Aberration-free laser spot scanning equipment for optical microscopic imaging

Abstract

The invention provides aberration-free laser spot scanning equipment. The aberration-free laser spot scanning equipment comprises a resonance galvanometer, a first concave mirror, a second concave mirror, an X-axis galvanometer scanning mirror, a third concave mirror, a fourth concave mirror and a Y-axis galvanometer scanning mirror which are sequentially arranged in the light beam propagation direction. Existing laser spot scanning equipment generally uses a lens as an optical relay element between scanning mirrors, which inevitably affects the quality of light beams and causes aberration. According to the aberration-free laser spot scanning equipment provided by the invention, the concave mirror is used for replacing a lens to complete optical relay between the scanning mirrors, so that the spherical aberration of a focusing transmission element can be avoided, and the optical power loss generated by light transmission through a glass element is reduced; the scanning device can be used as a scanning device in an optical microscopic imaging (such as laser confocal and two-photon imaging) system.

Description

Technical Field

[0001] The present application relates to the technical field of multiphoton microscopic imaging, and in particular to an aberration-free laser point scanning device for optical microscopic imaging. Background Art

[0002] Optical microscopy techniques (such as confocal imaging and multiphoton imaging) have high resolution and good signal-to-noise ratio, and have become indispensable research tools in life science and medical research. The scanning device composed of a galvanometer scanning mirror and a resonant galvanometer is an indispensable core component in this type of imaging system. Due to the need to expand the excitation beam and the large scanning angle range of the scanning mirror, an optical relay needs to be added between the scanning mirrors. Existing optical relays mostly use lenses as core components. However, due to the existence of spherical aberration / chromatic aberration, lens-based optical relays will affect the quality of the excitation beam and the point spread function. In addition, the projection element will inevitably cause light loss, thereby reducing the excitation light power. Summary of the invention

[0003] In view of this, it is necessary to provide an aberration-free laser point scanning device for optical microscopic imaging that reduces light loss and improves system efficiency in view of the technical defects of the prior art.

[0004] To solve the above problems, this application adopts the following technical solutions:

[0005] The present application provides an aberration-free laser point scanning device for optical microscopic imaging, comprising a laser control module (100) and a resonant galvanometer (1), a first concave mirror (2), a second concave mirror (3), an X-axis galvanometer scanning mirror (4), a third concave mirror (5), a fourth concave mirror (6), a Y-axis galvanometer scanning mirror (7), a reflector (8), a first lens (9), a second lens (10), a dichroic mirror (11), an objective lens (12) and a detection module (13) arranged in sequence along the propagation direction of a light beam emitted by the laser control module (100). When the resonant galvanometer (1) is in a stationary state, the X-axis galvanometer scanning mirror (4) and the Y-axis galvanometer scanning mirror (7) are used to perform full-width scanning within the entire field of view; when the X-axis galvanometer scanning mirror (4) completes deflection at a specified angle and is in a stationary state, the resonant galvanometer (1) and the Y-axis galvanometer scanning mirror (7) are combined to perform rapid scanning within a partial field of view, wherein:

[0006] When the X-axis galvanometer scanning mirror (4) and the Y-axis galvanometer scanning mirror (7) are used to perform full-width scanning imaging within the entire field of view, the resonant galvanometer (1) is in a stationary state: the excitation light beam emitted by the laser control module (100) is incident on the surface of the resonant galvanometer (1), and is reflected by the first concave mirror (2) and the second concave mirror (3) in sequence, and then focused on the X-axis galvanometer scanning mirror (4); the excitation light beam reflected by the X-axis galvanometer scanning mirror (4) is reflected by the third concave mirror ( 5) and the fourth concave mirror (6), and then focused on the Y-axis galvanometer scanning mirror (7); the excitation light beam is reflected by the Y-axis galvanometer scanning mirror (7), and then sequentially passes through the reflector (8), the first lens (9), the second lens (10), the dichroic mirror (11) and the objective lens (12) to be focused in the tissue to be tested; the emission light generated by the excitation is collected by the objective lens (12), reflected by the dichroic mirror (11), and finally detected by the detection module (13), thereby completing the scanning imaging task;

[0007] When the resonant galvanometer (1) and the Y-axis galvanometer scanning mirror (7) are used to perform rapid scanning imaging within a partial field of view, the X-axis galvanometer scanning mirror (4) is in a stationary state: the excitation light beam emitted by the laser control module (100) is incident on the surface of the resonant galvanometer (1), and is reflected by the first concave mirror (2) and the second concave mirror (3) in sequence, and then focused on the X-axis galvanometer scanning mirror (4); the excitation light beam reflected by the X-axis galvanometer scanning mirror (4) is reflected by the third concave mirror (3) in sequence, and then focused on the X-axis galvanometer scanning mirror (4); (5) and the fourth concave mirror (6), and then focused on the Y-axis galvanometer scanning mirror (7); the excitation light beam is reflected by the Y-axis galvanometer scanning mirror (7), and then sequentially passes through the reflector (8), the first lens (9), the second lens (10), the dichroic mirror (11) and the objective lens (12) to be focused in the tissue to be tested; the emission light generated by the excitation is collected by the objective lens (12), reflected by the dichroic mirror (11), and finally detected by the detection module (13), thereby completing the scanning imaging task.

[0008] In some embodiments, the first concave mirror is identical to the second concave mirror, and the two are symmetrically arranged to perform optical relay between the resonant galvanometer and the X-axis scanning mirror.

[0009] In some of the embodiments, the third concave mirror is identical to the fourth concave mirror, and the two are symmetrically arranged to perform optical relay between the X-axis scanning mirror and the Y-axis scanning mirror.

[0010] This application adopts the above technical solution, and its beneficial effects are as follows:

[0011] The aberration-free laser point scanning device provided in the present application avoids the spherical aberration / chromatic aberration caused by the lens and reduces the light loss to almost zero. This technology can achieve aberration-free laser point scanning without increasing the manufacturing cost. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings required for use in the embodiments of the present application or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present application. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0013] Figure 1 A schematic diagram of the structure of an aberration-free laser point scanning device for optical microscopic imaging provided in this application. DETAILED DESCRIPTION

[0014] Embodiments of the present application are described in detail below, and examples of the embodiments are shown in the accompanying drawings, wherein the same or similar reference numerals throughout represent the same or similar elements or elements having the same or similar functions. The embodiments described below with reference to the accompanying drawings are exemplary and are intended to be used to explain the present application, and should not be construed as limiting the present application.

[0015] In the description of the present application, it should be understood that the terms "upper", "lower", "horizontal", "inside", "outside", etc., indicating orientations or positional relationships, are based on the orientations or positional relationships shown in the accompanying drawings, and are only for the convenience of describing the present application and simplifying the description, rather than indicating or implying that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation, and therefore should not be understood as a limitation on the present application.

[0016] In addition, the terms "first" and "second" are used for descriptive purposes only and should not be understood as indicating or implying relative importance or implicitly indicating the number of the indicated technical features. Thus, a feature defined as "first" or "second" may explicitly or implicitly include one or more of the features. In the description of this application, the meaning of "plurality" is two or more, unless otherwise clearly and specifically defined.

[0017] In order to make the objectives, technical solutions and advantages of the present application more clearly understood, the present application is further described in detail below in conjunction with the accompanying drawings and embodiments.

[0018] See also Figure 1The present embodiment provides an aberration-free laser point scanning device for optical microscopic imaging, comprising a laser control module (100) and a resonant galvanometer (1), a first concave mirror (2), a second concave mirror (3), an X-axis galvanometer scanning mirror (4), a third concave mirror (5), a fourth concave mirror (6), a Y-axis galvanometer scanning mirror (7), a reflector (8), a first lens (9), a second lens (10), a dichroic mirror (11), an objective lens (12) and a detection module (13) arranged in sequence along the propagation direction of a light beam emitted by the laser control module (100), wherein when the resonant galvanometer (1) is in a stationary state, the X-axis galvanometer scanning mirror (4) and the Y-axis galvanometer scanning mirror (7) are used to perform full-width scanning within the entire field of view; when the X-axis galvanometer scanning mirror (4) completes the deflection of a specified angle and is in a stationary state, the resonant galvanometer (1) and the Y-axis galvanometer scanning mirror (7) are combined to perform rapid scanning within a partial field of view, wherein:

[0019] When the X-axis galvanometer scanning mirror (4) and the Y-axis galvanometer scanning mirror (7) are used to perform full-width scanning imaging within the entire field of view, the resonant galvanometer (1) is in a stationary state: the excitation light beam emitted by the laser control module (100) is incident on the surface of the resonant galvanometer (1), and is reflected by the first concave mirror (2) and the second concave mirror (3) in sequence, and then focused on the X-axis galvanometer scanning mirror (4); the excitation light beam reflected by the X-axis galvanometer scanning mirror (4) is reflected by the third concave mirror ( 5) and the fourth concave mirror (6), and then focused on the Y-axis galvanometer scanning mirror (7); the excitation light beam is reflected by the Y-axis galvanometer scanning mirror (7), and then sequentially passes through the reflector (8), the first lens (9), the second lens (10), the dichroic mirror (11) and the objective lens (12) to be focused in the tissue to be tested; the emission light generated by the excitation is collected by the objective lens (12), reflected by the dichroic mirror (11), and finally detected by the detection module (13), thereby completing the scanning imaging task.

[0020] When the resonant galvanometer (1) and the Y-axis galvanometer scanning mirror (7) are used to perform rapid scanning imaging within a partial field of view, the X-axis galvanometer scanning mirror (4) is in a stationary state: the excitation light beam emitted by the laser control module (100) is incident on the surface of the resonant galvanometer (1), and is reflected by the first concave mirror (2) and the second concave mirror (3) in sequence, and then focused on the X-axis galvanometer scanning mirror (4); the excitation light beam reflected by the X-axis galvanometer scanning mirror (4) is reflected by the third concave mirror (3) in sequence, and then focused on the X-axis galvanometer scanning mirror (4); (5) and the fourth concave mirror (6), and then focused on the Y-axis galvanometer scanning mirror (7); the excitation light beam is reflected by the Y-axis galvanometer scanning mirror (7), and then sequentially passes through the reflector (8), the first lens (9), the second lens (10), the dichroic mirror (11) and the objective lens (12) to be focused in the tissue to be tested; the emission light generated by the excitation is collected by the objective lens (12), reflected by the dichroic mirror (11), and finally detected by the detection module (13), thereby completing the scanning imaging task.

[0021] This application takes two-photon neurocalcitonin imaging of living mouse brain as an example, and the working method is as follows:

[0022] GCaMP is used to mark the calcium activity of nerve cells. When the X / Y axis scanning mirror is used to perform full-width scanning imaging in the entire field of view (the resonant galvanometer is in a stationary state): the 920-nanometer excitation light beam is incident on the surface of the resonant galvanometer, and is reflected by the first concave mirror and the second concave mirror in turn (the first concave mirror and the second concave mirror form an optical relay), and then focused on the X axis scanning mirror; the excitation light beam reflected by the X axis scanning mirror is reflected by the third concave mirror and the fourth concave mirror in turn (the third concave mirror and the fourth concave mirror form an optical relay), and then focused on the Y axis scanning mirror; the excitation light beam is reflected by the Y axis scanning mirror and enters the microscopic imaging module (composed of the first lens, the second lens and the objective lens), and is focused in the tissue to be tested; the fluorescence generated by the excitation is collected by the objective lens, and after being reflected by the dichroic mirror, it is finally detected by the light detection module to complete the imaging task. This imaging task generally needs to be carried out in a continuous cycle to record changes in neural calcium activity.

[0023] However, the imaging speed based on X / Y galvanometer scanning mirror is slow and cannot meet the time resolution requirements for detecting neural calcium activity.

[0024] When the resonant galvanometer and the Y-axis scanning mirror are used to perform rapid scanning imaging within a partial field of view (the X-axis scanning mirror is in a stationary state): the 920-nanometer excitation light beam is incident on the surface of the resonant galvanometer, and is reflected by the first concave mirror and the second concave mirror in sequence (the first concave mirror and the second concave mirror form an optical relay), and then focused on the X-axis scanning mirror; the excitation light beam reflected by the X-axis scanning mirror is reflected by the third concave mirror and the fourth concave mirror in sequence (the third concave mirror and the fourth concave mirror form an optical relay), and then focused on the Y-axis scanning mirror; the excitation light beam is reflected by the Y-axis scanning mirror and enters the microscopic imaging module (composed of the first lens, the second lens and the objective lens), and is focused on the tissue to be tested; the emission light (such as fluorescence) generated by the excitation is collected by the objective lens, reflected by the dichroic mirror, and finally detected by the light detection module to complete the scanning imaging task. This imaging task generally needs to be performed in a continuous cycle to record the changes in neural calcium activity over time.

[0025] The aberration-free laser point scanning device provided in the embodiment of the present application avoids the spherical aberration / chromatic aberration caused by the lens and reduces the light loss to almost zero. This technology can realize aberration-free laser point scanning without increasing the manufacturing cost.

[0026] It can be understood that the technical features of the above-described embodiments can be arbitrarily combined. In order to make the description concise, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0027] The above are only preferred embodiments of the present application, and only specifically describe the technical principles of the present application. These descriptions are only for explaining the principles of the present application and cannot be interpreted as limiting the scope of protection of the present application in any way. Based on the explanation here, any modifications, equivalent substitutions and improvements made within the spirit and principles of the present application, and other specific implementation methods of the present application that can be associated with the technicians in this field without creative work, should be included in the scope of protection of the present application.

Claims

1. An aberration-free laser point scanning device for optical microscopy imaging, characterized in that: The invention comprises a laser control module (100), and a resonant galvanometer (1), a first concave mirror (2), a second concave mirror (3), an X-axis galvanometer scanning mirror (4), a third concave mirror (5), a fourth concave mirror (6), a Y-axis galvanometer scanning mirror (7), a reflector (8), a first lens (9), a second lens (10), a dichroic mirror (11), an objective lens (12), and a detection module (13) which are sequentially arranged along the propagation direction of a light beam emitted by the laser control module (100); when the resonant galvanometer (1) is in a stationary state, the X-axis galvanometer scanning mirror (4) and the Y-axis galvanometer scanning mirror (7) are used to perform full-width scanning within the entire field of view; when the X-axis galvanometer scanning mirror (4) has completed deflection at a specified angle and is in a stationary state, the resonant galvanometer (1) and the Y-axis galvanometer scanning mirror (7) are combined to perform rapid scanning within a partial field of view, wherein: When the X-axis galvanometer scanning mirror (4) and the Y-axis galvanometer scanning mirror (7) are used to perform full-width scanning imaging within the entire field of view, the resonant galvanometer mirror (1) is in a stationary state: the excitation light beam emitted by the laser control module (100) is incident on the surface of the resonant galvanometer mirror (1), and is reflected by the first concave mirror (2) and the second concave mirror (3) in sequence, and then focused on the X-axis galvanometer scanning mirror (4); the excitation light beam reflected by the X-axis galvanometer scanning mirror (4) is reflected by the third concave mirror (5 ) and the fourth concave mirror (6), and then focused on the Y-axis galvanometer scanning mirror (7); the excitation light beam is reflected by the Y-axis galvanometer scanning mirror (7), and then passes through the reflector (8), the first lens (9), the second lens (10), the dichroic mirror (11) and the objective lens (12) in sequence and is focused on the tissue to be tested; the emission light generated by the excitation is collected by the objective lens (12), and after being reflected by the dichroic mirror (11), it is finally detected by the detection module (13), thereby completing the scanning imaging task; When the resonant galvanometer (1) and the Y-axis galvanometer scanning mirror (7) are used to perform rapid scanning imaging within a partial field of view, the X-axis galvanometer scanning mirror (4) is in a stationary state: the excitation light beam emitted by the laser control module (100) is incident on the surface of the resonant galvanometer (1), and is reflected by the first concave mirror (2) and the second concave mirror (3) in sequence, and then focused on the X-axis galvanometer scanning mirror (4); the excitation light beam reflected by the X-axis galvanometer scanning mirror (4) is reflected by the third concave mirror (3) in sequence, and then focused on the X-axis galvanometer scanning mirror (4); (5) and the fourth concave mirror (6), and then focused on the Y-axis galvanometer scanning mirror (7); the excitation light beam is reflected by the Y-axis galvanometer scanning mirror (7), and then sequentially passes through the reflector (8), the first lens (9), the second lens (10), the dichroic mirror (11) and the objective lens (12) to be focused in the tissue to be tested; the emission light generated by the excitation is collected by the objective lens (12), reflected by the dichroic mirror (11), and finally detected by the detection module (13), thereby completing the scanning imaging task.

2. The aberration-free laser point scanning device for optical microscopic imaging according to claim 1, characterized in that: The first concave mirror is identical to the second concave mirror, and the two are symmetrically arranged for optical relay between the resonant galvanometer mirror and the X-axis scanning mirror.

3. The aberration-free laser point scanning device for optical microscopic imaging according to claim 1, characterized in that: The third concave mirror is identical to the fourth concave mirror, and the two are symmetrically arranged for optical relay between the X-axis scanning mirror and the Y-axis scanning mirror.

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