Abstract:
A confocal microscopic equipment for measuring the solid shape of a sample at high speeds, and which does not require scanning of an irradiating light beam, wherein are provided a light source, apertures on which the output beam from the light source is irradited, an objective lens that forms stationary images of the apertures on the sample and collects the reflected or fluorescent light from the sample, a photo detector, and a beam splitter that makes the beam that has passed through the apertures incident to the objective lens by means of transmission or reflection and focuses the output beam from the objective lens on the photo detector by reflection or transmission.

Description:
RELATED APPLICATION 
     This is a division of Ser. No. 09/433,903 filed Nov. 5, 1999, soon to now U.S. Pat. No. 6,111,690, which was a division of Ser. No. 09/003,068 filed Jan. 5, 1998, now U.S. Pat. No. 6,031,661 issued Feb. 9, 2000. 
    
    
     BACKGROUND OF THE INVENTION 
     1. Field of Invention 
     This invention relates to confocal microscopic equipment that can measure the solid shape of a sample at high speed; and more particularly, to equipment that does not require scanning of an irradiating beam. 
     2. Description of the Prior Art 
     The confocal microscopic equipment has resolution in the direction of an optical axis as well as resolution on a sample surface by scanning the irradiating beam on the sample and detecting light reflected from the sample through a pinhole or slit. The resolution in the direction of the optical axis obtained by detecting light through a slit is described in “Three-dimensional optical transfer function analysis for a laser scan fluorescence microscope with an extended detector” by S. Kawata, R. Arimoto, and O. Nakamura, J. Opt. Soc. Am. A. Vol 8, No. 1, (1991). 
     FIG. 1 shows a conventional confocal microscopic equipment, wherein an output light beam from laser  1  is made incident to galvo-scanner  2  and then a reflected beam from galvano-scanner  2  is made incident to beam splitter  4  via mirror  3 . Beam splitter  4  reflects the incident beam from galvano-scanner  2  and makes the reflected light irradiate sample  6  via objective lens  5 . The reflected light from sample  6  is again made incident to objective lens  5  and then made incident to one dimensional charge coupled device (CCD) camera  7  via beam splitter  4 . 
     The operation of the FIG. 1 equipment will be described with reference to FIG. 2, which refers to each pixel of CCD camera  7 . The output beam of laser  1  is scanned by galvano-scanner  2  and directed to scan sample  6  in a one dimensional direction. The reflected beam from sample  6 , scanned in a one dimensional direction, is made incident to CCD camera  7 . In CCD camera  7 , the reflected beam scanning direction is set to match the direction marked “A” in FIG.  2 . For example, the reflected beam is detected, for example, in turn with the pixels marked “B”, “C”, and “D” in FIG.  2 . In this case, since each pixel can detect the reflected beam only in the range of the height of pixel marked “E” in FIG. 2, this means that the pixels detect substantially only the reflected beam that has passed through a slit of width “E”. 
     That is to say, if it is assumed that the direction of the optical axis is the z-axis, the plane orthogonal to the optical axis is the x-y plane and the direction of scanning is the x-axis, then the equipment has resolution which is also in the z-axis direction. Thus, the fault shape of the x-z plane can be measured by scanning sample  6  in the z-axis direction. In addition, measurement of the solid shape of the sample  6  is made possible by scanning sample  6  in the z-axis direction while moving sample  6  in the y-axis direction in turn. 
     As a result, the resolution in the direction of the optical axis as well as the resolution on the sample surface can be obtained by scanning a laser beam in a one dimensional direction and by detecting the reflected light with a one dimensional line CCD camera  7 . 
     However, there are problems one encounters when using conventional equipment in that expensive optical equipment such as a laser  1  is necessary and a scanning means such as a galvano-scanner  2  is required. 
     Moreover, there are other problems. For example, the wavelength becomes longer and the resolution worsens because the laser incorporated in the confocal microscopic equipment can be used only in a red color system due to size and price. Also, since the laser beam is a coherent light beam, speckle noise is generated , which degrades the signal to noise ratio (S/N). 
     SUMMARY OF THE INVENTION 
     Accordingly, an object of the invention is to overcome the aforementioned and other disadvantages, problems and deficiencies of the prior art. 
     Another object is to provide a confocal microscopic equipment which can measure the solid shape of a sample with high speed. 
     A further object is to provide a confocal microscopic equipment that does not require scanning of an irradiating beam. 
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS 
     FIG. 1 a diagram depicting a conventional confocal microscopic equipment. 
     FIG. 2 is diagram depicting the details of each pixel in the CCD camera shown in FIG.  1 . 
     FIG.  3 (A) a diagram depicting an illustrative embodiment of the invention; and FIG.  3 (B) a graph of intensity and focal point. 
     FIGS.  4 (A) and  4 (B) are a perspective view of a sample having a three dimensional shape and an image of a cross sectional view of a sample, respectively. 
     FIGS.  5 (A)- 5 (E) a views depicting aperture shapes. 
     FIG. 6 a diagram depicting another illustrative embodiment of the invent using a cylindrical lens array. 
     FIG. 7 is diagram depicting a further illustrative embodiment of the invention using a polarized beam splitter. 
     FIG. 8 is diagram depicting another illustrative embodiment of the invention using an illuminating system of microscopes. 
     FIG. 9 is diagram depicting a still further illustrative embodiment of the invention. 
     FIG. 10 is a view depicting slits to which color filters are attached. 
    
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS 
     FIG. 3 shows a confocal microscopic equipment wherein an output beam from white light source  8 , which is an incoherent light source, is made incident to slit-array  10  in which a plurality of slits, or apertures, are provided (called “slit array”) through lens  9 . The beams that pass through each slit of slit array  10  are made incident to beam splitter  11 , such as a half mirror dichroic mirror. Beams that are transmitted through beam splitter  11  irradiate sample  13  through objective lens  12 . The reflected beam from sample  13  is again made incident to beam splitter  11  through objective lens  12  and the beam reflected by beam splitter  11  is made incident to two dimensional CCD camera  14  which is a photo detector. Sample  13  is scanned in the direction of the optical axis (see double headed arror “A ” in FIG.  3 (A)) by means not shown in FIG.  3 (A). 
     Operation of the embodiment of FIG.  3 (A) is described with reference to FIG.  3 (B). Slit array  10  is placed on the image forming plane of objective lens  12  and the slit width is the same as that of the Airy first dark band on the image forming plane. The distance between slits is about 10 times the foregoing slit width. If light is collected with a lens, actually it is not neatly focused into a point, but forms a fine image in which light and dark rings are arranged alternately surrounding the center light part. These rings are called “Airy disks” and the dark ring adjacent to the center part is the Airy first dark band, referred to above. 
     Let the width of the Airy first dark band be “b”, wavelength be “λ” and the numerical aperture be “NA”, then 
     
       
           b= 1.22×λ/NA  (1) 
       
     
     For example, if it is assumed that the magnification of objective lens  12  is 100, the numerical apertures NA of objective lens  12 =0.9 and λ=0.5 μm, then since the numerical apertures in the place of “B” in FIG.  3 (A) becomes 1/100, 
     
       
           B= 1.22×0.5/(0.9/100)=68 μm  (2) 
       
     
     Therefore, a slit having such a width can be easily prepared by etching or other methods. 
     When the output beam of white light source  8  is made incident on slit array  10 , a plurality of slit images of slit array  10  are formed on sample  13  by means of objective lens  12 . The slit images on sample  13  are made incident on CCD camera  14 . CCD camera  14  is a two dimensional CCD camera and multiple pixels are arranged on a plane. By considering the pixel array corresponding to the slit width, the camera detects only the reflected light passing through a slit of fixed width, as described before. 
     For example, the slit image marked “C” in FIG.  3 (A) is measured by the pixel array in the place where the image is formed. If the width of the pixel array in the x-direction in FIG.  3 (A) is assumed to be “W”, the pixel array detects only the beam passing through the slit whose width is “W”. Thus, the equipment has resolution in the direction of the optical axis as well as resolution on the sample surface. In this case, if the sample surface is located at the focal plane of objective lens  12 , the slit images are clear. On the other hand, if sample  13  is moved upward or downward from the focal plane, such as by a means not shown in FIG.  3 (A), the slit image will be out of focus. 
     If the slit images are detected by pixels of width “W” on CCD camera  14 , a maximum intensity of light is made incident in the case where the focal position coincides with the focal plane as shown in FIG.  3 (B). The incident intensity of light decreases as the focal position is shifted from the focal plane. In other words, the focal position in which the incident intensity of light becomes the maximum shows the height of the sample in the direction of the optical axis. 
     Accordingly, a change of the surface of sample  13  in the direction of the optical axis can be obtained in the following way. That is, the focal position at which the intensity of light measured by the CCD camera  14  becomes maximum is determined by scanning sample  13  in the direction of the optical axis marked “A” in FIG.  3 (A) with a means not shown in FIG.  3 (A). For example, if a sample of a three dimensional shape having a step “L” as shown in FIG.  4 (A) is measured by the invention, slit images are formed as shown with the broken lines in FIG.  4 (A). If the plane of surface “A” in FIG.  4 (A) is the focal plane, the slit image of plane “B” in FIG.  4 (A) will be out of focus. Conversely, if the plane of “B” in FIG.  4 (A) is the focal plane, the slit image of plane “A” in FIG.  4 (A) will be out of focus. Thus, a tomographic image, as shown in FIG.  4 (B), can be produced by determining focal positions in which the maximum intensity of light is inputted to each pixel of CCD camera  14 , respectively. 
     As a result, the resolution in the direction of the optical axis is obtained without scanning an irradiating beam using incoherent light. That is, this can be achieved by forming slit images on the sample surface and by determining, respectively, focal positions in which the maximum intensity of light is inputted to each pixel of CCD camera  14 . 
     In addition, the equipment of the invention has resolution in the direction of the optical axis as well as resolution which is equivalent to that of an ordinary optical microscope in a direction orthogonal to the optical direction. Also, the objective lens  12  may focus fluorescence from sample  13 . It is not limited to reflected beams from sample  13 . 
     In the embodiment of FIG.  3 (A), the transmitted beam through beam splitter  11  is focused on sample  13  and the reflected beam from beam splitter  11  is made incident on CCD camera  14 . However, the reflected beam from beam splitter  11  may be focused on sample  13  and the transmitted beam through beam splitter  11  may be made incident to CCD camera  14 . 
     In addition, if the slit width is increased, the intensity of light is increased and spatial resolution is decreased. On the other hand, if the slit width is decreased, the spatial resolution is improved but the intensity of light is decreased. Hence, the slit width need not be limited to the width of the first Airy first dark band, but, it is sufficient for the width to be from approximately 1/20 up to 20 times the width of the Airy first dark band, as determined by experimentation. 
     Similarly, if the pitch of the slits is increased, the resolution in the direction of the optical axis is improved because interference by adjacent slits is reduced but the number of slits that can be simultaneously measured is decreased. On the other hand, if the above distances are made smaller, the number of slits is increased and interference between the slits is generated, which makes correct measurement difficult to achieve. Therefore, the pitch of the slits is not limited to 10 times the Airy first dark band width, but is is sufficient for the distance to be about two or more times the Airy first dark band width, based on experimentation. 
     Furthermore, the photo detector is not limited to a CCD camera, but a camera tube, film or the like, may also be used and direct observation by the human eye using an ocular device can be adapted. 
     As shown in FIG.  5 (A), the shape of the apertures is not limited to slits. For example, a reticulate shape can be arranged by using multiple slits orthogonal to each other. Such an aperature shape can measure the three dimensional shape of a sample more accurately. As shown in FIG.  5 (B) a spherical shape can be adapted using a plurality of rings. Another shape is shown in FIG.  5 (C) using radial slits. If the shape of the sample is known beforehand, an aperture shape, such as that shown in FIG.  5 (D), can be used wherein the place requiring measurement is limited, for example. If the reflectively of the sample surface varies depending on the location thereof, the width of the slit “W” corresponding to places of small reflectivity may be made wider, as shown in FIG.  5 (E). 
     If the apertures consist of pinholes of a diameter approximately the same as that of the Airy first dark band, coherent light can be used because speckle noise is not generated even if coherent light is used. The aperature may comprise an etched metal plate or a light shielding film formed on a glass substrate partially removed by etching. Also, the apertures may be made of a transmitting liquid crystal panel. In this case, such aperatures are especially effective because the shape of the aperatures can be electrically and conveniently changed. 
     If the shapes between the slits are to be measured, images between slits can be interpolated by moving slits in a direction orthogonal to the optical axis, obtaining multiple solid images, and composing them thereafter. In this case, a stage or the like on which a sample or slits are provided may be moved, or the measurements can be realized by moving the apertures, changing the shapes, etc by controlling the transmitting liquid crystal panel. 
     In the embodiment of FIG.  3 (A), the ratio of slit width to slit interval is 1:10. This indicates that the ratio of aperture areas is 10%, and that only 10% of the incident beam is utilized. For this reason, the embodiment of FIG. 6 may be convenient and useful. The FIG. 6 embodiment comprises a white light source  8  which provides an output beam which is made incident on cylindrical lens array  15 , which is a focusing means, through lens  9  and the beam is converged by cylindrical lens array  15  and made incident on each slit of slit array  10 . Each slit of slit array  10  is placed at each focal point of the cylindrical lenses, respectively. By using the embodiment of FIG. 6, the incident beam can be effectively utilized. In addition, as the focusing means, both the ordinary convex lenses and. Fresnel lenses and refractive index distributed lenses can be used. Furthermore, if a transmitting liquid crystal panel is used for the apertures, micro-lenses may be added to each liquid crystal element. 
     The beam splitter  11  is shown as being a half mirror, but, a polarized beam splitter can also be used. FIG. 7 shows an illustrative embodiment using a polarized beam splitter. For example, polarizer  16  converts the transmitted beam to an S polarized light. Polarized beam splitter  17  transmits S polarized light and reflects P polarized light. The light passing slit array  10  becomes S polarized light by being transmitted through polarizer  16  and is made incident to polarized beam splitter  17 . The S polarized light is made incident to quarter wave plate  18  after transmission through beam splitter  17 . The incident light thus becomes circularly polarized light, which irradiates sample  13  through objective lens  12 . The reflected light from sample  13  is again made incident to quarter wave plate  18  through objective lens  12 . As a result, the circularly polarized light returns to linearly polarized light and is made incident to polarized beam splitter  17  as a 90 degree rotated P polarized light. Thus, the P polarized light is reflected by polarized beam splitter  17  and is made incident on CCD camera  14 . 
     As a result, 100% of the incident light from slit array  10  can be made incident to the CCD camera, compared with the case where a half mirror is used. 
     The scanning of the sample  13  in the direction of the optical axis is done by moving the microscope using pulse motor or the like. However, sample  13  or slit array  10  may also be moved using piezo-electric elements. In addition, objective lens  12  may be moved in the direction of the optical axis. Although slit array  10  is mounted on the image forming plane of objective lens  12  in the embodiment of FIG.  3 (A), an illuminating system for the microscopes can be used. 
     FIG. 8 shows an embodiment wherein an illuminating system of microscopes is used, wherein an output beam of illuminating light source  19  is focused with lens  20  and is made incident to mirror  22  through the iris of light source  21 . This incident beam is reflected by mirror  22  and is then made incident on lens  24  through the iris of condenser  23 . Light transmitted through lens  24  illuminates sample position  25  and is finally made incident to lens  29  through lenses  26  and  27  and field stop  28 ,respectively, and is then observed. 
     The operation of the embodiment of FIG. 8 is as follows. Since an image located in the position of the iris of light source  21  is formed on sample position  25 , the apertures, such as slit arry  10 , are provided in the position of the iris of the light source. Since the image of the apertures, such as slit array  10 , is projected on sample position  25  in the foregoing manner, observation of the image using an ordinary ocular device or camera makes it possible to easily change an ordinary microscope into a confocal system. Although FIG. 8 illustrates a microscope having an illuminating system of transmitted light illumination, this concept is also applicable to an illuminating system of reflected lumination. 
     In FIGS.  3 (A) and  7 , the system has a configuration wherein the beam transmitted through slit array  10  is made incident on beam splitter  11  or polarized beam splitter  17 . However, it may have the configuration shown in FIG.  9 . The difference between the embodiments of FIGS.  9  and  3 (A) is that in FIG. 9 the beam splitter  11  is located between light source  8  and slit array  10 , whereas in FIG. 3 (A), the beam splitter  11  is located between slit array  10  and sample  13 . In addition, lens  30  becomes necessary in FIG. 9 to focus the branched light on CCD camera  14 . 
     As described above, when the shapes between the slits are to be measured, the images between the slits are interpolated by composing cross sectional images obtained by moving slits in a direction orthogonal to the optical axis. This is because the distance between the slits is made to be about twice the width of the Airy first dark band or more to suppress interference between adjacent slits. However, the following method can also be used for this purpose. 
     As shown in FIG. 10, the slits to which different color filters are attached are located close to each other and the same color filters as relevant slits are also arranged on the CCD camera located on the photo detecting side. For example, in FIG. 10, red filters are attached to the slits marked “A” and “D”, a green filter to the slit marked “B”, and a blue filter to the slit marked “C”. Interpolation between slits becomes possible by performing data processing for each filter color on the cross sectional images obtained in such embodiment. Thus, spatial resolution is improved three fold. In addition, filter colors are not limited to these three colors, but, a similar effect can also be obtained by using color liquid crystals as color filters and by using a color CCD camera. 
     The invention is not limited to usage as microscopes. The shapes of large sized objects, such as mechanical parts,can also be measured using an ordinary camera lense in the place of the objective lens. Advantageously, this invention provides a confocal microscopic equipment which does not require scanning of an irradiating beam. In the invention, slit images are formed on the sample surface, and the focal positions, in which the maximum intensity of light is inputted to each pixel of the CCD camera, are determined, respectively. Moreover, advantageously, in the invention which uses incoherent light as an irradiating beam the resolution is improved because of the spectral light components of shorter wavelengths, such as blue color, or ultra violet light can be used, without generating speckle noise. 
     The foregoing description is illustrative of the principles of the invention. Numerous extensions and modifications thereof would be apparent to the worker skilled in the art. All such extensions and modifications are to be considered to be within the spirit and scope of the invention.