技术领域technical field
本发明涉及一种显微镜，特别涉及一种新型外置荧光模块的显微镜。The invention relates to a microscope, in particular to a novel microscope with an external fluorescent module.
背景技术Background technique
目前的荧光显微镜的荧光模块都是在物镜与目镜之间，采用汞灯、氙灯或是卤素灯作为光源，激发光通过柯拉照明系统到达滤色块组(滤色块组由激发滤光片、二向分色片、发射滤光片组成)，光源经激发滤光片得到所需激发光谱，经二向分色片反射经过物镜射至样品上，被荧光染料染色的样品受到激发后，发射波长大于激发光的光谱。发射光经过物镜、二向分色片、发射滤光片(发射光以外波段的光谱被截止)到达目镜或成像装置。The fluorescence module of the current fluorescence microscope is all between the objective lens and the eyepiece, using a mercury lamp, a xenon lamp or a halogen lamp as a light source, and the excitation light reaches the color filter group through the Kohler illumination system (the color filter group consists of an excitation filter , dichroic dichroic film, and emission filter), the light source is passed through the excitation filter to obtain the required excitation spectrum, reflected by the dichroic dichroic film and shot onto the sample through the objective lens, after the sample dyed by the fluorescent dye is excited, The emission wavelength is greater than the spectrum of the excitation light. The emitted light passes through the objective lens, the dichroic filter, and the emission filter (the spectrum of the wavelength band other than the emitted light is cut off) to the eyepiece or imaging device.
目前，公开号为CN104035195A的专利公开了一种荧光显微镜，包括架体、载物台和底座，所述架体、载物台和底座从上到下依次设置，所述架体周侧设有一个以上的调节旋钮，所述架体下部设有物镜，所述载物台位于物镜的正下方位置，所述载物台底部位置依次设置有聚光器、光阑和固定支架，所述底座上的中部位置安装有LED荧光激发光源，所述LED荧光激发光源正对载物台的中部位置，所述LED荧光激发光源周围设有一个以上的反光镜旋转架，所述LED荧光激发光源包括有灯罩，所述灯罩上设有屏蔽罩，所述屏蔽罩与LED荧光激发光源活动连接，所述各个反光镜旋转架上皆安装有反光镜；该荧光显微镜通过设置有多个聚光设备来加强荧光效果。公开号为CN202735583A的专利公开了一种荧光显微镜，包括物镜，物镜安装在物镜转化器上，物镜转化器与目镜筒连接，目镜筒安装在镜臂上，镜臂安装在固定台上，载物台与固定台连接，载物台上方安装有标本推进尺，标本推进尺与标本推进尺螺旋连接，载物台下方安装有聚光器，聚光器与光阑连接，固定台上安装有粗调螺旋和细调螺旋，固定台安装在镜座上，所述的镜座上安装有LED荧光激发光源，其照明方式为落射式，即光源要通过物镜投射于样品上，但这样会导致光损耗加大。At present, the patent with the publication number CN104035195A discloses a fluorescent microscope, which includes a frame, a stage and a base, and the frame, the stage and the base are arranged in sequence from top to bottom, and the periphery of the frame is provided with More than one adjustment knob, the lower part of the frame body is provided with an objective lens, the stage is located directly below the objective lens, the bottom of the stage is provided with a condenser, a diaphragm and a fixed bracket in sequence, and the base An LED fluorescent excitation light source is installed at the middle position on the top, and the LED fluorescent excitation light source is facing the middle position of the stage, and more than one reflector rotating frame is arranged around the LED fluorescent excitation light source, and the LED fluorescent excitation light source includes There is a lampshade, and a shielding cover is arranged on the lampshade, and the shielding cover is movably connected with the LED fluorescent excitation light source, and reflectors are installed on each of the reflector rotating frames; the fluorescence microscope is equipped with multiple light-gathering devices to Enhance the fluorescent effect. The patent with the publication number CN202735583A discloses a fluorescence microscope, including an objective lens, the objective lens is installed on the objective lens converter, the objective lens converter is connected with the eyepiece tube, the eyepiece tube is installed on the mirror arm, and the mirror arm is installed on the fixed table, and the object The stage is connected with the fixed stage, the specimen advancing ruler is installed on the top of the stage, the specimen advancing ruler is screw connected with the specimen advancing ruler, the condenser is installed under the stage, the condenser is connected with the diaphragm, and the coarse Adjusting screw and fine-tuning screw, the fixed table is installed on the mirror base, and the LED fluorescent excitation light source is installed on the mirror base. Loss increased.
传统显微镜的荧光模块位于显微镜物镜与成像透镜之间，因光阑、各种透镜以及物镜的影响，所得到的光斑光损耗很大，有时达到90％，导致光强不够或光源视场小。The fluorescence module of a traditional microscope is located between the microscope objective lens and the imaging lens. Due to the influence of the diaphragm, various lenses and objective lenses, the resulting spot light loss is very large, sometimes reaching 90%, resulting in insufficient light intensity or a small field of view of the light source.
但对于某些生物实验而言，需要大面积、光场均匀的样品照明。如数字PCR，其样品为包含有几万甚至上百万份反应单元，要对这些反应单元同时进行观测分析，样品照明须同时满足以下三个条件：(1)大视场光源，(2)光强大，(3)光强均匀。同时满足这三个条件才可以对这些大数量的反应单元的样品进行同时观测分析，尤其面积大，光强大是首要考虑的因素。而传统的照明装置视场很小，无法满足这一需求；虽然传统的显微镜可以通过采用低倍率的物镜来增大样品视场，但这样得到的样品照明的强度以视场的增大倍数的二次幂减小，同样无法满足数字PCR分析的需要。因而促使我们开发一种新型外置荧光模块的显微镜照明装置。But for some biological experiments, sample illumination with large area and uniform light field is required. Such as digital PCR, the sample contains tens of thousands or even millions of reaction units. To observe and analyze these reaction units at the same time, the sample illumination must meet the following three conditions at the same time: (1) large field of view light source, (2) The light is strong, and (3) the light intensity is uniform. Only when these three conditions are met at the same time can the samples of these large number of reaction units be observed and analyzed at the same time, especially when the area is large, the light intensity is the primary consideration. The field of view of the traditional lighting device is very small, which cannot meet this requirement; although the traditional microscope can increase the field of view of the sample by using a low-magnification objective lens, the intensity of the sample illumination obtained in this way is equal to the increase of the field of view. The reduction of the power of two also cannot meet the needs of digital PCR analysis. Therefore, we are prompted to develop a new type of microscope illumination device with an external fluorescent module.
发明内容Contents of the invention
本发明的目的是为解决传统显微镜光损耗大、光源视场小、光强不足等问题，提供一种光源视场大、光强大、光强均匀的照明装置，其技术方案如下：The purpose of the present invention is to solve the problems of traditional microscopes such as large light loss, small light source field of view, and insufficient light intensity, and provide a lighting device with a large light source field of view, strong light, and uniform light intensity. The technical solution is as follows:
新型外置荧光模块的显微镜，其照明装置包括一个外置光源和滤光片组构成的荧光模块。A new type of microscope with an external fluorescent module, the illuminating device of which comprises a fluorescent module composed of an external light source and a filter set.
所述荧光模块置于物镜与样品之间，位于物镜的正下方。The fluorescent module is placed between the objective lens and the sample, directly below the objective lens.
外置光源选自汞灯、卤素灯、氙灯或LED灯中的一种。The external light source is selected from one of mercury lamp, halogen lamp, xenon lamp or LED lamp.
外置光源为LED灯时，所述新型外置荧光模块的显微镜还包括集光装置。When the external light source is an LED lamp, the microscope of the novel external fluorescent module also includes a light collecting device.
所述的滤光片组由激发滤光片、二向分色片、发射滤光片组成。激发滤光片与发射滤光片成90度垂直，二向分色片平分激发滤光片与发射滤光片所成的直角，二向分色片与显微镜的光轴成45度角。The filter group is composed of an excitation filter, a dichroic dichroic filter and an emission filter. The excitation filter is 90 degrees perpendicular to the emission filter, the dichroic dichroic filter bisects the right angle formed by the excitation filter and the emission filter, and the dichroic dichroic filter is at a 45 degree angle to the optical axis of the microscope.
激发滤光片的作用是将激发光滤波，得到窄带激发波段，避免光源中的长波段光进入滤光片组。二向分色片的作用是将短波长光反射，长波长光透射。发射滤光片是将荧光发射的长波长光透射，滤掉短波长的光。The function of the excitation filter is to filter the excitation light to obtain a narrow-band excitation band, so as to prevent the long-wave band light in the light source from entering the filter group. The function of the dichroic filter is to reflect short-wavelength light and transmit long-wavelength light. The emission filter transmits the long-wavelength light emitted by the fluorescence and filters out the short-wavelength light.
所述滤光片组针对不同荧光基团采用不同光谱特性的滤光片。外置光源为LED灯时，此时不同光谱特性的滤光片对应不同波段的LED灯。The filter set uses filters with different spectral characteristics for different fluorescent groups. When the external light source is an LED light, the filters with different spectral characteristics correspond to LED lights of different bands.
本发明的新型外置荧光模块的显微镜的光源路径依次为外置光源，荧光模块，样品，荧光模块，物镜，目镜。相比传统的显微镜，本发明的新型外置荧光模块的显微镜光损耗非常小，得到的光斑面积及光强度大。The light source path of the novel microscope with an external fluorescent module of the present invention is an external light source, a fluorescent module, a sample, a fluorescent module, an objective lens, and an eyepiece in sequence. Compared with the traditional microscope, the microscope light loss of the novel external fluorescent module of the present invention is very small, and the obtained light spot area and light intensity are large.
本发明尤其适用于需要样品视场大，荧光信号弱的生物信号检测。The invention is especially suitable for the detection of biological signals that require a large sample field of view and weak fluorescence signals.
本发明提供的外置荧光模块的照明装置有如下优点：The lighting device with an external fluorescent module provided by the present invention has the following advantages:
1、光斑面积大，传统的显微镜，光斑面积不超过0.5cm2，而本发明的显微镜能产生至少2cm2的光斑，还可定制荧光模块，产生9cm2以上的光斑。1. The spot area is large. Traditional microscopes have a spot area of no more than 0.5 cm2 , but the microscope of the present invention can produce a spot of at least 2 cm2 . The fluorescent module can also be customized to produce a spot of more than 9 cm2 .
2、光损耗小，光强大，本发明的显微镜将荧光模块置于物镜与样品之间，其光损耗仅为30％，相比传统显微镜高达90％的光损耗，本发明的显微镜可有效提高光源的强度。2. The light loss is small and the light is strong. The microscope of the present invention places the fluorescent module between the objective lens and the sample, and its light loss is only 30%. Compared with the light loss of traditional microscopes up to 90%, the microscope of the present invention can effectively improve The intensity of the light source.
3、如采用LED灯作为外置光源还有功率低、价格低、寿命长、开机无需预热、关机无需冷却等优势。3. If LED lights are used as external light sources, there are advantages such as low power, low price, long life, no need to warm up when starting up, and no need to cool down when turning off.
附图说明Description of drawings
图1为本发明新型外置荧光模块的显微镜的结构图。Fig. 1 is a structural diagram of a microscope with a novel external fluorescence module of the present invention.
图2为本发明新型外置荧光模块的显微镜的光路流程图。Fig. 2 is a flow chart of the optical path of the microscope with a new type of external fluorescence module of the present invention.
图3为传统显微镜的结构图。Figure 3 is a structural diagram of a conventional microscope.
图4为传统显微镜的光路流程图。Figure 4 is a flow chart of the optical path of a conventional microscope.
具体实施方式detailed description
现结合附图和实施例对本发明作进一步说明。The present invention will be further described in conjunction with accompanying drawing and embodiment now.
实施例1Example 1
参照图1和图2所示，荧光模块1包括激发滤光片11、二向分色片12、发射滤光片13组成。荧光模块置于物镜3与样品2之间，位于物镜3的正下方，激发滤光片11与发射滤光片13成90度垂直，二向分色片12平分激发滤光片11与发射滤光片13所成的直角，二向分色片12与显微镜的光轴成45度角。Referring to FIGS. 1 and 2 , the fluorescent module 1 includes an excitation filter 11 , a dichroic filter 12 and an emission filter 13 . The fluorescence module is placed between the objective lens 3 and the sample 2, and is located directly below the objective lens 3. The excitation filter 11 is perpendicular to the emission filter 13 at 90 degrees, and the dichroic filter 12 bisects the excitation filter 11 and the emission filter. The light sheet 13 forms a right angle, and the dichroic sheet 12 forms an angle of 45 degrees with the optical axis of the microscope.
首先，外置光源5发出的光进入荧光模块1，经激发滤光片11滤波，得到特定波段短波长的光，该特定波段短波长的光射到与显微镜光轴成45度角放置的二向分色片12上，此时二向分色片12表现出反射特性，将该短波长的光反射到样品2上，在样品2上受到激发后，发射波长大于激发光的光谱，然后再透过二向分色片12，经发射滤光片13，被物镜3接收，再进入目镜4或探测装置。First, the light emitted by the external light source 5 enters the fluorescent module 1, and is filtered by the excitation filter 11 to obtain light with a specific band and short wavelength. On the dichroic film 12, at this time, the dichroic dichroic film 12 shows reflection characteristics, and the short-wavelength light is reflected onto the sample 2, and after being excited on the sample 2, the emission wavelength is greater than the spectrum of the exciting light, and then It passes through the dichroic filter 12, passes through the emission filter 13, is received by the objective lens 3, and then enters the eyepiece 4 or the detection device.
本发明的外置荧光模块的显微镜光源路径依次为外置光源，荧光模块，样品，荧光模块，物镜，目镜。The microscope light source path of the external fluorescent module of the present invention is an external light source, a fluorescent module, a sample, a fluorescent module, an objective lens, and an eyepiece in sequence.
采用本实施例的方法，由于将荧光模块置于物镜3与样品2之间，使得光源到达样品的光路变短，避免了光源经过光阑、各种透镜以及物镜的汇聚作用而导致的光损耗，因此，可以把光损耗降到最低。当激发滤光片11、二向分色片12、发射滤光片13的尺寸为3cm×3cm时，可得到的光斑面积为9cm2，采用3W的蓝光LED，此时光强达到20mW/cm2以上。With the method of this embodiment, since the fluorescent module is placed between the objective lens 3 and the sample 2, the optical path from the light source to the sample is shortened, and the light loss caused by the convergence of the light source through the diaphragm, various lenses and the objective lens is avoided. , therefore, the optical loss can be minimized. When the dimensions of the excitation filter 11, the dichroic filter 12, and the emission filter 13 are 3cm×3cm, the available spot area is 9cm2 , and a 3W blue LED is used, and the light intensity reaches 20mW/cm2 above.
实施例2Example 2
参照图3和图4所示，首先，外置光源5发出的光透过光阑6和透镜7，进入荧光模块1，经激发滤光片11滤波，得到短波长的光，短波长的光射到二向分色片12上，此时二向分色片表现出反射特性，将短波长的光反射到物镜3上，经物镜3照射到样品2上，在样品2上激发出长波长的光再透过物镜3进入荧光模块1，经发射滤光片13，然后进入目镜4或探测装置。3 and 4, first, the light emitted by the external light source 5 passes through the diaphragm 6 and the lens 7, enters the fluorescent module 1, and is filtered by the excitation filter 11 to obtain short-wavelength light, short-wavelength light irradiated on the dichroic film 12, at this time the dichroic film exhibits reflective properties, reflecting the short-wavelength light to the objective lens 3, and then irradiating it to the sample 2 through the objective lens 3, and excites the long-wavelength light on the sample 2 The light then enters the fluorescent module 1 through the objective lens 3, passes through the emission filter 13, and then enters the eyepiece 4 or the detection device.
传统显微镜的光源路径依次为外置光源，荧光模块，物镜，样品，物镜，荧光模块，目镜。The light source path of a traditional microscope is an external light source, a fluorescent module, an objective lens, a sample, an objective lens, a fluorescent module, and an eyepiece.
传统显微镜的光斑的面积一般不足1cm2，其光路图需要透过多个光阑和透镜，加之物镜的影响，所得到的光斑光损耗很大，有时达到90％，采用3W的蓝光LED，光强仅为1mW/cm2。The area of the light spot of a traditional microscope is generally less than 1cm2 , and its light path diagram needs to pass through multiple diaphragms and lenses. In addition to the influence of the objective lens, the light loss of the obtained light spot is very large, sometimes reaching 90%. Using a 3W blue LED, the light The intensity is only 1mW/cm2 .
以上对本发明实施例所提供的新型外置荧光模块的显微镜，进行了详细介绍，本文中应用了具体个例对本发明的原理及实施方式进行了阐述，以上实施例的说明只是用于帮助理解本发明的方法及其核心思想；同时，对于本领域的一般技术人员，依据本发明的思想，在具体实施方式及应用范围上均会有改变之处，综上所述，本说明书内容不应理解为对本发明的限制。The above is a detailed introduction to the microscope with a new type of external fluorescent module provided by the embodiment of the present invention. In this paper, specific examples are used to illustrate the principle and implementation of the present invention. The description of the above embodiment is only used to help understand the present invention. The method of the invention and its core idea; at the same time, for those of ordinary skill in the art, according to the idea of the present invention, there will be changes in the specific implementation and scope of application. In summary, the content of this specification should not be understood To limit the present invention.
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510338469.0ACN104865688B (en) | 2015-06-17 | 2015-06-17 | The microscope of external fluorescent moieties |
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510338469.0ACN104865688B (en) | 2015-06-17 | 2015-06-17 | The microscope of external fluorescent moieties |
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| CN104865688A CN104865688A (en) | 2015-08-26 |
| CN104865688Btrue CN104865688B (en) | 2016-01-27 |
| Application Number | Title | Priority Date | Filing Date |
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| CN201510338469.0AActiveCN104865688B (en) | 2015-06-17 | 2015-06-17 | The microscope of external fluorescent moieties |
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