JP3948962B2 - Imaging apparatus having a light source for irradiating a subject - Google Patents

Description

[0001]
BACKGROUND OF THE INVENTION
The present invention relates to an imaging apparatus that images a specimen containing bacteria on a stage, and more particularly to an imaging apparatus having a light source for irradiating a subject.
[0002]
[Prior art]
A sample observation camera for observing the state of a bacterium by placing a sample containing the bacterium on a stage and projecting a specific wavelength on the sample, in order to distinguish the state of the bacterium There is known an imaging technique in which a special reagent is given to a specimen containing, the excitation light corresponding to the state of the bacteria is generated, and this is imaged to count the number of solids of the bacteria.
[0003]
In such an imaging apparatus, since the luminance from bacteria that generate excitation light is usually weak, it is necessary to irradiate the stage uniformly and brightly in order to receive excitation light having an illuminance of a predetermined amount or more.
[0004]
Since a light source for irradiating light onto a stage in such an imaging apparatus requires light in a specific wavelength region, a light emitting diode can be used. Recently, since a light emitting diode having high emission luminance in a wide wavelength range has been developed, when such a light emitting diode is used as a light source, it is necessary to combine it with a color filter that cuts light in an unnecessary wavelength range.
[0005]
By the way, in such an imaging apparatus, since it is necessary to direct the excitation light of the bacterium in the direction of observing, when observing from above the stage as in microscopic photography, there is no irradiation method that illuminates from below the stage and brightens the background. . For this reason, illumination is performed obliquely with respect to the upper side to be observed. However, irradiation from a plurality of positions is necessary due to the necessity of irradiating light with a uniform illuminance and a predetermined illuminance on the stage.
[0006]
In the prior art, in order to irradiate the stage with light with high illuminance, the irradiation illuminance is improved by condensing the irradiation light from the light source with a lens and irradiating the stage with the light.
[0007]
[Problems to be solved by the invention]
In the irradiation method of condensing with the lens as described above, the illuminance is improved if the subject is at the focal position, but the shape of the light source itself is projected because of the focal position, and an image is formed on the subject. In order to avoid this, a light source called Koehler illumination that has a so-called defocused state that does not produce a focus at the expense of illuminance and has a compound lens that is configured by combining a plurality of lenses to maximize illuminance. There is.
[0008]
In such a light source, the purpose is to improve the illuminance by using two light sources for one wavelength and to make the illuminance of the area to be inspected as uniform as possible. The illuminance on the stage surface of light needs to be the same.
[0009]
Then, the center of the spot that can be regarded as uniform illuminance needs to be positioned substantially at the center of the stage on which the specimen is mounted. Furthermore, when two wavelengths must be switched and used in order to specify the state of the bacterium, two pairs of four are required, so that the position adjustment of the light source is not easy. In other words, since the overlapping position when assuming that two pairs of light sources are simultaneously irradiated should coincide with the approximate center of the area to be inspected as described above, the size of the narrowed light source spot is the inspection area. However, it cannot be said that the size has a margin.
[0010]
The present invention irradiates the upper surface of a stage on which a specimen is mounted uniformly with a predetermined amount of light, and easily adjusts the position of the light source in relation to the spot center of the light source and the optical axis of the imaging camera. It aims at providing the composition which can be.
[0011]
[Means for Solving the Problems]
For this reason, the present invention provides a stage for loading a subject, a plurality of light source means for converging light from a light emitting element that emits light to illuminate the stage with light rays, and light from the subject irradiated with the light rays. Lens means for converting the condensed two-dimensional image light into an electrical signal, and a holder for positioning the light source means integrally with the lens means, the holder comprising the lens means Fixing means to be fixed to the fixing means, a plurality of arm portions extending radially from the fixing means, and an attachment portion provided at an end of each arm portion for attaching the light source means, When the fixing means is attached and fixed to the lens means, the light from each light source means is directed obliquely from a plurality of directions symmetrical to the lens means to the stage. And a step for substantially positioning the light source means so that the converging position of all the light rays from each light source means is located on the optical axis of the lens means on the stage, and the attachment portion of the holder A light source unit, to which the light source means is attached, is attached to the lens means via the fixing means, so that the light source means is integrated with the lens means and positioned. Provided is an imaging device having a light source for the purpose.
[0012]
Here, the lens means is arranged so that its optical axis coincides with an orthogonal line passing through the upper surface of the central portion of the stage. As a result, the respective light source means and lens means are fixedly arranged with respect to the center of the stage. Moreover, in one embodiment, a light source means is comprised from a pair of light source arrange | positioned in the position where the optical axis opposes the said orthogonal axis as a center. For this reason, the light source means irradiates light from a plurality of positions obliquely above the stage, so that the entire surface of the stage can be irradiated with light with an illuminance equal to or higher than a predetermined level.
[0013]
In one embodiment, the light beam means constituting the light source means includes a light emitting element provided at one end of a rod-shaped hollow case, and the inside of the hollow case and / or the other end of the hollow case. And a filter means for controlling a wavelength region that passes through the lens.
[0014]
In the above configuration, since the holder has a step as a positioning portion for positioning the light source means at a predetermined mounting position of the holder, the relationship between the light source spot center of the imaging device and the optical axis of the lens means This makes it possible to easily adjust the position.
[0016]
DETAILED DESCRIPTION OF THE INVENTION
Prior to describing the details of the imaging apparatus, the technical background of the specimen to be imaged by the imaging apparatus will be described.
(1) Technical Background of the Imaging Device In the imaging device, a resin receptor is prepared in a membrane portion including a specimen to be counted. Since the illuminance of the light emitted from the bacteria to be photographed is extremely low, it cannot be grasped unless the exposure is increased by performing continuous exposure for a certain period of time. However, since the resin color of the receptor is not a perfect black body, the level increases with the image of bacteria. As a result, the contrast between the background portion and the bacterial portion in the finally acquired entire image has a very low S / N.
[0017]
Therefore, if the subsequent image processing is performed without imposing a burden, a malfunction will be caused. In addition, white noise processing for superposing a low-cost configuration is superimposed even when processing of weak signals is involved.
Therefore, it is necessary to sharpen the bacterial image before image processing by appropriately performing background correction.
[0018]
(2) Explanation background of bacteria emitting excitation light of a specific wavelength The background (background) is a film having a resinous specimen-containing body or an adhesive layer for adhering bacteria. Both are colored as dark as possible in order to efficiently obtain excitation light from bacteria (with good contrast). However, when the light is not completely black but dark gray, reflected light is generated when light is irradiated.
[0019]
In this bacterial device, the inspection is not performed with the reflected light of the irradiated light, but a special chemical solution is contained in the bacteria, and ultraviolet light or green light is irradiated in this state. In response to the ultraviolet light or green light, excitation light is generated from bacteria containing the chemical solution. This excitation light is received for inspection.
[0020]
Specifically, as shown in FIGS. 5A and 5B, when ultraviolet light having a wavelength of 370 nm (NSHU590) is irradiated, excitation light (exposure 1) is emitted from bacteria at a wavelength of 461 nm. When green light having a wavelength of 525 nm (NSPG500S) is irradiated, excitation light (exposure 2) is generated at a wavelength of 617 nm.
[0021]
FIG. 5C shows the spectral characteristics of the filter. In addition to the weakness of the excitation light with respect to the irradiation light, the wavelengths of the irradiation light must be cut in the optical path for receiving light because the wavelengths are close to each other.
[0022]
For this reason, ultraviolet light is irradiated with a cut filter having a characteristic of abruptly decaying from 360 nm, so that the transmittance becomes 0% at a point of about 400 nm. Moreover, the light source NSPG 500S has almost no power at the 400 nm point. Then, a filter that transmits a wavelength band of 420 nm or more, called L42, is inserted on the light receiving side. If it does so, the excitation light (exposure 1) whose principal wavelength is 461 nm can be acquired appropriately. This excitation light is generated for both live and dead bacteria.
[0023]
Similarly, a filter is used for green light. For green light, a band pass filter BP535 that transmits only the wavelength band from 450 nm to 550 nm is attached to the green light source (536 nm). NSPG500S as a light source emits light in a wavelength band from 450 nm to 650 nm, but the base is cut by the bandpass filter described above. A filter called O58 that transmits a wavelength band of 580 nm or more is inserted on the light receiving side. Excitation light (exposure 2) having a dominant wavelength of 617 nm can be obtained. This excitation light is generated only in dead bacteria.
[0024]
As described above, the excitation light can be acquired, but the wavelength regions that pass through each of them naturally remain. Although the irradiation light source is filtered, the amount of light source is extremely large. In particular, the green light source has about 2000 lux on the specimen irradiation surface.
[0025]
For this reason, only the excitation light cannot be picked up, and since the excitation light is weak, an exposure time of about 2 seconds is required, and the background level rises to about 128 at the time of resolution of 255 at the maximum. Since the number of pumping light outputs is 10 in this, it is preferable to remove the background level for subsequent image recognition.
[0026]
Hereinafter, an embodiment of an imaging device according to the present invention based on the above basic technique will be described in detail with reference to the drawings.
[0027]
FIG. 1 is a side view of the entire imaging apparatus 1. The main configuration of this apparatus is a stage 2 on which a subject is placed, a light source unit 4 that emits light to the subject, and two-dimensional image light emitted from the subject emitted from the light source unit 4 is incident. Then, the camera unit 3 that converts an electrical signal into an image signal, and further, the stage 2 is moved to the operation position when the sample is placed on the previous stage 2, and the area to be examined is subdivided. It comprises a stage moving unit 9 that changes the shooting area position that enables fine inspection by shooting with the camera unit 3 of the magnifying optical system.
[0028]
FIG. 2 is a front view of the light source unit 4 as viewed from above and above the apparatus. FIG. 3 is an enlarged view of a portion where the camera 3 and the rod light source 6 are attached to the holder 51.
[0029]
The light source unit 4 includes four pairs of a rod light source 6a and a rod light source 6b as first light source means and a pair of a rod light source 6c and a rod light source 6d as second light source means. Each rod light source 6 includes substrates 50a, 50b, 50c, and 50d at end portions, and light emitting diodes are respectively disposed on the substrate toward the inside of the rod.
[0030]
As the first light source means, the rod light source 6a and the rod light source 6b (NSPG500S) emit light of the same wavelength (ultraviolet light having a wavelength of 370 nm) and work as a pair. As the second light source means, the rod light source c and the rod light source d (NSPG 500S) emit light of the same wavelength (green light having a wavelength of 525 nm) and work as a pair.
[0031]
As described above, the reason why the two rod light source pairs (rod light sources 6a and 6b) are placed at the target position is to complement each other to make the irradiated surface substantially uniform in illuminance. The pair of the rod light source 6a and the rod light source 6b and the pair of the rod light source 6c and the rod light source 6d do not light simultaneously and do not need to be orthogonal.
[0032]
With this configuration, the pair of rod light sources 6 irradiate the irradiation stage with the imaging means disposed above it so that it has the same illuminance at the same wavelength obliquely from two positions symmetrical to the stage on the side of the imaging means. The projection area matches the irradiation stage so that there is no uneven illumination.
[0033]
The stage 2 is positioned below the light emitting end opposite to the substrate mounting to which the light from the light source unit 4 is transmitted. Above the light emitting end, a camera 3 of lens means is provided, and a filter unit 55 that advances and retreats in parallel with the surface of the stage 2 is located between the camera 3 and the light emitting end. The filter unit 55 is provided with an L42 filter 55a that cuts a wavelength band below 420 nm and an O58 filter 55b that cuts a wavelength band below 580 nm.
[0034]
With this filter unit 55, it is possible to obtain clear excitation light with the interfering wavelength band cut off in the camera 3.
[0035]
Specimens for inspecting bacteria include cases where the target is agitated in a solution and then water-absorbed and hydrated, and then fixed to a specimen device called a membrane kit capable of containing a reagent for examination, and an adhesive surface containing the reagent In some cases, the film is collected directly with a film having Further, since the fixing device having a different shape may be considered, the shape of the fixing device is not particular. As already described, since the magnified optical system forms a magnified image with respect to the image sensor, the distance to the subject is extremely strict because the depth of focus is small.
[0036]
For this reason, the stage 2 has three adjustment points, and adjusts not only the height adjustment but also the level. As shown in the adjustment margin that can be adjusted at these adjustment points, an attachment portion (not shown) for the frame of the camera 3 has a long hole so that rough adjustment is possible. These adjustments determine the position of the stage and the position of the camera.
[0037]
The rod light source 6a, the rod light source 6b, the rod light source 6c, and the rod light source 6d, which are four rod-shaped light sources, are configured to be attached to one mounting holder 51. The attachment holder 51 is X-shaped when viewed from above, and includes a ring portion 57 as a fixing means for attaching the camera 3, and an arm portion 56 a, an arm portion 56 b, an arm portion 56 c, and an arm that protrude radially from the ring portion 57. The unit 56d is configured.
[0038]
The arm portions 56a, 56b, 56c, and 56d can be fixed by screwing the fixtures 62a, 2b, 62c, and 62d provided on the rod light sources 6a, 6b, 6c, and 6d, respectively, in an adjustable manner. Has a rod mounting portion.
[0039]
The four attachment portions of the holder 51 are formed so that when the rod light source 6 is attached, the irradiation light of the rod light source 6 has an angle positioned on the optical axis of the lens means. In this example, it is composed of four, but may be six, for example. That is, in this case, the ultraviolet light is provided at a position corresponding to the apex position of the equilateral triangle, and the green light is provided at a position corresponding to the apex position of the equilateral triangle.
[0040]
The pair of rods that irradiate the irradiation light having the same wavelength irradiates the camera 3 on the stage 2 at a symmetrical angle and in an oblique direction from the opposite position, so that the irradiated surface has a substantially uniform brightness. Each rod light source 6 is fixed to one holder 51 and formed on the outer wall of each rod so that the irradiation light of the two rod pairs is uniform over the entire photographing area on the stage 2 for photographing the subject. The steps 52a, 52b, 52c, and 52d thus engaged with the holder 51 can be positioned.
[0041]
Furthermore, not only the position but also the LED, which is a light-emitting element, has a large variation in product accuracy, and when used at the same operating voltage, there are illuminances that are about twice as high. The current is adjusted for each line so that the illuminance is uniform.
[0042]
The rod light source 6 and the holder 51 are attached by engaging the step 52 of the rod light source 6 with the attachment portion of the holder 51 and adjusting the position, and then screwed to the attachment portion of the holder 51 with the attachment tool 62. This step is machined so that the tip of the rod light source 6 is positioned at a desired position with almost no adjustment in design. If it is necessary to further adjust the position of the light source after screwing, the screws are loosened so that the adjustment can be performed easily.
[0043]
Thus, the light source means of the present invention is configured to be adjustable so that the light convergence position is located on the optical axis of the lens means. In the light source means, a unit assembly process of the rod light source 6 and the holder 51 is performed in the light source means assembly process before the assembly process to the lens means. At this stage, the adjustment is almost completed, and the adjustment of the positions of the lens means and the light source means is completed by assembling the light source unit to the lens means, thus simplifying the adjustment process.
[0044]
FIG. 4 is a cross-sectional view showing the structure of the rod light source 6 using the rod light source 6a as an example. The rod light source 6 differs between a 370 nm light source such as a light emitting diode color, a filter color, and a lens, and a 525 nm light source, but the basic structure is the same. When roughly divided, it is composed of a rod 60a and a rod 61a having a smaller diameter. A light emitting diode 64a is fixed to the substrate by soldering at one end of the rod 60a to the substrate 50a via a diode spacer 63a.
[0045]
In front of the light emitting diode 64a in the light emitting direction, there is a lens 65a fixed by a ring 72a. A rod 61a is inserted into the other end of the rod 60a. At the end inserted into the rod 60 of the rod 61a, there are a lens 66a and a band cut filter 73a fixed by a ring 74a and a spacer 67a.
[0046]
The band cut filter 73a has a function of cutting a hindrance in the respective light emission distributions. The U380 filter 73a is in the rod light sources 6a and 6b, and the L42 filter 55a is in the rod light sources 6c and 6d. Equipped to sharpen the light emission distribution.
[0047]
At the other end of the rod 61a, there are a lens 68a and a lens 69a fixed by a ring 70a and a spacer 71a. Further, an attachment portion 62 a for attaching to the holder 51 is provided. Then, a step 52 a created by the difference in diameter between the rod 60 a and the rod 61 a engages the holder 51. With this step 52a, the attachment of the rod 61 is accurately positioned with respect to the rod attachment position of the holder 51.
[0048]
As described above in detail, in the present invention, the light switching unit for switching the light beam irradiated from the light source unit for each light beam having a plurality of different wavelength regions, and the light beam irradiated by the light beams of the plurality of different wavelength regions. An imaging apparatus comprising: lens means for condensing light from a subject; and an image sensor that converts the two-dimensional image light collected by the lens means into an electrical signal. First light source means for irradiating light having a wavelength region of the first light source means for irradiating light from a plurality of obliquely upper locations, and second light emitting means for irradiating light having the second wavelength region obliquely upward. Second light source means for irradiating from a plurality of locations, holder means for supporting the first light source means and the second light source means, and the holder means in the lens means By providing a fixing means for fixing at a position, it is possible to irradiate the upper surface of the stage on which the specimen is mounted with a uniform amount of light more than a predetermined amount. The position of the light source can be adjusted in relation to the optical axis of the camera.
[0049]
Further, as already described, the rod light sources are not fixed to four, but not only the light source used by switching to two wavelengths but also the spot irradiation as a light beam without switching, and this spot position is Since the object is to achieve the purpose of aligning the center with the optical axis of the lens means, the present invention is not limited to an apparatus that images excitation light.
[Brief description of the drawings]
FIG. 1 shows a side view of an imaging apparatus having a light source for irradiating a subject according to the present invention.
FIG. 2 is a configuration diagram of light source means when the imaging apparatus of FIG. 1 is viewed from above.
FIG. 3 is a side view showing a state where a camera and a rod light source are attached to a holder.
FIG. 4 shows a cross-sectional view of the internal structure of a rod light source.
FIG. 5 is a diagram illustrating reagent characteristics, LED emission characteristics, and filter spectral characteristics.
[Explanation of symbols]
DESCRIPTION OF SYMBOLS 1 Imaging device 2 Stage 3 Camera (lens means)
4 Light source unit 6 Rod light source 20 Table 50 Substrate 51 Holder 55 Filter unit 55a L42 filter 56 Arm 62 Attaching tool 64 Light emitting diode (light emitting means)

Claims (6)

A stage to load the subject,
A plurality of light source means for converging light from the light emitting elements to emit light and illuminating the stage with light rays;
Lens means for condensing light from a subject irradiated with the light beam and converting the collected two-dimensional image light into an electrical signal;
A holder for positioning the light source means integrally with the lens means;
With
The holder includes a fixing means that is attached and fixed to the lens means, a plurality of arm portions that extend radially from the fixing means, and an attachment portion that is provided at an end of each arm portion and for attaching the light source means. And
Each of the attachment portions is configured such that when the fixing means is attached and fixed to the lens means, light beams from the light source means are directed obliquely from a plurality of directions symmetrical to the lens means to the stage, and A step for substantially positioning the light source means so that the convergence position of all the light rays from each light source means is located on the optical axis of the lens means on the stage;
A light source unit formed by attaching the light source means to the attachment portion of the holder is attached to the lens means via the fixing means, whereby the light source means is integrated and positioned with the lens means. An imaging apparatus having a light source for irradiating a subject. The light source means is
First irradiating means comprising light means for emitting light in the first wavelength region;
A second irradiation means comprising a light beam means for emitting a light beam in the second wavelength region;
The imaging apparatus having a light source for irradiating a subject according to claim 1.   The imaging apparatus having a light source for irradiating a subject according to claim 2, wherein a light beam irradiation direction of the first irradiation unit is different from a light beam irradiation direction of the second irradiation unit.   The imaging apparatus having a light source for irradiating a subject according to claim 2, further comprising a light emission switching unit that controls light emission by switching the plurality of light beam units. The light beam means
A light emitting device provided at one end of a rod-shaped hollow case;
A lens provided inside the hollow case and / or at the other end of the hollow case;
Filter means for controlling the wavelength region that passes through the lens;
The imaging apparatus having a light source for irradiating a subject according to claim 2. The fixing means includes
The imaging apparatus having a light source for irradiating a subject according to claim 1, wherein a predetermined position fixed to the lens unit is adjustable.

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