CN111474141A - A kind of interference microscopy imaging method and interference microscope - Google Patents

Abstract

The invention provides an interference microscopic imaging method and an interference microscope, in particular to an interference microscope, which comprises a light source generating device, a compensation interference cavity, a detection arm and a signal acquisition and processing unit, wherein the light source generating device is used for generating a light source signal; the compensating interference cavity receives and reflects the light beam generated by the light source generating device to form a reflected light beam which enters the detection arm; the detection arm focuses the reflected light beam to form sample light and sends the sample light to the signal acquisition and processing unit; the detection unit is provided with a second microscope objective which is used for focusing light on a sample and modulating the structural information of the sample into light rays to return. According to the interference microscope, the first microscope objective is inserted into the compensation interference cavity, so that the reference light and the sample light pass through the completely same microscope objective only in different sequences, the optical distances of the sample light and the reference light are ensured to be completely equal, and the optical distances of some points on an interference surface caused by inconsistency of the sample light and the reference light passing through an optical device are avoided.

Description

A kind of interference microscopy imaging method and interference microscope

technical field

The invention belongs to an interference microscopic imaging technology, in particular to an interference microscopic imaging method and an interference microscope.

Background technique

Interference microscope is a microscope that uses coherent light interference technology, which can see the structure distribution of transparent sample tissue compared to traditional microscopes. It uses the interference principle to convert the phase difference distribution into an intensity distribution for the naked eye to observe. Using the low coherence of a broad-spectrum light source also enables optical sectioning, which images only thin sections of the sample within a specific optical path range, and the section thickness depends on the temporal coherence length of the light source. Microscopes using low-coherence interference technology have been used in integrated circuit microscopic inspection, clinical pathological diagnosis and other fields, and have the advantages of non-contact and non-destructive inspection.

The current low-coherence interference microscope (patent 201680034161.X) mostly adopts the Michelson interference structure, and the microscope is inserted between the reference arm and the sample arm. This structure is a light splitter structure. When the two arms are far apart, different phase changes will be formed under the environmental vibration, which seriously affects the normal interference pattern acquisition, so that high-quality sample structure diagrams cannot be obtained. In practice, an air flotation platform is often added to filter out environmental interference, so it is bulky and inconvenient to use, which limits its practical use. There is also a common optical path interference system using a Mirau objective lens, but when the numerical aperture is large, the beam splitter in it will cause a large aberration (Stanley Siu-chor Chim, G S Kino. Correlation microscope [J]. Opt. Lett ., 1990, 15(10):579-581.), which brought great difficulties to the structural design of the Mirau objective.

Because the reference light and the sample light are transmitted in the same channel in the interference system using the common optical path structure, the phase change caused by external interference will not change the phase difference, so the interference pattern is stable, and it has the structural advantage of anti-interference. In addition to the common optical path design using the Mirau objective above, there are also some common optical path designs using two interferometers in series (such as Benoita laGuillaume E, Martins F, Boccara C, et al. High-resolution handheld rigidendomicroscope based on full-field optical coherence tomography[J].Journal of biomedical optics), these designs either cannot make the numerical aperture of the objective lens too large, or the reference optical path and the sample optical path cannot be completely symmetrical to bring additional optical path difference.

In order to improve some defects of the current interference microscope, the present invention proposes a completely symmetrical optical path design. The reference light and the sample light pass through the exact same path, but in different order, which eliminates the additional optical path difference caused by the system structure, and at the same time The numerical aperture is not limited, and it is not sensitive to environmental vibration.

SUMMARY OF THE INVENTION

In view of this, one of the objectives of the present invention is to provide an interference microscope, which, by inserting the first microscope objective lens in the compensation interference cavity, ensures that the optical paths of the sample light and the reference light are completely equal, and avoids Inconsistencies in the optics cause some points on the interference surface to have unequal optical paths.

For achieving the above object, the technical scheme of the present invention is:

An interference microscope, comprising a light source generating device, a compensation interference cavity, a detection arm, and a signal acquisition and processing unit;

The compensation interference cavity receives the light beam generated by the light source generating device and reflects it into a reflected light beam into the detection arm;

The detection arm focuses the reflected beam to form sample light, and sends the sample light to the signal acquisition and processing unit; wherein, the detection unit is provided with a second microscope objective lens, and the second display unit is provided with a second microscope objective lens. Microobjectives are used to focus light on a sample and modulate the structural information of the sample into light back.

Further, the light source generating device includes a light emitting device having a spectral width and a first lens.

The light source generating device includes a light emitting device with a spectral width and a first lens; wherein,

The compensation interference cavity includes a first beam splitter, a first reflector, a second reflector and a first microscope objective lens; the first microscope objective lens is located between the first beam splitter and the second reflector , the second reflector is at the focal plane of the first microscope objective, the main optical axis of the first microscope objective and the surface of the first beam splitter are at an angle of 40°-50°, and the first reflector passes through the The virtual image formed by the first beam splitter and the second reflector are parallel to each other, and the first reflector and the second reflector are located on two sides of the first beam splitter respectively.

Further, the detection arm includes a second lens, a second beam splitter, a third beam splitter and a second microscope objective lens; wherein, the third beam splitter and the main optical axis of the second microscope objective lens Vertically, the second beam splitter forms an angle of 40°-50° with the main optical axis of the second microscope objective lens and an angle of 80°-100° with the first beam splitter in the compensation interference cavity.

Further, the rear surfaces of the second beam splitter and the third beam splitter are semi-permeable films with a reflectivity of 5%-15%, and the front surfaces are anti-reflection films.

Further, the signal acquisition and processing unit includes a photoelectric sensor and a signal processor, the signal processor is electrically connected to the photoelectric sensor, and the sample light reaches the photoelectric sensor through the second lens.

Further, the photoelectric sensor includes an array photoelectric sensor and an area array photoelectric sensor.

Further, the photoelectric sensor front is conjugated to the sample through the second lens in the detection arm, the second microscope objective lens, and also through the second lens in the detection arm, the first microscope objective lens in the compensation interference cavity and the first microscope objective lens. The two mirrors are conjugated.

In view of this, the second purpose of the present invention is to provide an interference microscopy imaging method, which can ensure that the sample light and the reference light pass through the exact same microscope objective lens through adjustment, but the order is different. The optical paths are completely equal, which avoids the unequal optical path of some points on the interference surface caused by the inconsistency of the optical devices, and finally obtains a high-precision three-dimensional structure diagram inside the sample.

For achieving the above object, the technical scheme of the present invention is:

An interference microscopic imaging method, comprising the following steps:

The light beam generated by the light source generating device is received, the light beam is divided into two beams by the first beam splitter, one beam is reflected by the first reflector and then transmitted by the first beam splitter, and the other beam is passed through the first objective lens and then transmitted by the first beam splitter. second mirror reflection;

The two beams reach the second beam splitter, are reflected by the second beam splitter to the third beam splitter, and then a part of the beam is reflected by the third beam splitter to become the reference beam, and the other part is reflected by the third beam splitter. The beam mirror transmits, and the transmitted beam is focused on the sample by the second microscope objective lens and modulated by the sample to become sample light and reflected back to the second microscope objective lens;

The sample light and the reference light reach the front of the photoelectric sensor through the second beam splitter and the second lens;

The signal processor collects electrical signals from the photoelectric sensor to obtain structural information of the sample section.

Further, the sample light carries the result information of the sample.

Further, the step that the sample light and the reference light reach the front of the photoelectric sensor through the second beam splitter and the second lens specifically include:

Adjusting the position of the first mirror so that the optical path difference between the sample light and the reference light is within the coherence length range of the light source;

The sample light and the reference light reach the surface of the photoelectric sensor to form interference fringes.

Further, it also includes the steps:

By moving the position of the sample, the signal processor collects the interference fringes multiple times to form an internal three-dimensional structure diagram of the sample.

beneficial effect

The invention provides an interference microscope. The interference microscope inserts a first microscope objective lens into a compensation interference cavity, so that the reference light and the sample light pass through the exact same optical element, but the sequence is different, wherein the sample light first passes through the reflecting mirror and then passes through the The second microscope objective in the detection arm goes to the sample. The reference light first passes through the second microscope objective in the detection arm, then passes through the reflector, and then is reflected by the beam splitter. Using the same two microscope objectives can ensure two paths of light. The optical paths are completely equal, which avoids that not every point on the interference surface has the same optical path due to the inconsistency of the optical devices. At the same time, the present invention also provides an interference microscopic imaging method, in which the reference light and the The sample light passes through the exact same microscope objective lens, but the order is different, to ensure that the optical path of the sample light and the reference light are completely equal, and a high-precision three-dimensional structure map of the sample is obtained.

Description of drawings

In order to illustrate the embodiments of the present invention or the technical solutions in the prior art more clearly, the following briefly introduces the accompanying drawings that are required to be used in the description of the embodiments or the prior art. Obviously, the drawings in the following description are some embodiments of the present invention, and for those of ordinary skill in the art, other drawings can also be obtained from these drawings without creative effort.

1 is a schematic structural diagram of an embodiment of an interference microscope of the present invention;

FIG. 2 is a schematic structural diagram of another embodiment of an interference microscope of the present invention.

Detailed ways

In order to make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be described clearly and completely below with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are some, but not all, embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

The examples are given to better illustrate the present invention, but the content of the present invention is not limited to the examples. Therefore, those skilled in the art make non-essential improvements and adjustments to the embodiments according to the above-mentioned contents of the invention, which still belong to the protection scope of the present invention.

Example 1

Referring to FIG. 1 , a schematic structural diagram of an embodiment of an interference microscope of the present embodiment is shown. Specifically, an interference microscope includes:

Light source generating device, compensation interference cavity, detection arm, signal acquisition and processing unit; wherein,

The light source generating device includes a light-emitting device S with a spectral width and a first lens L1. In this embodiment, the light-emitting device S adopts an LED light source, the center wavelength λ0 = 850 nm, and the spectral width Δλ = 30 nm. The light source is collimated by the first lens L1 and becomes Quasi-parallel light;

The compensation interference cavity includes a first beam splitter BS1, a first mirror M1, a second mirror M2, a first microscope objective MS1, and the first microscope objective MS1 is located at the first beam splitter BS1 and the second mirror M2 In between, the second mirror M2 is located at the focal plane of the first microscope objective lens MS1, and the main optical axis of the first microscope objective lens MS1 and the surface of the first beam splitter BS1 are at an angle of 45°. The angle between the main optical axis of the microscope objective MS1 and the surface of the first beam splitter BS1 can be in the range of 40°-50°, and the first mirror M1 is formed by the first beam splitter BS1. The virtual image and the second mirror M2 are parallel to each other, the first reflection mirror M1 and the second reflection mirror M2 are located on both sides of the first beam splitter BS1 respectively, and the first reflection mirror M1 is also fixedly connected with the piezoelectric ceramic PZT. In this embodiment, the first reflection mirror M1 and the second reflector M2 are total reflectors;

The detection arm comprises a second lens L2, a second beam splitter BS2, a third beam splitter BS3, a second microscope objective MS2, the rear surfaces of the second beam splitter BS2 and the third beam splitter BS3 are semi-permeable membranes, The reflectivity is 5%-15%, the front surface is anti-reflection coating, the third beam splitter BS3 is perpendicular to the main optical axis of the second microscope objective MS2, and the second beam splitter BS2 is perpendicular to the main optical axis of the second microscope objective MS2 At an angle of 45° and at an angle of 80°-100° with the first beam splitter BS1 in the compensation interference cavity.

The signal acquisition and processing unit includes an array photoelectric sensor CAM and a signal processor PS. The photoelectric sensor CAM front is conjugated with the sample Sample through the second lens L2 and the microscope objective MS2 in the detection arm, and also passes through the second lens in the detection arm. The lens L2 and the first microscope objective lens MS1 in the compensation interference cavity are conjugated with the second mirror M2, so that both the sample and the second mirror M2 are imaged on the surface of the photoelectric sensor. The position of a beam splitter BS1 can ensure that the second mirror surface M2 and the sample Sample are imaged on the front surface of the photoelectric sensor CAM through the same optical path. The interference fringes are formed on the surface, the signal processor PS is electrically connected with the sensor CAM, and the signal processor PS is also connected with the electric ceramic PZT.

Referring to the arrows in FIG. 1 , an interference microscopic imaging method in this embodiment is:

The wide-spectrum light source emitted by the light-emitting device S is collimated by the first lens L1 and then divided into two beams by the first beam splitter BS1 in the compensation interference cavity, and one beam is reflected by the first mirror M1 and then split by the first beam. The mirror BS1 transmits it, and the first microscope objective lens MS1 passing through the other way is reflected by the second reflecting mirror M2 and then reflected by the first beam splitter BS1 to be emitted.

After the two beams from the compensation interference cavity enter the detection arm, they are both reflected by the second beam splitter BS2 in the detection arm to the third beam splitter BS3, and then both are reflected by the third beam splitter BS3 with a reflectivity of 10% and 90% transmittance (the reflectivity of the third beam splitter BS3 is set to be 10% in a specific embodiment), the transmitted light is focused on the sample by the second microscope objective lens MS2 and reflected back after being modulated by the sample. The second microscope objective MS2 and 10% of the reflected light from the third beam splitter BS3 are combined at the third beam splitter BS3 to form four beams (the four beams include two beams reflected by the third beam splitter BS3 at 10%) , two beams of light reflected by the sample from the second microscope objective lens MS2), these four beams then reach the front of the sensor CAM through the second beam splitter BS2 and the second lens L2 through the same path. Among these four beams, Two of the beams carry the structural information of the sample and become the sample light (the sample light is the light that reaches the sample through the second microscope objective MS2 and is reflected after being adjusted by the sample), and the other two beams do not have the structural information of the sample and become the reference light (reference light). The light is the light reflected by the third beam splitter BS3), and one of the sample light and the reference light has passed through the microscope objective lens once. By adjusting the position of the reference mirror (mirror M1) in the compensation interference cavity, the sample light and the reference light can be guaranteed. The optical path difference is within the range of the coherence length of the light source, so interference fringes can be formed on the surface of the photoelectric sensor, and the other two paths of light form background light superimposed on the interference fringes. The signal processor PS collects the electrical signal from the photoelectric sensor, and controls the voltage loaded on the PZT to drive the first mirror M1 to make a small displacement each time the interferogram is collected, and the displacement range shall not exceed one wavelength. The displacement of the first mirror M1 will change the optical path difference between the sample light involved in the interference and the reference light, and then move the interference fringes on the sensor CAM front. After collecting multiple interference images, the sample can be restored through numerical fitting. The reflectivity of the inner section perpendicular to the optical axis, and then the structural information of the section can be obtained. By moving the sample to scan the inner section, the obtained tomographic image (section structure information map) can be displayed on the display MN, and finally the internal structure of the sample is formed. Three-dimensional structural diagram, the interference microscope in this embodiment is also applicable to tissue biopsy.

An interference microscope in this embodiment adopts a common optical path design, and the detection arm can be designed to be very long and used for real-time detection in vivo, without the need for tissue biopsy sampling; thanks to the first microscope objective lens inserted in the compensation interference cavity MS1, so that the reference light and the sample light pass through the exact same optical elements, but the order is different, in which the sample light first passes through the first mirror M1 and then passes through the second microscope objective MS2 before being reflected by the sample, and the reference light first passes through the microscope. The objective lens MS1 is then reflected by the second reflector M2 and then reflected by the third beam splitter BS3. The use of two identical microscope objective lenses can ensure that the optical paths of the two paths of light are completely equal, avoiding the inconsistency caused by the inconsistency of the optical devices. Not every point on the interference surface has the same optical path.

Example 2

As shown in FIG. 2 , this embodiment adopts the same compensation interference cavity as that of Embodiment 1, except that the image transmission unit Green lens rod or the image transmission fiber bundle (F-B) is inserted into the detection arm. Both the reference light and the sample light are transmitted in the imaging unit without changing the optical path difference, so endoscopic interference imaging can be realized. The specific imaging process is the same as that in Embodiment 1, and will not be repeated here.

The embodiments of the present invention have been described above in conjunction with the accompanying drawings, but the present invention is not limited to the above-mentioned specific embodiments, which are merely illustrative rather than restrictive. Under the inspiration of the present invention, without departing from the scope of protection of the present invention and the claims, many forms can be made, which all belong to the protection of the present invention.

Claims (10)

1. an interference microscope, is characterized in that, comprises light source generating device, compensation interference cavity, detection arm, signal acquisition and processing unit; The compensation interference cavity receives the light beam generated by the light source generating device and reflects it into a reflected light beam into the detection arm; The detection arm is used for focusing the reflected beam to form sample light, and sending the sample light to the signal acquisition and processing unit; wherein, the detection unit is provided with a second microscope objective lens, and the first Two microscope objectives are used to focus light on the sample and modulate the structural information of the sample into light return. 2 . The interference microscope according to claim 1 , wherein the light source generating device comprises a light-emitting device with a spectral width and a first lens, and the compensation interference cavity comprises a first beam splitter, a second a reflector, a second reflector, and a first microscope objective lens; wherein, The first microscope objective is located between the first beam splitter and the second reflector, the second reflector is located at the focal plane of the first microscope objective, and the main optical axis of the first microscope objective is connected to the first The surface of the beam mirror is at an angle of 40°-50°, and the virtual image formed by the first mirror through the first beam splitter and the second mirror are parallel to each other, and the first mirror and the second mirror are parallel to each other. The reflecting mirrors are respectively located on both sides of the first beam splitter. 3 . The interference microscope according to claim 2 , wherein the detection arm comprises a second lens, a second beam splitter, a third beam splitter and a second microscope objective lens; wherein the first The third beam splitter is perpendicular to the main optical axis of the second microscope objective, and the second beam splitter is at an angle of 40°-50° with the main optical axis of the second microscope objective, and is at an angle of 40°-50° with the first splitter in the compensation interference cavity. The beam mirror is at an angle of 80°-100°. 4 . The interference microscope according to claim 3 , wherein the rear surfaces of the second beam splitter and the third beam splitter are semi-permeable films with a reflectivity of 5%-15%. 5 . , the front face is anti-reflection coating. 5. The interference microscope according to claim 4, wherein the signal acquisition and processing unit comprises a photoelectric sensor and a signal processor, the signal processor is electrically connected to the photoelectric sensor, and the sample light passes through The second lens reaches the photosensor. 6. The interference microscope according to claim 5, wherein the photoelectric sensor front is conjugated to the sample through the second lens and the second microscope objective lens in the detection arm, and also passes through the second lens in the detection arm and the second microscope objective lens. Two lenses, the first microscope objective in the compensation interference cavity and the second mirror are conjugated. 7. A method of interference microscopy imaging, characterized in that, comprising the following steps: The light beam generated by the light source generating device is received, the light beam is divided into two beams by the first beam splitter, one beam is reflected by the first reflector and then transmitted by the first beam splitter, and the other beam is passed through the first objective lens and then transmitted by the first beam splitter. second mirror reflection; The two beams reach the second beam splitter, are reflected by the second beam splitter to the third beam splitter, and then a part of the beam is reflected by the third beam splitter to become the reference beam, and the other part is reflected by the third beam splitter. The beam mirror transmits, and the transmitted beam is focused on the sample by the second microscope objective lens and modulated by the sample to become sample light and reflected back to the second microscope objective lens; The sample light and the reference light reach the front of the photoelectric sensor through the second beam splitter and the second lens; The signal processor collects electrical signals from the photoelectric sensor to obtain structural information of the sample section. 8 . The interference microscopy imaging method according to claim 7 , wherein the sample light carries the result information of the sample. 9 . 9 . The interference microscopy imaging method according to claim 7 , wherein the step that the sample light and the reference light reach the front of the photoelectric sensor through the second beam splitter and the second lens specifically comprises: 10 . Adjusting the position of the first mirror so that the optical path difference between the sample light and the reference light is within the coherence length range of the light source; The sample light and the reference light reach the surface of the photoelectric sensor to form interference fringes. 10. A method of interference microscopy imaging according to claim 9, characterized in that, further comprising the steps of: By moving the position of the sample, the signal processor collects the interference fringes multiple times to form an internal three-dimensional structure diagram of the sample.

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