label part 47.
• the head 41 can move above the fixed part 29 comprising the test pattern 30.
• the garage position of the head 41 is vertical to the test pattern 30.
• the acquisition of successive images of high magnification is carried out automatically for all the available objectives and for the entire blade 46. This makes it possible to constitute a complete database.
• Each of the high magnification acquisitions is then carried out much faster than current devices, with much greater precision.
• the optical quality of the images and the precision of the measurements that can be carried out are evaluated in a reliable and objective manner which will be described below.
• the acquisition of successive images of high magnification is carried out online, selectively by an observer.
• the low magnification device 42 begins by acquiring the navigation image. It is therefore possible for an observer to select a pathologically or diagnostically interesting part and to limit the scanning of the high resolution head 41 to a restricted area of the slide 46. It will also be possible for him to very quickly select the desired objective.
• the automatically and immediately scanned navigation image can be reproduced on a display either in the immediate vicinity of the microscope according to the invention, or remotely via a computer network.
• the control of the reconstruction means is completely digital and can therefore also be carried out remotely via a computer network.
• the observer is therefore advantageously no longer forced to be in the immediate vicinity of the microscope.
• This first phase already has two major advantages over existing homologous solutions (scanner).
• the resolution is much better.
• the microscope according to the invention can be placed at a location remote from the observer.
• the adjustments of lighting and positioning of the lens on the optical axis are made automatically with the idea of the autofocusing device and the target placed on the blade holder, as this has been described previously in relation to FIG. 2.
• the different settings are stored by the reconstruction means and associated with the image concerned.
• the microscope according to the invention can perform a certain number of objective tests as to the optical quality of the microscope. These tests can be performed for different types of objectives.
• the reconstruction means 50 performs a measurement by acquiring, in the manner described above (combined displacements of the blade holder and of the head 42) an image of one or more selected parts of the test pattern 30. Next, a comparison with benchmark data to assess the stability of the system.
• the means of reconstruction can, depending on conditions previously fixed possibly modifiable by an observer, either indicate to the observer a perfect state of functioning of the device, or indicate to him a malfunction, by specifying to him which parameter (s) is (are) deviant (s). If the tests reveal a malfunction, provision may be made for automatic correction by the computer system, either by purely digital processing, or by a modification of the acquisition conditions, for example an increase or a decrease in the intensity of the light source of illumination, or a displacement of the objective. However, provision may also be made to limit the possibilities of automatic intervention of the system or to submit them for approval by an observer.
• An advantage in dissociating high magnification 41 and low magnification heads 42, in addition to the reduction in complexity is as follows.
• the head 42 is relatively stable and pre-adjusted during its construction.
• the quality of the images provided by the head 41 must be optimal. Its function must therefore be checked using tests carried out as described above. Dissociating the heads makes it possible to acquire the navigation image during at least part of the tests on the head 41, which shortens the acquisition time.
• the images obtained using a microscope according to the invention are advantageously perfectly reproducible. Their optical quality is optimized, and the exact conditions of their acquisition are memorized absolutely by the reconstruction means. This makes it possible to overcome the problems described above of individual variations linked to the use by a given observer.
• the microscope according to the invention can evaluate the optical quality of the images or even improve it, by keeping a record of this improvement.
• Another advantage of the present invention is interactivity with an observer. Indeed, unlike a a scanner-type device, the observer can intervene at any time via the reconstruction means to modify the acquisition conditions. In addition, the system can absolutely store these changes. Another advantage of the present invention is that it makes it possible to combine the observation system and an archiving system.
• a microscope according to the present invention is that it is both an observation system allowing a remote observer or not to locate the general appearance of a preparation and also to constitute a measuring instrument.
• This measuring instrument is advantageously an automatically standardized instrument. Indeed, by means of the test pattern, and tests as described above, it is possible to detect and automatically assess, or even correct, any possible drift in the system. This limits the possibilities of differences in results. .
• Another advantage of a microscope according to the present invention is its ergonomics. Indeed, the whole of the device described above will fit into a reader, the size of which on a workspace is approximately 30 ⁇ 20 cm. It has the advantage of being much less bulky in terms of height than a common microscope.
• the blades typically have a width of 25 to 30 mm, generally standardized to 26 mm, for a length of the order of 75 to 80 mm, generally standardized to 76 mm, with a label area of the order of 15 to 20 mm, usually 16 mm.
• the slot will have a width of between 3 and 5 mm.
• the blade holder will be designed with a width and length varying from 2 mm to the dimensions of the blade.
• the combination of lugs, claw, pusher and springs will allow repositioning of the same microscopic preparation to within half a micrometer.
• the associated stepper motor allows the blade holder to be moved with an accuracy of 0.1 ⁇ m, at a speed of 50 mm / s.
• the digital camera is of the CMOS type, with a size of at least 1280 ⁇ 1024 image points (pixels) such as, for example, the PB ⁇ MV13 Megapixel CMOS device marketed by the company Photobit of Pasadema, California, United States of North America, having a frame rate limit of around 2 ms per frame (500 frames per second), the acquisition performance with the various objectives is as follows.
• the motors of the high and low magnification heads are similar. Each moves the associated head at a speed between 1 and 10 cm / s. The choice of speed will be made according to the acquisition conditions (active objective, type of light source, etc.).
• the reconstruction means selects the speed either automatically or on the instruction of an observer.
• a 4x objective it takes a total time of between 4 and 30 s, for example around 13 s, to acquire a navigation image with an optical resolution of the order of 3 ⁇ m.
• the optical resolution is of the order of 1.2 ⁇ m, and the entire useful part of the microscopic preparation is acquired in a time between 60 and 200 s, for example of the order of 60 s.
• the acquisition of a succession of images, with an optical resolution of 0.5 ⁇ m, is carried out, for the entire useful part of the microscopic preparation, in a time between 400 and 1000 s, for example of the order of 700 s.
• the passage time of a high resolution image obtained with lOx, 20x or 40x objectives from a position defined in the navigation image is between 0.1 and 1 s, for example of the order of 0.25 s.
• the passage time from a high resolution image obtained with lOx, 20x or 40x objectives to another adjacent image obtained with the same objective is between 0, 1 and 0.5 s, for example of the order of 0 , 25 s.

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