CN109884012B - Fluorescent microsphere test piece, manufacturing method and manufacturing assembly thereof - Google Patents
Fluorescent microsphere test piece, manufacturing method and manufacturing assembly thereof Download PDFInfo
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Abstract
本发明提供荧光微球测试片的制作方法，包括制作原料容置区，将荧光微球溶液滴入原料容置区内自然流平风干形成平铺一层的荧光微球，在所述荧光微球上滴加树脂，盖上盖玻片以使原料容置区形成封装空间内填充满树脂无空隙，固化树脂后剥离盖玻片即可得到荧光微球测试片。该方法中通过使荧光微球溶液滴入原料容置区内自然流平风干形成平铺一层的荧光微球，从而保证了荧光微球在任一纵向排布上不层叠、不堆积，进而可获得较准确的检测结果；通过测量尺来控制最终制得的荧光微球测试片的厚度，从而可根据需求获得不同厚度的荧光微球测试片。
The invention provides a method for making a fluorescent microsphere test piece, which includes making a raw material accommodating area, dropping a fluorescent microsphere solution into the raw material accommodating area, and naturally leveling and air-drying to form a flat layer of fluorescent microspheres. Drop resin on the ball, cover with a cover glass so that the raw material accommodation area forms an encapsulation space filled with resin without voids, and peel off the cover glass after curing the resin to obtain a fluorescent microsphere test piece. In this method, the fluorescent microspheres solution is dripped into the raw material accommodation area and naturally leveled and air-dried to form a flat layer of fluorescent microspheres, thereby ensuring that the fluorescent microspheres are not stacked or stacked in any longitudinal arrangement, and thus can be Accurate detection results are obtained; the thickness of the finally prepared fluorescent microsphere test piece is controlled by a measuring ruler, so that fluorescent microsphere test pieces of different thicknesses can be obtained according to requirements.
Description
Technical Field
The invention relates to the technical field of performance testing of confocal endoscopes, in particular to a fluorescent microsphere testing sheet, a manufacturing method and a manufacturing assembly thereof.
Background
The fluorescent microsphere test piece has very wide application in the field of integrated detection of confocal endoscopes, including detection for longitudinal resolution, detection for fluorescent stained cells and the like, however, the structure of the existing fluorescent microsphere test piece comprises a glass slide, a cover glass and fluorescent microspheres between the glass slide and the cover glass, the fluorescent microspheres of the structure have certain defects, taking the detection of axial resolution as an example, on one hand, due to the fact that gray scale change of the fluorescent microspheres needs to be obtained along the longitudinal direction during detection, the fluorescent microspheres need not to be stacked and piled up on any longitudinal arrangement, but the fluorescent microspheres in the existing fluorescent microsphere test piece are scattered and distributed in the glass slide and the cover glass, and therefore, the fluorescent microsphere test piece meeting the requirements is troublesome to manufacture; on the other hand, the thickness of the cover glass is 170um, so that the thickness of the whole fluorescent microsphere test piece is larger than 170um, and for a confocal endoscope with the working distance smaller than 170um, if the fluorescent microsphere test piece with the structure is used, the fluorescent microspheres covered by the cover glass cannot be imaged, and the axial resolution cannot be detected.
Therefore, it is a problem to be solved to provide a fluorescent microsphere test strip which is simple to prepare and accurate in detection result.
Disclosure of Invention
The invention aims to solve at least one technical problem in the prior art and provides a method for manufacturing a fluorescent microsphere test piece. In the method, the fluorescent microsphere solution is dripped into the raw material accommodating area to naturally level and air-dry to form a layer of tiled fluorescent microspheres, so that the fluorescent microspheres are prevented from being stacked and piled on any longitudinal arrangement, and accurate detection results can be obtained; the thickness of the finally prepared fluorescent microsphere test piece is controlled by the measuring scale, so that fluorescent microsphere test pieces with different thicknesses can be obtained according to requirements.
The specific scheme of the invention is as follows:
a method for manufacturing a fluorescent microsphere test piece comprises the following steps:
s1, horizontally placing a glass slide, and horizontally placing two or more than two measuring scales on the glass slide at intervals to form a raw material accommodating area;
s2, dripping a fluorescent microsphere solution into the raw material containing area in the step S1, and naturally leveling the dripped fluorescent microsphere solution to enable fluorescent microspheres in the fluorescent microsphere solution to be tiled into a layer;
s3, air-drying the fluorescent microsphere solution in the step S2 to obtain fluorescent microspheres;
s4, dripping resin into the raw material accommodating area on the fluorescent microspheres obtained in the step S3, and then covering a cover glass to ensure that the closed space formed by the glass slide, the measuring ruler and the cover glass is filled without gaps and the fluorescent microspheres are completely wrapped;
s5 curing the resin in the closed space in the step S4;
s6, peeling the cover glass on the resin cured in the step S5 to form a fluorescent microsphere test piece comprising the resin, the fluorescent microsphere layer and the glass slide.
The fluorescent microsphere test piece with the preset thickness and the non-laminated and non-accumulated fluorescent microspheres in any longitudinal arrangement can be obtained through the steps.
The invention also provides a fluorescent microsphere test piece which is manufactured by the manufacturing method and comprises a glass slide, resin and fluorescent microspheres, wherein the resin is arranged on the glass slide, the fluorescent microspheres are embedded into the resin so that the resin completely wraps the fluorescent microspheres, and the fluorescent microspheres are tangent to the glass slide.
The invention also provides a manufacturing assembly for manufacturing the fluorescent microsphere test strip, which comprises:
the raw material accommodating area forming assembly comprises a glass slide and measuring scales, wherein the measuring scales are vertically arranged on the glass slide, and 2 or more measuring scales are arranged at intervals in parallel to form a raw material accommodating area;
the glass slide comprises a raw material, a fluorescent microsphere solution and resin, wherein the fluorescent microsphere solution is dripped into the raw material accommodating area, air-dried and then forms a single-layer fluorescent microsphere on the glass slide, and the resin is dripped into the raw material accommodating area from the upper part of the fluorescent microsphere; the cover glass is arranged on the measuring scale, so that resin in a closed space formed by the glass slide, the measuring scale and the cover glass is filled without gaps, and the fluorescent microspheres are completely wrapped;
a curing device for curing the resin in the enclosed space;
and the stripping device is used for stripping the cover glass on the cured resin.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings needed to be used in the embodiments will be briefly described below, it should be understood that the following drawings only illustrate some embodiments of the present invention and therefore should not be considered as limiting the scope, and for those skilled in the art, other related drawings can be obtained according to the drawings without inventive efforts.
FIG. 1 is a flow chart of a method for manufacturing a fluorescent microsphere test strip according to the present invention;
FIG. 2 is a schematic diagram of a manufacturing assembly of a fluorescent microsphere test strip according to the present invention;
FIG. 3 is a schematic structural diagram of a fluorescent microsphere test strip provided in the present invention;
FIG. 4 is an image of a fluorescent microsphere at a longitudinal cross-section tested for axial resolution using the fluorescent microsphere test strip of FIG. 3;
FIG. 5 is axial resolution data obtained using the fluorescent microsphere test strip of FIG. 3.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are some, but not all, embodiments of the present invention. The components of embodiments of the present invention generally described and illustrated in the figures herein may be arranged and designed in a wide variety of different configurations.
Thus, the following detailed description of the embodiments of the present invention, presented in the figures, is not intended to limit the scope of the invention, as claimed, but is merely representative of selected embodiments of the invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
It should be noted that: like reference numbers and letters refer to like items in the following figures, and thus, once an item is defined in one figure, it need not be further defined and explained in subsequent figures.
In the description of the present invention, it should be noted that the terms "middle", "upper", "lower", "left", "right", "inner", "outer", etc. indicate orientations or positional relationships based on the orientations or positional relationships shown in the drawings or the orientations or positional relationships that the products of the present invention are usually placed in when used, and are only used for convenience of describing the present invention and simplifying the description, but do not indicate or imply that the devices or elements referred to must have specific orientations, be constructed in specific orientations, and be operated, and thus, should not be construed as limiting the present invention. Furthermore, the terms "first," "second," "third," and the like are used solely to distinguish one from another and are not to be construed as indicating or implying relative importance.
Furthermore, the terms "horizontal", "vertical", "overhang" and the like do not imply that the components are required to be absolutely horizontal or overhang, but may be slightly inclined. For example, "horizontal" merely means that the direction is more horizontal than "vertical" and does not mean that the structure must be perfectly horizontal, but may be slightly inclined.
In the description of the present invention, it should also be noted that, unless otherwise explicitly specified or limited, the terms "disposed," "mounted," "connected," and "connected" are to be construed broadly and may, for example, be fixedly connected, detachably connected, or integrally connected; can be mechanically or electrically connected; they may be connected directly or indirectly through intervening media, or they may be interconnected between two elements. The specific meanings of the above terms in the present invention can be understood in specific cases to those skilled in the art.
As shown in FIG. 1, the present invention provides a method for manufacturing a fluorescent microsphere test strip, comprising:
s1, horizontally placing a glass slide, and horizontally placing two or more than two measuring scales on the glass slide at intervals to form a raw material accommodating area;
s2, dripping a fluorescent microsphere solution into the raw material containing area in the step S1, and naturally leveling the dripped fluorescent microsphere solution to enable fluorescent microspheres in the fluorescent microsphere solution to be tiled into a layer;
s3, air-drying the fluorescent microsphere solution in the step S2 to obtain fluorescent microspheres;
s4, dripping resin into the raw material accommodating area on the fluorescent microspheres obtained in the step S3, and then covering a cover glass to ensure that the closed space formed by the glass slide, the measuring ruler and the cover glass is filled without gaps and the fluorescent microspheres are completely wrapped;
s5 curing the resin in the closed space in the step S4;
s6, peeling the cover glass on the resin cured in the step S5 to form a fluorescent microsphere test piece comprising the resin, the fluorescent microsphere layer and the glass slide.
In the method, a raw material accommodating area with preset height and size is formed through a glass slide and a measuring ruler, then a fluorescent microsphere solution is dripped into the raw material accommodating area, the fluorescent microsphere solution is preferably paved into a layer after natural leveling, then the fluorescent microsphere solution is air-dried to obtain fluorescent microspheres, then resin is added onto the fluorescent microspheres, the amount of the resin is added to fill a closed space formed by the raw material accommodating area after a cover glass is covered, so that no gap is formed, and the fluorescent microspheres are preferably completely wrapped; and further curing the resin and peeling the cover glass to obtain the fluorescent microsphere test piece. The method is simple and easy to operate, ensures that the fluorescent microspheres are not stacked or piled on any longitudinal arrangement, and can control the thickness of the finally prepared fluorescent microsphere test piece through a measuring scale, thereby obtaining the fluorescent microsphere test pieces with different thicknesses according to requirements.
In this embodiment, an axial resolution of the confocal endoscope is measured as an example, and further, a measuring scale with a thickness smaller than a working distance of the confocal endoscope is selected for the measuring scale in step S1, in this embodiment, since the working distance of the confocal endoscope to be detected belongs to a micrometer scale, a micrometer-scale feeler gauge is selected as the measuring scale, a refractive index range of the resin is 1.33-1.40, and the thickness of the feeler gauge is smaller than the working distance of the confocal endoscope, so that the fluorescent microspheres can be imaged during measurement, thereby ensuring smooth detection; the resin is selected in the range, and mainly when the confocal endoscope is designed, the object space of the objective lens of the confocal endoscope is assumed to be water, and the refractive index of the water is about 1.33, so that the test strip can be ensured to be matched with the working condition of the micro objective lens at the far end of the confocal endoscope.
Further, the specific method for obtaining the fluorescent microsphere solution in step S2 is as follows: the fluorescent microsphere solution with the solution concentration of 0.3 percent is diluted by 1000-10000 times. When the concentration of the directly purchased fluorescent microsphere solution is too high, it needs to be diluted, such as InSpeck from SammerfellTMGreen series.
Further, the specific method for curing in step S5 is as follows: the resin was irradiated with a uniform ultraviolet light source in the direction from the slide to the cover glass. The "irradiation from the slide to the cover glass" can cure the resin close to the slide more thoroughly, so that the resin is more firmly bonded to the slide and more easily separated from the cover glass.
Specifically, to finally obtain a fluorescent microsphere test piece with a resin thickness of 40um, the parameters are selected as follows: the feeler in the step S1 needs to be a feeler of 40um, and the refractive index range of the resin is 1.33; the fluorescent microsphere solution with the solution concentration of 0.3% is diluted by 10000 times.
The thickness of the feeler and the refractive index of the resin can be adjusted according to specific conditions, the manufacturing method of the corresponding fluorescence test piece is the same as the refractive index of the resin of 1.33 and the thickness of the feeler of 40um, and only the refractive index of the resin or the thickness parameter of the feeler is modified. Whether the concentration of the fluorescent microsphere solution needs to be diluted or not and how many times the concentration needs to be diluted are determined according to actual conditions, and the concentration is determined by only meeting the requirement that the dripped fluorescent microsphere solution naturally levels to enable the fluorescent microspheres in the fluorescent microsphere solution to be tiled into a layer.
As shown in FIG. 2, the present invention further provides a manufacturing assembly for manufacturing the above-mentioned fluorescent microsphere test strip, comprising:
the raw material accommodating area forming assembly comprises a glass slide 1 and measuring scales 2, wherein the measuring scales are vertically arranged on the glass slide, and 2 or more measuring scales are arranged at intervals in parallel to form a raw material accommodating area;
the glass slide comprises a raw material, a fluorescent microsphere solution 3 and resin 4, wherein the fluorescent microsphere solution is dripped into the raw material accommodating area, air-dried and then forms a single-layer fluorescent microsphere on the glass slide, and the resin is dripped into the raw material accommodating area from the upper part of the fluorescent microsphere;
the cover glass 5 is arranged on the measuring scale, so that resin in a closed space formed by the glass slide, the measuring scale and the cover glass is filled without gaps, and the fluorescent microspheres are completely wrapped;
a curing device 6 for curing the resin in the closed space;
and the stripping device is used for stripping the cover glass on the cured resin.
The working principle of the manufacturing assembly is as follows: the assembly is formed through the raw material containing area to construct the raw material containing area, raw materials are added into the raw material containing area according to a certain sequence and requirements, then a cover glass is covered, so that a closed space which is filled with resin and has no gap and is completely wrapped by the fluorescent microspheres is formed, and finally the cover glass is peeled through a peeling device after the resin is cured through a curing device. The curing device in the manufacturing assembly can be an ultraviolet lamp or other devices capable of achieving resin curing, and the stripping device can be tweezers or other devices for realizing resin stripping of the cover glass.
Furthermore, because the working distance of the confocal endoscope to be detected in the embodiment belongs to the micron level, a micron-level clearance gauge is selected as the measuring scale, the thickness of the clearance gauge is 20um to 50um, and the refractive index of the resin is 1.33 to 1.40; the concentration of the fluorescent microsphere solution is 0.00001-0.001%, and the thickness of the resin in the final fluorescent microsphere test piece is within the range by using the feeler gauge within the thickness range, so that the finally obtained fluorescent microsphere test piece can be ensured to be a confocal endoscope with the measuring working distance of 70-100 um; the concentration of the fluorescent microsphere solution is 0.00001% -0.001%, so that when the fluorescent microsphere solution is directly dripped, fluorescent microspheres in the solution are not stacked after the solution is leveled; the resin is selected in the range, and mainly when the confocal endoscope is designed, the object space of the objective lens of the confocal endoscope is assumed to be water, and the refractive index of the water is about 1.33, so that the test strip can be ensured to be matched with the working condition of the micro objective lens at the far end of the confocal endoscope.
Further, the curing device is a uniform ultraviolet light source. In this example, an LED UV lamp with a wavelength of 365nm and a power of 5W is selected, and the curing time is 3-5 minutes.
Further, the device comprises an air drying device which is an oven and is used for air drying the fluorescent microsphere solution to obtain the fluorescent microspheres. The time for preparing the fluorescent microsphere test piece can be shortened by using an oven for air drying.
As shown in fig. 3, the invention further provides a fluorescent microsphere test piece, which is manufactured by the manufacturing assembly shown in fig. 2 according to the manufacturing method shown in fig. 1, and comprises a glass slide 1, resin 31 and fluorescent microspheres 21, wherein the resin is arranged on the glass slide, the fluorescent microspheres are embedded in the resin so that the resin completely wraps the fluorescent microspheres, and the fluorescent microspheres are tangent to the glass slide. The fluorescent microspheres are tangent to the glass slide, so that the fluorescent microspheres are not stacked or piled on any longitudinal arrangement. The fluorescent microsphere test piece is prepared from resin and fluorescent microspheres, and the performance of the resin and the fluorescent microspheres is stable, so that the fluorescent microsphere test piece can be stored for a long time.
Further, the refractive index of the resin is 1.33, and the thickness of the resin is 40 um. The thickness of the resin is 40um, so that the finally obtained fluorescent microsphere test piece can be used for measuring a confocal endoscope with the working distance of 70-100 um. The resin is selected in the range, and mainly when the confocal endoscope is designed, the object space of the objective lens of the confocal endoscope is assumed to be water, and the refractive index of the water is about 1.33, so that the test strip can be ensured to be matched with the working condition of the micro objective lens at the far end of the confocal endoscope.
Although the refractive index of the resin is 1.33 and the thickness of the resin is 40um to manufacture the fluorescent microsphere test piece in this embodiment, according to the actual requirement, the refractive index of the resin is 1.4 or 1.37, the thickness of the resin is 20um or 30um, and the like, and the manufacturing method and the structure thereof are both 1.33 and 40um, and only the refractive index of the resin or the thickness parameter of the resin is modified.
FIG. 4 shows an image of a fluorescent microsphere at a longitudinal cross-section taken with the fluorescent microsphere test strip of FIG. 3 for axial resolution testing, where the lighter piece represents the image of the fluorescent microsphere and the background is the image of the end face of the image-transmitting fiber bundle; fig. 5 is axial resolution data obtained by processing a series of fluorescent microsphere images, specifically, setting a movement step (i.e., distance/pixel) of a motor to 1um, and then obtaining light intensity (i.e., gray value) of the fluorescent microsphere every other step, i.e., obtaining a curve with abscissa as distance/pixel and ordinate as gray value of the fluorescent microsphere, where the curve is manually identified to have full width at half maximum (or gaussian curve fitting), so as to obtain an axial resolution of the confocal endoscope obtained by using the fluorescent microsphere test strip of 7 um. Fig. 4 and 5 actually demonstrate that the thickness of the fluorescent microsphere test strip is 40um, and the fluorescent microsphere test strip provided by the invention can be used for measuring the axial resolution of a confocal endoscope with the working distance of less than 100 um.
The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (8)
1. The manufacturing method of the fluorescent microsphere test strip is characterized in that the fluorescent microsphere test strip is used for measuring the axial resolution of a confocal endoscope with the working distance of 70-100 mu m, and comprises the following steps:
s1, horizontally placing a glass slide, and horizontally placing two or more than two measuring scales on the glass slide at intervals to form a raw material accommodating area, wherein the measuring scales are micrometer-sized clearance gauges with the thickness of 20-50 um;
s2, diluting the fluorescent microsphere solution with the solution concentration of 0.3% by 1000-10000 times, dripping the fluorescent microsphere solution into the raw material accommodating area in the step S1, and naturally leveling the dripped fluorescent microsphere solution to enable the fluorescent microspheres in the fluorescent microsphere solution to be scattered and spread into a layer;
s3, air-drying the fluorescent microsphere solution in the step S2 to obtain fluorescent microspheres;
s4, dripping resin on the fluorescent microspheres obtained in the step S3 to the raw material containing area, wherein the refractive index of the resin is 1.33-1.40, and then covering a cover glass to ensure that the closed space formed by the glass slide, the measuring ruler and the cover glass is filled without gaps and the fluorescent microspheres are completely wrapped;
s5 curing the resin in the closed space in the step S4;
s6, peeling the cover glass on the resin cured in the step S5 to form a fluorescent microsphere test piece comprising the resin, the fluorescent microsphere layer and the glass slide.
2. The method as claimed in claim 1, wherein the fluorescent microsphere test strip is used for measuring the axial resolution of a confocal endoscope, and the measuring scale in step S1 is a measuring scale with a thickness smaller than the working distance of the confocal endoscope.
3. The manufacturing method according to claim 2, wherein the specific method for curing in step S5 is: the resin was irradiated with a uniform ultraviolet light source in the direction from the slide to the cover glass.
4. A fluorescent microsphere test piece is characterized by being manufactured by any one of the manufacturing methods of claims 1 to 3 and comprising a glass slide, resin and fluorescent microspheres, wherein the refractive index of the resin is 1.33 to 1.40, the thickness of the resin is 20um to 50um, the resin is arranged on the glass slide, the fluorescent microspheres are embedded into the resin so that the resin completely wraps the fluorescent microspheres, and the fluorescent microspheres are tangent to the glass slide.
5. A manufacturing assembly for manufacturing the fluorescent microsphere test strip of claim 4, wherein the fluorescent microsphere test strip is used for measuring the axial resolution of a confocal endoscope with a working distance of 70-100 μm, the manufacturing assembly comprises:
the raw material accommodating area forming assembly comprises a glass slide and measuring scales, wherein the measuring scales are vertically arranged on the glass slide, 2 or more measuring scales are arranged at intervals in parallel to form a raw material accommodating area, the measuring scales are micrometer-sized clearance gauges, and the thickness of the measuring scales is 20-50 micrometers;
the glass slide comprises a raw material, a fluorescent microsphere solution and resin, wherein the fluorescent microsphere solution is dripped into the raw material accommodating area, air-dried and then forms a single-layer scattered fluorescent microsphere on the glass slide, and the resin is dripped into the raw material accommodating area from the upper part of the fluorescent microsphere;
the cover glass is arranged on the measuring scale, so that resin in a closed space formed by the glass slide, the measuring scale and the cover glass is filled without gaps, the fluorescent microspheres are completely wrapped, and the refractive index of the resin is 1.33-1.40;
a curing device for curing the resin in the enclosed space;
and the stripping device is used for stripping the cover glass on the cured resin.
6. The fabrication assembly of claim 5, wherein the concentration of the fluorescent microsphere solution is 0.00001% to 0.001%.
7. The fabrication assembly of claim 5, wherein the curing device is a uniform ultraviolet light source.
8. The manufacturing assembly of claim 5, further comprising an air drying device, wherein the air drying device is an oven for air drying the fluorescent microsphere solution to obtain fluorescent microspheres.
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