WO2021189453A1 - 一种微型荧光显微成像模块 - Google Patents

一种微型荧光显微成像模块 Download PDF

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Publication number
WO2021189453A1
WO2021189453A1 PCT/CN2020/081796 CN2020081796W WO2021189453A1 WO 2021189453 A1 WO2021189453 A1 WO 2021189453A1 CN 2020081796 W CN2020081796 W CN 2020081796W WO 2021189453 A1 WO2021189453 A1 WO 2021189453A1
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Prior art keywords
lens
objective lens
excitation light
light source
fluorescence microscopy
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PCT/CN2020/081796
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English (en)
French (fr)
Inventor
胡庆磊
黄凯
李宁
李梦婷
李培
Original Assignee
肯维捷斯（武汉）科技有限公司
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Priority to PCT/CN2020/081796 priority Critical patent/WO2021189453A1/zh
Publication of WO2021189453A1 publication Critical patent/WO2021189453A1/zh

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    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/06—Means for illuminating specimens
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/36—Microscopes arranged for photographic purposes or projection purposes or digital imaging or video purposes including associated control and data processing arrangements

Definitions

  • the invention relates to the field of optical imaging, in particular to a miniature fluorescent microscopic imaging module.
  • Fluorescence microscope In the field of life sciences, fluorescence microscopes are often used. Fluorescence microscope usually includes light source, excitation filter, dichroic lens, objective lens, emission filter, tube lens, detector and other devices.
  • the light source passes through the excitation filter to obtain the excitation light, which is reflected by the dichroic mirror into the objective lens, and is focused on the sample surface.
  • the excited fluorescence signal is collected by the objective lens, and then imaged by the tube lens to the detector target surface after passing through the dichroic mirror and the filter.
  • This kind of microscope usually has a certain magnification, which will inevitably result in short object distance and long image distance. There must be enough space in the image space to meet the requirements of this optical path setting.
  • the light source needs to be connected to the mains power
  • the detectors such as CCD or CMOS also need to be connected to the mains or computer power supply
  • the image display and storage also need to be connected to the computer.
  • the microscope must also have a focusing mechanism. Whether it is manual focusing or automatic focusing, its mechanical structure is relatively complicated and takes up a lot of space. For these reasons, fluorescence microscopes are usually large in size, complex in structure, high in cost, and not portable.
  • the core components that can be imaged by the fluorescence microscope are the objective lens and the tube lens.
  • the main factor restricting its inability to miniaturize is the large magnification of the system, which results in the long focal length of the tube lens.
  • the filter set still uses the previous settings for miniaturization, without further consideration of its miniaturization design.
  • the technical problem to be solved by the present invention is to provide a miniature fluorescent microscopic imaging module in view of the defects of the prior art.
  • the technical solution adopted by the present invention to solve its technical problem is: constructing a miniature fluorescent microscopic imaging module, including an imaging lens module and an illumination device;
  • the imaging lens module includes an objective lens and a tube lens arranged symmetrically from the object side to the image side, an image sensor with a high spatial sampling rate arranged on the side of the tube mirror, and an image sensor arranged in front of the objective lens, or The emission filter between the objective lens and the tube lens, or between the tube lens and the image sensor;
  • the illumination device is arranged separately from the imaging lens module to form uniform illumination of the sample
  • the excitation light path of the illumination device is set separately from the detection light path of the imaging lens module; the detection light path from the object side to the image side is the sample, the emission filter, the objective lens, the tube lens and the image sensor; or the sample, Objective lens, emission filter, tube lens and image sensor; or sample, objective lens, tube lens, emission filter and image sensor.
  • the objective lens and the tube lens form a lens group, and the distance from the object surface of the lens group to the image surface on the optical axis is TTL, and the The distance from the object side surface of the lens group to the image side surface on the optical axis is TD, which satisfies the following relationship: 0.15 ⁇ TD/TTL ⁇ 0.9;
  • the length of the optical tube of the lens group satisfies the following relationship: 0.2 ⁇ TTL/( f 1 +f 2 ) ⁇ 15;
  • the distance L Obj from the principal plane of the object side of the objective lens to the object surface satisfies the following relationship: 0.5f 1 ⁇ L Obj ⁇ 1.5f 1 ;
  • the distance L Ima from the principal plane of the image side of the tube lens to the image surface satisfies the following relationship: 0.5f 2 ⁇ L Ima ⁇ 1.5f 2 .
  • the illumination device includes an excitation light source arranged on the periphery of the objective lens and a uniform light guide plate corresponding to the excitation light source;
  • the light emitted by the excitation light source passes through the uniform light guide plate to form uniform illumination of the sample.
  • the excitation light source includes a plurality of excitation light sources, which are uniformly distributed on the periphery of the objective lens in a ring shape; And the end surface of the ring-shaped light guide plate is substantially flush with the end surface of the objective lens close to the sample.
  • the illumination device includes an excitation light source arranged on one side of the sample; the light emitted by the excitation light source illuminates the target area from the side of the sample .
  • the micro-fluorescence microscopy imaging module further includes a support arm provided on one side of the imaging lens module; the excitation light source is mounted on the support arm .
  • the illumination device is arranged under the sample, and includes a first excitation light source arranged correspondingly under the periphery of the objective lens, and the first excitation light source.
  • the first uniform light guide plate corresponding to the excitation light source and the shielding baffle set corresponding to the edge of the field of view of the objective lens, the light is irradiated to the target area from below the sample side.
  • the lighting device further includes a second excitation light source correspondingly arranged under the field of view of the objective lens, and a second excitation light source corresponding to the second excitation light source.
  • Uniform light guide plate the light is irradiated to the target area from directly below the sample;
  • the shielding baffle separates the first excitation light source and the second excitation light source, the first uniform light guide plate and the second uniform light guide plate, respectively.
  • the micro-fluorescence microscopy imaging module further includes a support base, the support base includes a support arm, and a base fixed at the end of the support arm; The support arm fixes the sample from its side.
  • the excitation light source is a laser diode or a light emitting diode
  • the objective lens and the tube lens are respectively composed of at least three lenses.
  • the miniature fluorescence microscope imaging module further includes a drive circuit board with a USB interface for driving the illumination device and the image sensor; and or
  • a protective glass set in front of the objective lens is
  • the micro-fluorescence microscopy imaging module further includes a focusing motor, with an animal lens, or a tube lens, or is composed of an objective lens, an emission filter, and a tube lens The overall movement to achieve focusing.
  • a focusing motor with an animal lens, or a tube lens, or is composed of an objective lens, an emission filter, and a tube lens The overall movement to achieve focusing.
  • the invention is based on the structure of a traditional infinity-corrected fluorescence microscope, adopts an approximately symmetrical structure to design an objective lens and a tube lens, and combines a sensor with a high spatial sampling rate to realize high-resolution imaging at a low magnification.
  • This design can effectively shorten the conjugation distance and solve the problem that the image distance is still very long after the microscope is miniaturized.
  • the three excitation lighting schemes eliminate the need to add excitation filters and dichroic mirrors, which can meet the lighting requirements for different observation samples.
  • the three different positions of the emission filter reduce the aberration and ensure the imaging quality.
  • the driving circuit of the entire microscope can be powered by a simple USB connection, and the image of the image sensor can also be connected to a computer or mobile phone through the USB for display and storage.
  • the microscope imaging module can also add a focusing motor. Due to the small size and light weight of the objective lens and tube lens, mature focusing motors such as voice coil motors can be used to drive the lens group to focus.
  • Fig. 1 is a schematic structural diagram of a first embodiment of a miniature fluorescence microscopy imaging module of the present invention
  • Fig. 2 is a schematic structural diagram of a second embodiment of the miniature fluorescence microscopy imaging module of the present invention.
  • Fig. 3 is a schematic structural diagram of a third embodiment of a miniature fluorescence microscopy imaging module of the present invention.
  • Fig. 4 is a schematic structural diagram of a fourth embodiment of the miniature fluorescence microscopy imaging module of the present invention.
  • orientation or positional relationship indicated by “front”, “rear”, “upper”, “lower”, etc. is based on the orientation or positional relationship shown in the drawings, constructed and operated in a specific orientation, and is only for It is convenient to describe the technical solution, rather than indicating that the pointed device or element must have a specific orientation, so it cannot be understood as a limitation of the present utility model.
  • the present invention constructs a miniature fluorescent microscopy imaging module, which includes an imaging lens module and an illumination device;
  • the imaging lens module includes an objective lens 18 and a tube lens 16 arranged symmetrically from the object side to the image side, an image sensor 28 with a high spatial sampling rate arranged on the image side of the tube lens 16, and an image sensor 28 arranged in front of the objective lens 18, or The emission filter 17 between the objective lens 18 and the tube lens 16 or between the tube lens 16 and the image sensor 28.
  • the illuminating device and the imaging lens module are separately arranged to form uniform illumination of the sample 10.
  • the excitation light path of the illumination device is set separately from the detection light path of the imaging lens module; the detection light path from the object side to the image side is the sample 10, the emission filter 17, the objective lens 18, the tube lens 16 and the image sensor 28; or the sample 10, The objective lens 18, the emission filter 17, the barrel lens 16 and the image sensor 28; or the sample 10, the objective lens 18, the barrel lens 16, the emission filter 17 and the image sensor 28.
  • the objective lens 18 and the tube lens 16 are respectively composed of at least three lenses.
  • the material of the lens can be glass or plastic, and the surface of the lens can be set to a spherical or aspherical surface according to aberration correction requirements.
  • the objective lens 18 and the tube lens 16 form a lens group.
  • the lens group is an infinity-corrected microscope and adopts an approximately symmetrical design to facilitate optimization of aberrations. Under low magnification, a resolution of 1 ⁇ m can be achieved. Low magnification can effectively reduce the length of the image distance, realize the miniaturization of the microscope, and can increase the field of view.
  • the distance from the object surface of the lens group to the image surface on the optical axis is TTL
  • the distance from the object side surface of the lens group to the image side surface on the optical axis is TD, which satisfies the following relationship: 0.15 ⁇ TD/TTL ⁇ 0.9 ;
  • the optical tube length of the lens group satisfies the following relationship: 0.2 ⁇ TTL/( f 1 +f 2 ) ⁇ 15;
  • the distance L Obj from the principal plane of the object side of the objective lens 18 to the object surface satisfies the following relationship: 0.5f 1 ⁇ L Obj ⁇ 1.5f 1 ;
  • the distance L Ima from the principal plane of the image side of the tube lens 16 to the image surface satisfies the following relationship: 0.5f 2 ⁇ L Ima ⁇ 1.5f 2 .
  • the emission filter 17 may be an absorption filter or an interference filter.
  • the distance between the objective lens 18 and the tube lens 16 can be shortened, which helps light
  • the pupil is connected to better correct aberrations; when it is set between the objective lens 18 and the tube lens 16, because the light incident on the filter is parallel light, aberrations can be avoided; it is set on the tube lens 16 and the image sensor When it is between 28, the light incident angle is small when working in afocal mode, which is beneficial to better realize the function of filtering light.
  • the micro fluorescence microscopy imaging module further includes a driving circuit board 11 with a USB interface for driving the lighting device, the image sensor 28 and the focusing motor 27. Its USB interface can be connected to computers, mobile phones and other devices to achieve power supply and display images.
  • the lighting device includes an excitation light source 19 and a uniform light guide plate 29 corresponding to the excitation light source 19.
  • the excitation light source 19 can be a laser diode or a light emitting diode
  • the material of the homogenizing light guide plate 29 can be PC (polycarbonate), PMMA (acrylic), glass and other materials, and the homogenization effect is achieved by processing the surface of the material into a frosted surface.
  • the illumination method of the illuminating device is as follows: 1) Annular light reflective dark field illumination above the sample, the annular light is distributed on the periphery of the objective lens and is flush with the lower surface of the objective lens;
  • the side of the sample is illuminated, and the light from the 0° direction of the light source is directed to the center of the sample field of view within the range of an angle of 10° with the horizontal plane.
  • the light from the 0° light emitting direction of the light source is incident on the center of the sample field of view along the horizontal direction;
  • the ring light is distributed under the sample and outside the field of view.
  • the above three illumination methods can achieve uniform illumination of the sample surface while avoiding the excitation light directly entering the lens to form stray light, thereby affecting the imaging effect.
  • the three lighting methods eliminate the configuration of dichroic mirrors and excitation filters, which can effectively reduce the space and increase the compactness of the structure.
  • a protective glass 15 is provided in front of the objective lens 18, which can effectively protect the entire imaging module from external dust, moisture and other pollution.
  • the structure is miniature, compact, simple and easy to install and adjust. Because the microscope adopts an approximately symmetrical structure, combined with a high spatial sampling rate image sensor, a large field of view and high resolution imaging can be realized at low magnification, which solves the problem of long image distance, which is beneficial to shorten the conjugate distance and realize imaging Miniaturization of modules. In addition, the aforementioned excitation filter and dichroic mirror do not need to be provided, which simplifies the structure and further reduces the space occupied, thereby realizing a more compact design.
  • the fluorescence microscopy imaging module only needs to be powered by USB to work, saving the trouble of connecting to the mains.
  • it can be directly connected to a mobile phone to display and store images, which is convenient to carry and use outdoors.
  • the micro-fluorescence microscopy imaging module may further include a focusing motor 27, and optionally the focusing motor 27 may be a voice coil motor, a stepping motor, an ultrasonic motor, a memory alloy motor, and the like.
  • the focusing motor 27 can carry the objective lens 18 to achieve focusing without affecting the magnification of the system. If the influence on the magnification of the system is not considered, the focusing motor 27 can also drive the barrel lens 16 or the overall movement composed of the objective lens 18, the emission filter 17, and the barrel lens 16 to achieve focusing.
  • the focusing motor 27 When the focusing motor 27 is equipped with the animal lens 18 or tube lens 16 to realize focusing, it changes the position of the object plane of the miniature fluorescence microscopy imaging module.
  • the object plane closest to the micro-fluorescence microscopy imaging module is the near-focus object plane
  • the object plane farthest from the micro-fluorescence microscopy imaging module is the far-focus object plane
  • the direction away from the imaging lens module is the positive direction.
  • the objective lens 18 and the tube lens 16 form a lens group, which includes a limit surface on the object side of the lens group.
  • the object side surface of the protective glass 15 is the limit surface;
  • the end surface of the shell of the imaging lens module or other mechanical structure matched with the imaging lens module is the limit surface.
  • the near-focus object surface is within ⁇ 50 ⁇ m of the limit surface, and the distance between the far-focus object surface and the limit surface is ⁇ 220 ⁇ m.
  • the stroke of the focusing motor 27 is ⁇ 300 ⁇ m and ⁇ 600 ⁇ m, and when the focusing motor 27 moves with the objective lens 18, the focusing distance is the same as the stroke of the focusing motor 27.
  • the minimum distance between the objective lens 18 and the tube lens 16 is ⁇ 50 ⁇ m.
  • the minimum distance between the objective lens 18 and the protective glass 15 is ⁇ 50 ⁇ m.
  • the near-focus object surface can cover the short-distance area of the protective glass 15 and can image close objects. At the same time, the far-focus object surface can surpass the cover glass commonly used in microscopes to meet the needs of biomedical imaging;
  • the stroke of the focusing motor 27 can effectively cover the dimensional tolerances of the components in the module due to processing and installation, which improves the manufacturability of mass production;
  • the near-focus object surface at this time is the object surface that the lens group focuses on when the distance between the objective lens 18 and the tube lens 16 is the smallest;
  • the near-focus object plane at this time is the object plane on which the lens group focuses when the distance between the objective lens 18 and the tube lens 16 is the smallest.
  • the detection light path from the object side to the image side is the sample 10, the protective glass 15, the objective lens 18, the tube lens 16, the emission filter 17, and the image sensor 28 in order.
  • the objective lens 18 is fixed in the voice coil motor 27, and the tube lens 16 is fixed in the tube lens fixing member 12.
  • the distance TTL from the object surface to the image surface of the lens group on the optical axis is 10.2 mm
  • the distance TD from the object side surface to the image side surface on the optical axis is 7.2 mm.
  • the emission filter 17 is fixed in the housing 26, and the image sensor 28 is directly integrated on the driving circuit board 11.
  • the lighting device includes an excitation light source 19 arranged on the periphery of the objective lens 18 and a uniform light guide plate 29 corresponding to the excitation light source 19.
  • the light emitted by the excitation light source 19 passes through the uniform light guide plate 29 to form uniform illumination for the sample 10.
  • the excitation light source 19 includes a plurality of excitation light sources 19, which are uniformly distributed around the periphery of the objective lens 18 in a ring shape.
  • the end surface of 18 close to the sample 10 is basically flush, forming a ring-shaped light reflection dark field illumination above the sample.
  • the excitation light source 19 is an LED lamp, and the LED lamp and the uniform light guide plate 29 are fixed on the housing 26.
  • the LED lamp is at least two LED light sources distributed at equal angular intervals along the circumferential direction. The specific number of light sources can be illuminated according to the sample surface. Parameter settings such as uniformity and illuminance value.
  • USB interface on the driving circuit board 11, which can be directly connected to a computer USB port or connected to a mobile phone through an OTG cable.
  • the LED lamp 19 emits excitation light, which irradiates the sample 10 through the uniform light guide plate 29 and the protective glass 15 to form uniform illumination.
  • the fluorescent substance is excited to generate a fluorescent signal, which is collected by the objective lens 18 after passing through the protective glass 15, and then sequentially focused by the tube lens 16 and filtered by the emission filter 17, and finally imaged on the image sensor 28.
  • the signal of the image sensor 28 is processed by the driving circuit board 11, and displayed on a computer or a mobile phone via a USB data line, which allows the operator to view the fluorescent image in real time and save it as needed.
  • the voice coil motor 27 is fixed to the housing 26 around the objective lens 18.
  • the driving circuit board 11 drives the voice coil motor 27, and the energized coil inside the voice coil motor 27 generates ampere force under the action of the magnetic field, so that the objective lens 18 can move axially, realizing the automatic focusing function.
  • the lighting device in the second embodiment, as shown in FIG. 2, compared with the first embodiment, the lighting device, the position of the emission filter 17 and the setting of the focusing object are different.
  • the illuminating device includes an excitation light source 19 arranged on the side of the sample 10, and light emitted by the excitation light source 19 is irradiated from the side of the sample 10 to the target area.
  • the miniature fluorescence microscopy imaging module also includes a support arm 24 arranged on one side of the imaging lens module, and the excitation light source 19 is mounted on the support arm 24.
  • the micro-fluorescence microscopy imaging module further includes a support base, and the support base includes a support arm 24 and a base 23 fixed at the end of the support arm 24.
  • the excitation light source 19 is a light-emitting diode, which is fixed on the support arm 24, and the light emitted from it illuminates the target area from the side of the sample.
  • the structure is simple and can achieve the effect of uniform illumination.
  • the number of excitation light sources 19 can be distributed along the periphery of the sample according to actual lighting requirements.
  • Support arms 24 are connected to the drive circuit board 11 to provide circuit support for the excitation light sources 19.
  • the base 23 provides support for the entire fluorescence microscopy imaging module.
  • the emission filter 17 is arranged between the objective lens 18 and the tube lens 16 and fixed on the housing 26.
  • the emission filter 17 filters out other unwanted wavelengths of light passing through the objective lens 18 to avoid aberrations.
  • the voice coil motor 27 is fixed on the housing 26 around the tube lens 16 to drive the tube lens 16 to move axially to realize automatic focusing.
  • the lighting device, the position of the emission filter 17, and the setting of the focusing object are different.
  • the illuminating device is arranged under the sample 10, and includes at least two first excitation light sources 19 corresponding to the periphery of the field of view of the objective lens 18, a first uniform light guide plate 29 corresponding to the first excitation light source 19, and a field of view corresponding to the objective lens 18
  • the shielding plate 30 is set at the edge, and light is irradiated to the target area from the bottom of the sample 10 side.
  • the lighting device also includes at least one second excitation light source 19A correspondingly arranged under the field of view of the objective lens 18 and a second uniform light guide plate 29A corresponding to the second excitation light source 19A.
  • the shielding plate 30 separates the first excitation light source 19 and the second excitation light source 19A, the first uniform light guide plate 29 and the second uniform light guide plate 29A, respectively.
  • the excitation light source includes a plurality of excitation light sources, which are uniformly distributed in the outer periphery of the field of view of the objective lens 18 in a ring shape.
  • the uniform light guide plate is an annular light guide plate arranged in the light emission direction of the excitation light source.
  • the micro-fluorescence microscopy imaging module further includes a support base, and the support base includes a support arm 24 and a base 23 fixed at the end of the support arm 24.
  • the support arm 24 fixes the sample 10 from its side.
  • the first excitation light source 19 and the second excitation light source 19A are both LED lamps.
  • the LED lamps are fixed on the base 23 and located at the periphery of the field of view of the objective lens.
  • the lower part of the sample side illuminates the target area to form a transmitted dark field illumination.
  • This illumination method is suitable for uniform illumination of transparent samples, and can also prevent the illumination light from directly entering the objective lens.
  • the function of the shielding plate 30 is to prevent the light of the LED lamp 19 from being directly incident on the sample surface from the inside, and at the same time to isolate the first uniform light guide plate 29 and the second uniform light guide plate 29A.
  • directly below the sample there are also LED lights 19A, which can be independently controlled with the LED lights 19 respectively.
  • the support arm 24 is connected to the driving circuit board 11 and provides circuit support for the ring-shaped light source, that is, the first excitation light source 19 and the second excitation light source 19A.
  • the emission filter 17 is arranged in front of the objective lens 18, which can shorten the distance between the objective lens 18 and the tube lens 16, facilitate pupil connection, and better correct aberrations.
  • the position where the voice coil motor 27 is set is different.
  • the voice coil motor 27 is located on the periphery of the fixed parts of the objective lens 18, the tube lens 16, and the emission filter 17, which can drive the lens group to move as a whole to realize automatic focusing.
  • the voice coil motor 27 is reduced, and the objective lens 18 is fixed on the objective lens fixing member 13, so that the structure of the entire microscope is simpler and miniaturized.
  • the protective glass 15 is reduced to reduce the stray light entering the lens.
  • the light source 22 is a laser diode, which has a small divergence angle, and uses light to be fully concentrated and incident on the surface of the sample.

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Abstract

本发明公开了微型荧光显微成像模块，包括成像镜头模组以及照明装置；成像镜头模组包括从物方至像方依次对称设置的物镜和筒镜、设于筒镜像侧的具有高空间采样率的图像传感器、以及设于物镜前方、或物镜与筒镜之间、或筒镜与图像传感器之间的发射滤光片；照明装置与成像镜头模组分开设置，形成对样本的均匀照明；照明装置的激发光路与成像镜头模组的探测光路分开设置。本发明解决了显微镜微型化后像距依然很长的问题。

Description

一种微型荧光显微成像模块 技术领域
本发明涉及光学成像领域，尤其涉及一种微型荧光显微成像模块。
 背景技术
在生命科学领域，常需要使用到荧光显微镜。荧光显微镜通常包含光源、激发滤光片、二向色镜、物镜、发射滤光片、筒镜、探测器等器件。光源经过激发滤光片得到激发光，经二向色镜反射进入物镜，聚焦至样本表面。激发出的荧光信号经物镜收集，经过二向色镜、滤光片后由筒镜成像至探测器靶面。这种显微镜通常具有一定的放大倍数，必然造成物距短、像距长，像方空间必须留有足够的空间来满足这种光路设置要求。另外，光源需要接市电，探测器如CCD或者CMOS也需要接市电或者电脑供电，图像的显示和存储还需要连接电脑。再者，显微镜还必须带有调焦机构，无论是手动调焦还是自动调焦，其机械结构都较为复杂，占据不小的空间。正因为以上这些原因，荧光显微镜通常体积较大，结构复杂，造价高，且不具备便携性。
荧光显微镜能够成像的核心部件是物镜和筒镜，制约其不能小型化的主要因素是系统存在较大放大倍率，导致筒镜的焦距通常很长。并且滤光片组依旧沿用之前的设置进行微缩，没有进一步考虑其小型化设计。
 技术问题
本发明要解决的技术问题在于，针对现有技术的缺陷，提供一种微型荧光显微成像模块。
 技术解决方案
本发明解决其技术问题所采用的技术方案是：构造一种微型荧光显微成像模块，包括成像镜头模组以及照明装置；
所述成像镜头模组包括从物方至像方依次对称设置的物镜和筒镜、设于所述筒镜像侧的具有高空间采样率的图像传感器、以及设于所述物镜前方、或所述物镜与所述筒镜之间、或所述筒镜与所述图像传感器之间的发射滤光片；
所述照明装置与所述成像镜头模组分开设置，形成对样本的均匀照明；
所述照明装置的激发光路与所述成像镜头模组的探测光路分开设置；所述探测光路从物方到像方依次为样本、发射滤光片、物镜、筒镜和图像传感器；或样本、物镜、发射滤光片、筒镜和图像传感器；或样本、物镜、筒镜、发射滤光片和图像传感器。
优选地，在本发明所述的微型荧光显微成像模块中，所述物镜和所述筒镜组成镜头组，所述镜头组的物面至像面于光轴上的距离为TTL，所述镜头组的物侧表面至像侧表面于光轴上的距离为TD，其满足如下关系：0.15≤TD/TTL≤0.9；
所述物镜的焦距f 1和所述筒镜的焦距f 2之间满足如下关系：0.1≤f 1/f 2≤10；
所述镜头组的光学筒长满足如下关系：0.2≤TTL/( f 1+f 2)≤15；
所述物镜的物方主平面至所述物面的距离L Obj满足如下关系：0.5f 1≤L Obj≤1.5f 1；
所述筒镜的像方主平面到所述像面的距离L Ima满足如下关系式：0.5f 2≤L Ima≤1.5f 2。
优选地，在本发明所述的微型荧光显微成像模块中，所述照明装置包括设置在所述物镜外围的激发光源、以及与所述激发光源对应的匀光导光板；
所述激发光源发出的光线经所述匀光导光板后形成对所述样本的均匀照明。
优选地，在本发明所述的微型荧光显微成像模块中，所述激发光源包括多个，并呈环形均匀分布在所述物镜外围；所述匀光导光板为设置在所述激发光源的出光方向上的环形导光板；并且所述环形导光板的端面与所述物镜靠近所述样本的端面基本齐平。
优选地，在本发明所述的微型荧光显微成像模块中，所述照明装置包括设置在所述样本一侧的激发光源；所述激发光源发出的光线从所述样本侧方照射到目标区域。
优选地，在本发明所述的微型荧光显微成像模块中，微型荧光显微成像模块还包括设置在所述成像镜头模组一侧的支撑臂；所述激发光源安装在所述支撑臂上。
优选地，在本发明所述的微型荧光显微成像模块中，所述照明装置设置于所述样本下方，包括对应设置于所述物镜视场外围下的第一激发光源、与所述第一激发光源对应的第一匀光导光板、以及对应于所述物镜视场边缘设置的遮挡板，光线从所述样本侧下方照射到目标区域。
优选地，在本发明所述的微型荧光显微成像模块中，所述照明装置还包括对应设置于所述物镜视场下的第二激发光源、以及与所述第二激发光源对应的第二匀光导光板，光线从所述样本正下方照射到目标区域；
所述遮挡板分别将所述第一激发光源和所述第二激发光源、所述第一匀光导光板和所述第二匀光导光板隔离。
优选地，在本发明所述的微型荧光显微成像模块中，所述微型荧光显微成像模块还包括支撑座，所述支撑座包括支撑臂、以及固定在所述支撑臂末端的底座；所述支撑臂从所述样本侧方对其进行固定。
优选地，在本发明所述的微型荧光显微成像模块中，所述激发光源为激光二极管或发光二极管；
所述物镜和所述筒镜分别由至少三片透镜组成。
优选地，在本发明所述的微型荧光显微成像模块中，所述微型荧光显微成像模块还包括带有USB接口的驱动电路板，用于驱动所述照明装置和所述图像传感器；和或
设于所述物镜前方的保护玻璃。
优选地，在本发明所述的微型荧光显微成像模块中，所述微型荧光显微成像模块还包括调焦马达，带动物镜、或筒镜、或由物镜、发射滤光片和筒镜组成的整体移动以实现调焦。
 有益效果
通过实施本发明，具有以下有益效果：
本发明基于传统的无限远校正荧光显微镜的结构，采用近似对称结构设计物镜与筒镜，结合高空间采样率的传感器，在低放大倍率下实现高分辨率成像。这样的设计能够有效缩短共轭距，解决了将显微镜微型化后像距依然很长的问题。三种激发照明方案省去了添加激发滤光片和二向色镜的设置，可满足对不同观察样本的照明需求。发射滤光片设置的三个不同位置，减小了像差，保证了成像的质量。另外，出于携带方便，操作简单的考虑，整个显微镜的驱动电路只需简单的USB连接即可供电，图像传感器的图像也可以通过该USB连接至电脑、手机端进行显示和存储。此外，该显微成像模块还可以添加调焦马达，由于物镜和筒镜的体积小，重量轻，使用音圈马达等成熟的调焦马达便可以带动透镜组调焦。
 附图说明
下面将结合附图及实施例对本发明作进一步说明，附图中：
图1是本发明微型荧光显微成像模块第一实施例的结构示意图；
图2是本发明微型荧光显微成像模块第二实施例的结构示意图；
图3是本发明微型荧光显微成像模块第三实施例的结构示意图；
图4是本发明微型荧光显微成像模块第四实施例的结构示意图。
 本发明的实施方式
为了对本发明的技术特征、目的和效果有更加清楚的理解，现对照附图详细说明本发明的具体实施方式。
需要理解的是，“前”、“后”、“上”、“下”等指示的方位或位置关系为基于附图所示的方位或位置关系、以特定的方位构造和操作，仅是为了便于描述本技术方案，而不是指示所指的装置或元件必须具有特定的方位，因此不能理解为对本实用新型的限制。
如图1-4所示，本发明构造了一种微型荧光显微成像模块，包括成像镜头模组以及照明装置；
其中，成像镜头模组包括从物方至像方依次对称设置的物镜18和筒镜16、设于筒镜16像侧的具有高空间采样率的图像传感器28、以及设于物镜18前方、或物镜18与筒镜16之间、或筒镜16与图像传感器28之间的发射滤光片17。照明装置与成像镜头模组分开设置，形成对样本10的均匀照明。照明装置的激发光路与成像镜头模组的探测光路分开设置；探测光路从物方到像方依次为样本10、发射滤光片17、物镜18、筒镜16和图像传感器28；或样本10、物镜18、发射滤光片17、筒镜16和图像传感器28；或样本10、物镜18、筒镜16、发射滤光片17和图像传感器28。
在一些实施例中，物镜18和筒镜16分别由至少三片透镜组成，镜的材料可为玻璃、塑料，透镜的面型依据像差校正需求可设置为球面或者非球面。
物镜18和筒镜16组成镜头组，该镜头组为无限远校正显微镜，采用近似对称设计，便于优化像差，在低放大倍率下，可实现1μm的分辨率。低放大倍率可有效减小像距长度，实现显微镜的微型化，且能够增加视场范围。
其中，镜头组的物面至像面于光轴上的距离为TTL，镜头组的物侧表面至像侧表面于光轴上的距离为TD，其满足如下关系：0.15≤TD/TTL≤0.9；
物镜18的焦距f 1和筒镜16的焦距f 2之间满足如下关系：0.1≤f 1/f 2≤10；
镜头组的光学筒长满足如下关系：0.2≤TTL/( f 1+f 2)≤15；
物镜18的物方主平面至物面的距离L Obj满足如下关系：0.5f 1≤L Obj≤1.5f 1；
筒镜16的像方主平面到像面的距离L Ima满足如下关系式：0.5f 2≤L Ima≤1.5f 2。
在一些实施例中，发射滤光片17可以是吸收型滤光片或干涉型滤光片，其设于物镜18前方时，可缩短物镜18与筒镜16之间的间距，有助于光瞳衔接，更好地校正像差；其设于物镜18与筒镜16之间时，由于入射到滤光片的光线为平行光，可避免产生像差；其设于筒镜16与图像传感器28之间时，在工作在无焦模式的情况下时光线入射角度小，有利于更好地实现滤除光线的功能。
在一些实施例中，微型荧光显微成像模块还包括带有USB接口的驱动电路板11，用于驱动照明装置、图像传感器28和调焦马达27。其USB接口，可连接电脑、手机等设备实现供电并显示图像。
在一些实施例中，照明装置包括激发光源19、以及与激发光源19对应的匀光导光板29。激发光源19可以是激光二极管和发光二极管，匀光导光板29的材料可以是PC（聚碳酸酯）、PMMA（亚克力）、玻璃等材料，通过在材料表面处理成磨砂面来实现匀光作用。
在一些实施例中，照明装置的照明方式有以下几种：1）样本上方环形光反射式暗场照明，环形光分布在物镜外围，与物镜下表面平齐；
2）样本侧方照明，光源0°出光方向的光线在与水平面呈10°夹角的范围内射向样本视场中心。优选地，光源0°出光方向的光线沿水平方向入射至样本视场中心；
3）样本下方环形透射式暗场照明，环形光分布于样本下方，视场外围。上述三种照明方式在实现样本表面均匀照明的同时，避免激发光直接进入镜头形成杂散光，进而影响成像效果。
此外，三种照明方式省去二向色镜和激发滤光片的配置，可有效减小空间，增加结构的紧凑性。
在一些实施例中，物镜18的前方设有保护玻璃15，可有效保护整个成像模块不受外部灰尘、水汽等污染。
通过实施本发明，具有以下有益效果：
1）无需额外添加激发滤光和二向色镜。荧光物质的吸收谱通常是一个光谱带，而激光二极管和发光二极管的光谱都很窄，所以根据荧光物质的选择对应波长的光源即可，不需要添加激发滤光片。又因为激发光路与探测光路分开设置，不需要使用二向色镜将激发光与荧光做分离。这样的设置既可有效减小荧光显微镜的整体体积，又使得装置更加简单。
2）结构微型、紧凑、简单，便于装调。由于显微镜采用近似对称结构，结合高空间采样率的图像传感器，在低放大倍率下即可实现大视场高分辨率成像，解决了像距较长的问题，有利于缩短共轭距，实现成像模块微型化。加上前述的无需设置激发滤光片和二向色镜，简化结构的同时进一步减小所占空间，实现更紧凑的设计。
3）操作简单，方便携带。该荧光显微成像模块只需通过USB供电即可工作，省去连接市电的麻烦。另外，可直接连接手机显示并存储图像，方便携带至室外使用。
在一些实施例中，微型荧光显微成像模块还可包括调焦马达27，可选地调焦马达27可以是音圈马达、步进电机、超声马达、记忆合金马达等。调焦马达27可以带动物镜18实现调焦的同时，不影响系统放大倍率。如不考虑对系统放大倍率的影响，调焦马达27也可以带动筒镜16或者由物镜18、发射滤光片17、筒镜16组成的整体移动以实现调焦。
调焦马达27带动物镜18或筒镜16实现调焦时，即改变本微型荧光显微成像模块的物面位置。其中，距离微型荧光显微成像模块最近的物面为近焦物面，距离微型荧光显微成像模块最远的物面为远焦物面，并以远离成像镜头模组的方向为正方向。物镜18和筒镜16组成镜头组，在镜头组的物侧包含一个限位面，当镜头组的物侧有保护玻璃15时，保护玻璃15的物方表面为限位面；当镜头组的物侧没有保护玻璃15时，由成像镜头模组的外壳或与成像镜头模组配合的其他机械结构的端面为限位面。近焦物面位于限位面±50 μm范围内，远焦物面与限位面的距离≥220 μm。
调焦马达27的行程≥300 μm且≤600 μm，并且当调焦马达27带动物镜18移动时，调焦距离与调焦马达27的行程相同。在调焦范围内，物镜18和筒镜16的最小距离≥50 μm。当有保护玻璃15时，在调焦范围内，物镜18和保护玻璃15的最小距离≥50 μm。
采用上述参数具有以下有益效果：
1）近焦物面能够覆盖保护玻璃15的近距离区域，可对贴近的物体进行成像，同时远焦物面能够超越显微镜常用的盖玻片，满足于生物医学成像的需求；
2）调焦马达27行程能够有效覆盖模组内各组件由于加工和安装导致的尺寸公差，提升了量产的可制造性；
3）物镜18与保护玻璃15和筒镜16之间均保留了最小间隙，提升了本成像镜头模组的可靠性，避免由于调焦马达27超过额定行程时模组内部件相互撞击从而损坏；
4）在实现上述有益效果的同时，实现了模组的小型化。
另外，当调焦马达27带动物镜18移动时，筒镜16位置固定。此时的近焦物面为物镜18与筒镜16距离最小时，镜头组所对焦的物面；
调焦马达27带动筒镜16移动时，物镜18位置固定。此时的近焦物面为物镜18与筒镜16距离最小时，镜头组所对焦的物面。
下面结合附图和实施例，对本发明的具体实施方式进一步详细描述，以下实施例用于说明本发明，但不用来限制本发明/实用新型的范围。
在第一实施例中，如图1所示，探测光路从物方到像方依次是样本10、保护玻璃15、物镜18、筒镜16、发射滤光片17、图像传感器28。物镜18固定于音圈马达27中，筒镜16固定于筒镜固定件12中。其中，物镜18的焦距f 1=2.6 mm，筒镜16的焦距f 2=5.1 mm。镜头组的物面至像面于光轴上的距离TTL为10.2 mm，其物侧表面至像侧表面于光轴上的距离TD为7.2 mm。发射滤光片17固定于外壳26中，图像传感器28直接集成在驱动电路板11上。
照明装置包括设置在物镜18外围的激发光源19、以及与激发光源19对应的匀光导光板29。激发光源19发出的光线经匀光导光板29后形成对样本10的均匀照明。在本实施例中，激发光源19包括多个，并呈环形均匀分布在物镜18外围，匀光导光板29为设置在激发光源19的出光方向上的环形导光板，并且环形导光板的端面与物镜18靠近样本10的端面基本齐平，形成样本上方环形光反射式暗场照明。优选地，激发光源19为LED灯，LED灯和匀光导光板29固定在外壳26上，LED灯为沿圆周方向等角度间隔分布的至少2个LED光源，具体的光源个数可根据样本表面照明均匀度、照度值等参数设置。
驱动电路板11上有USB接口，可直接连接电脑USB口，或者通过OTG线与手机相连。当电路板上电后，荧光显微镜便处于工作状态。LED灯19发出激发光，经匀光导光板29和保护玻璃15后照射至样本10上，形成均匀照明。荧光物质受激发产生荧光信号，经过保护玻璃15后由物镜18收集，再依次经过筒镜16聚焦、发射滤光片17过滤后，最终成像在图像传感器28上。图像传感器28的信号经过驱动电路板11处理，通过USB数据线显示在电脑上或者手机上，可供操作者实时观看荧光图像，并根据需要进行保存。
音圈马达27在物镜18的周围，固定于外壳26上。驱动电路板11对音圈马达27行驱动，音圈马达27内部通电线圈在磁场作用下产生安培力，使得物镜18能够轴向运动，现实自动调焦功能。
在第二实施例中，如图2所示，与第一实施例相比，照明装置、发射滤光片17的位置和调焦对象的设置有所不同。照明装置包括设置在样本10一侧的激发光源19，激发光源19发出的光线从样本10侧方照射到目标区域。
微型荧光显微成像模块还包括设置在成像镜头模组一侧的支撑臂24，激发光源19安装在支撑臂24上。
在本实施例中，微型荧光显微成像模块还包括支撑座，支撑座包括支撑臂24、以及固定在支撑臂24末端的底座23。激发光源19为发光二极管，固定在支撑臂24上，其发出的光线从样本侧方照射到目标区域，结构简单又能实现均匀照明的效果。激发光源19的数量可按照实际照明需求沿样本四周分布支撑臂24连接驱动电路板11，为激发光源19提供线路支撑。底座23给整个荧光显微成像模块提供支撑。发射滤光片17设置于物镜18和筒镜16之间，固定于外壳26上。发射滤光片17则会过滤掉经过物镜18的其它不需要的波长的光，可避免产生像差。音圈马达27在筒镜16的周围，固定在外壳26上，带动筒镜16轴向运动，实现自动调焦。
在第三实施例中，如图3所示，与第一实施例相比，照明装置、发射滤光片17的位置和调焦对象的设置有所不同。照明装置设置于样本10下方，包括对应设置于物镜18视场外围下的至少两个第一激发光源19、与第一激发光源19对应的第一匀光导光板29、以及对应于物镜18视场边缘设置的遮挡板30，光线从样本10侧下方照射到目标区域。
照明装置还包括对应设置于物镜18视场下的至少一个第二激发光源19A、以及与第二激发光源19A对应的第二匀光导光板29A，光线从样本10正下方照射到目标区域。其中，遮挡板30分别将第一激发光源19和第二激发光源19A、第一匀光导光板29和第二匀光导光板29A隔离。
在本实施例中，激发光源包括多个，并呈环形均匀分布在物镜18视场外围下，匀光导光板为设置在激发光源的出光方向上的环形导光板。
在本实施例中，微型荧光显微成像模块还包括支撑座，支撑座包括支撑臂24、以及固定在支撑臂24末端的底座23。支撑臂24从样本10侧方对其进行固定。
在本实施例中，第一激发光源19和第二激发光源19A均为LED灯，LED灯固定在底座23上，位于物镜视场外围，其发出的光线经过第一匀光导光板29后，从样本侧下方照射到目标区域，形成透射暗场照明。这种照明方式适合透明样品的均匀照明，还可避免照明光直接入射进物镜。遮挡板30的作用是防止LED灯19的光直接从内侧入射至样本表面，同时将第一匀光导光板29和第二匀光导光板29A隔离。此外，在样本正下方，也分布着LED灯19A，其与LED灯19可分别独立控制。当LED灯29A开启时，光线从样本正下方通过第二匀光导光板29A后，直接照射样本实现荧光激发。上述两种光源照明方式可针对不同样本或者不同需求选择。支撑臂24 连接驱动电路板11，为环型光源，即第一激发光源19和第二激发光源19A提供线路支撑。发射滤光片17设置在物镜18前方，可缩短物镜18与筒镜16之间的间距，有助于光瞳衔接，更好地校正像差。音圈马达27设置的位置有所不同。音圈马达27在物镜18、筒镜16、发射滤光片17的固定件外围上，可带动透镜组整体运动，实现自动调焦。
在第四实施例中，如图4所示，第二实施例相比，减少了音圈马达27，物镜18固定在物镜固定件13上，使整个显微镜结构更加简单，微型化。减少了保护玻璃15，使其减少了杂散光进入镜头中。光源22为激光二极管，发散角小，有利用光线充分集中入射至样本表面。
 工业实用性
本发明是通过具体实施例进行说明的，本领域技术人员应当明白，在不脱离本发明范围的情况下，还可以对本发明进行各种变换和等同替代。另外，针对特定情形或具体情况，可以对本发明做各种修改，而不脱离本发明的范围。因此，本发明不局限于所公开的具体实施例，而应当包括落入本发明权利要求范围内的全部实施方式。

Claims (12)

  1. 一种微型荧光显微成像模块，其特征在于，包括成像镜头模组以及照明装置；
    所述成像镜头模组包括从物方至像方依次对称设置的物镜（18）和筒镜（16）、设于所述筒镜（16）像侧的具有高空间采样率的图像传感器（28）、以及设于所述物镜（18）前方、或所述物镜（18）与所述筒镜（16）之间、或所述筒镜（16）与所述图像传感器（28）之间的发射滤光片（17）；
    所述照明装置与所述成像镜头模组分开设置，形成对样本（10）的均匀照明；
    所述照明装置的激发光路与所述成像镜头模组的探测光路分开设置；所述探测光路从物方到像方依次为样本（10）、发射滤光片（17）、物镜（18）、筒镜（16）和图像传感器（28）；或样本（10）、物镜（18）、发射滤光片（17）、筒镜（16）和图像传感器（28）；或样本（10）、物镜（18）、筒镜（16）、发射滤光片（17）和图像传感器（28）。
  2. 根据权利要求1所述的微型荧光显微成像模块，其特征在于，所述物镜（18）和所述筒镜（16）组成镜头组，所述镜头组的物面至像面于光轴上的距离为TTL，所述镜头组的物侧表面至像侧表面于光轴上的距离为TD，其满足如下关系：0.15≤TD/TTL≤0.9；
    所述物镜（18）的焦距f 1和所述筒镜（16）的焦距f 2之间满足如下关系：0.1≤f 1/f 2≤10；
    所述镜头组的光学筒长满足如下关系：0.2≤TTL/( f 1+f 2)≤15；
    所述物镜（18）的物方主平面至所述物面的距离L Obj满足如下关系：0.5f 1≤L Obj≤1.5f 1；
    所述筒镜（16）的像方主平面到所述像面的距离L Ima满足如下关系式：0.5f 2≤L Ima≤1.5f 2。
  3. 根据权利要求1所述的微型荧光显微成像模块，其特征在于，所述照明装置包括设置在所述物镜（18）外围的激发光源（19）、以及与所述激发光源（19）对应的匀光导光板（29）；
    所述激发光源（19）发出的光线经所述匀光导光板（29）后形成对所述样本（10）的均匀照明。
  4. 根据权利要求3所述的微型荧光显微成像模块，其特征在于，所述激发光源（19）包括多个，并呈环形均匀分布在所述物镜（18）外围；所述匀光导光板（29）为设置在所述激发光源（19）的出光方向上的环形导光板；并且所述环形导光板的端面与所述物镜（18）靠近所述样本（10）的端面基本齐平。
  5. 根据权利要求1所述的微型荧光显微成像模块，其特征在于，所述照明装置包括设置在所述样本（10）一侧的激发光源（19）；所述激发光源（19）发出的光线从所述样本（10）侧方照射到目标区域。
  6. 根据权利要求5所述的微型荧光显微成像模块，其特征在于，微型荧光显微成像模块还包括设置在所述成像镜头模组一侧的支撑臂（24）；所述激发光源（19）安装在所述支撑臂（24）上。
  7. 根据权利要求1所述的微型荧光显微成像模块，其特征在于，
    所述照明装置设置于所述样本（10）下方，包括对应设置于所述物镜（18）视场外围下的第一激发光源（19）、与所述第一激发光源（19）对应的第一匀光导光板（29）、以及对应于所述物镜（18）视场边缘设置的遮挡板（30），光线从所述样本（10）侧下方照射到目标区域。
  8. 根据权利要求7所述的微型荧光显微成像模块，其特征在于，所述照明装置还包括对应设置于所述物镜（18）视场下的第二激发光源（19A）、以及与所述第二激发光源（19A）对应的第二匀光导光板（29A），光线从所述样本（10）正下方照射到目标区域；
    所述遮挡板（30）分别将所述第一激发光源（19）和所述第二激发光源（19A）、所述第一匀光导光板（29）和所述第二匀光导光板（29A）隔离。
  9. 根据权利要求8所述的微型荧光显微成像模块，其特征在于，所述微型荧光显微成像模块还包括支撑座，所述支撑座包括支撑臂（24）、以及固定在所述支撑臂（24）末端的底座（23）；所述支撑臂（24）从所述样本（10）侧方对其进行固定。
  10. 根据权利要求3-9任一项所述的微型荧光显微成像模块，其特征在于，所述激发光源（19）为激光二极管或发光二极管；
    所述物镜（18）和所述筒镜（16）分别由至少三片透镜组成。
  11. 根据权利要求1-9任一项所述的微型荧光显微成像模块，其特征在于，所述微型荧光显微成像模块还包括带有USB接口的驱动电路板（11），用于驱动所述照明装置和所述图像传感器（28）；和或
    设于所述物镜（18）前方的保护玻璃（15）。
  12. 根据权利要求1所述的微型荧光显微成像模块，其特征在于，所述微型荧光显微成像模块还包括调焦马达（27），带动物镜（18）、或筒镜（16）、或由物镜（18）、发射滤光片（17）和筒镜（16）组成的整体移动以实现调焦。
     
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