WO2012077936A2

WO2012077936A2 - Fluorescence-sensing probe, and fluorescence detection method, sentinel lymph node dissection method and target-guided surgery method which use the probe

Info

Publication number: WO2012077936A2
Application number: PCT/KR2011/009271
Authority: WO
Inventors: Seok Ki Kim; Byeong Il Lee
Original assignee: National Cancer Center
Priority date: 2010-12-07
Filing date: 2011-12-01
Publication date: 2012-06-14
Also published as: KR101166556B1; KR20120063342A; WO2012077936A3

Abstract

The present invention relates to a fluorescence-sensing probe which can detect a substance, which has been bound to or which contains therein a fluorescent dye that is distributed in a human or animal body, at high sensitivity in proximity to tissue, as well as a fluorescence detection method that uses the probe. This fluorescence-sensing probe can be used in sentinel lymph node dissection or target-guided surgery and is highly practical, because it can be suitably used for a surgical technique, such as pressing or turning the probe in proximity to the skin surface of a human or animal body during surgery.

Description

FLUORESCENCE-SENSING PROBE, AND FLUORESCENCE DETECTION METHOD, SENTINEL LYMPH NODE DISSECTION METHOD AND TARGET-GUIDED SURGERY METHOD WHICH USE THE PROBE

The present invention relates to a fluorescence-sensing probe, and a fluorescence detection method, a sentinel lymph node dissection method and a target-guided surgery method which uses the probe, and more particularly to a fluorescence-sensing probe capable of detecting a substance, which has been bound thereto or contains therein a fluorescent dye that is distributed in a human or animal body, at high sensitivity in proximity to tissue, and a fluorescence detection method, a sentinel lymph node dissection method and a target-guided surgery method which uses the probe.

1) Advantages, disadvantages and limitations of fluorescence technology

Fluorescence is harmless to a living body and involves relatively simple systems, and thus is highly cost efficient. However, there are problems due to the fact that the penetrating power of fluorescence is insufficient for application to a living body, particularly, to a human body, is significantly influenced by the surrounding light, such as sunlight or electric light, and can be influenced by various factors, including the kind of light source, the kind of filter and the relationship between the fluorescence and a living body. Hence, even though near infrared fluorescence is used for a biometric image, the fluorescence from the tissue with the depth of more than some centimeters cannot be sensed by a fluorescence camera according to the existing technology. Here, theoretically some centimeters may be the limit, but when actually applied to a living body, the sensing is difficult even in the depth of 1cm.

Also, fluorescence has a limitation associated with low sensitivity due to autofluorescence. Despite such disadvantages and limitations, there is a high possibility that fluorescence will be widely used in a living body, particularly a human body, because it has various advantages.

2) Status of related technologies and problems to be solved for applying fluorescence to living body

There are various types of technologies for applying fluorescence to a living body, and among them, the technology of distributing a fluorescent drug or agent in a living body by a variety of mechanisms, and non-invasively measuring the distributed location and amount of the fluorescence in the living body is an important technology in this area.

The most widely used method for non-invasively measuring distributed location and amount of the fluorescence in the living body is photographing fluorescence in the living body using a fluorescence camera. A representative method of detecting fluorescence in the living body is observing with eyes using the above described fluorescence camera and an optical filter. In the case of fluorescent light emitted from mainly visible ray, it is possible to use a method of directly observing with eyes using an optical filter through which only the wavelength region, to which the fluorescent ray to be measured belongs, passes. However, the fluorescence of the visible ray area is disadvantageous in sensitive measuring due to the low in-body penetration rate.

In this method, many unpredictable events can occur in a process during which fluorescence emitted from a fluorescent drug or agent reaches the fluorescence camera in which it is converted to an electrical signal to produce an image. Among them, the biggest variables are tissue-related variables, such as the thickness and type of tissue through which either excitation light itself, which causes fluorescence, or fluorescence light, passes. The fluorescence camera method, which photographs images with a certain distance with the body where there are fluorescent materials, is not optimal for sensing or accepting emitting fluorescence generated by light, but is appropriate for generating two-dimensional images using emitting fluorescence. Namely, if the thickness of tissue is high, it will be difficult to sense fluorescence with the fluorescence camera that is located far away from the tissue. This is because emitted fluorescence does not reach the camera if the tissue gets thicker because the emitting fluorescence is absorbed in proportion to the thickness of the tissue. Also, because the distance between tissue and the fluorescence camera is generally far, the amount of fluorescence that is additionally sensed is reduced, suggesting that this method has a significantly low sensitivity compared to a method of measuring fluorescence in proximity to tissue.

Even though the emitting fluorescence is not absorbed while passing the tissue, the emitting fluorescent light is diffusely-scattered within the tissue. Such diffused light cannot form an image, cannot be sensed by a fluorescent camera, and cannot be transmitted to the lens of the light camera if positioned a little away from the body because the direction is different from the direction of the excitation light. To solve this problem, various methods are used, including radiating with a strong excitation light so that the fluorescence camera can receive fluorescence light in as large an amount as possible, or using lenses having higher light-condensing capability, or using a type of fluorescent light that is less absorbed by tissue. However, very strong excitation light does not secure the safety for normal tissues, and thus it is difficult to raise up the strength of the excitation light, and there is a limit in improving performance of the condensing lens and the camera. Further, fluorescent conversion capacity of the fluorescent material itself is not excellent in the case of fluorescent materials that satisfy the condition that the human body or living body should not be harmed.

Herein, the fluorescent light that is less absorbed into tissue is advantageously fluorescent light in the near-infrared region, which has high penetrating power in vivo. The term near-infrared region refers to the region outside the color red in the light spectrum. Generally, the shortest wavelength region of infrared light (0.75-3 ㎛) is referred to as the near-infrared region.

This light in the near-infrared region includes an electromagnetic spectrum, shows thermal action, photographic action, photoelectric action and fluorescent action, exhibits sterilizing or disinfecting effects and acts to treat joints and muscles. Thus, it is generally used in industrial and medical applications. Particularly, light in the near-infrared region is relatively less absorbed in vivo than light in other wavelength ranges, and thus near-infrared light occurring in a deep area in vivo can also be detected ex vivo. Invisible fluorescence of the near-infrared region can be photographed by using a fluorescence camera. A fluorescence camera should be additionally used compared with visible fluorescence of the visible light region, but because in vivo permeability of near-infrared light is higher than light of other wavelength region, which is the characteristics of the near-infrared light, sensitive measuring is possible. Because invisible light is used, the observed part does not need to be darkened for the detection of the fluorescent light of interest.

The current method of detecting in vivo fluorescence is typically a method of visually observing fluorescence using the fluorescence camera and an optical filter. For fluorescent light that is emitted mainly in the visible light region, it is possible to use a method of visually observing the fluorescent light using an optical filter that passes only the wavelengths in which fluorescent light is present. However, fluorescence in the visible light region has low in vivo permeability and thus is disadvantageous for sensitive measurements. The method that uses a fluorescence camera is suitable for use if invisible fluorescence in the near-infrared region is used. Although this method should additionally use a fluorescence camera, it is advantageous for sensitive measurement, because near-infrared light has high in vivo permeability. Also, it uses invisible light, and thus eliminates the need to use a method that darkens the observation site.

The method of either visually observing fluorescence or observing fluorescence through an image using a camera is conveniently used to observe relatively wide sites, but is difficult to clinically use for relatively deep tissue, because it has low sensitivity due to autofluorescence and attenuation of in vivo tissue. Also, when the fluorescence is distributed or present in a narrow area rather than a wide area and should be observed, it is necessary to use a probe-type device in proximity to tissue rather than obtain an image.

3) Limitations of technology associated with sentinel lymph node biopsy and target-guided surgery

(A) Regarding sentinel lymph node biopsy

Lymph node dissection is necessarily required for surgical treatment of various cancers such as breast cancer. However, if lymph nodes are extensively dissected, complications such as lymphoedema can occur that significantly reduce the quality of life of the patient, even though the tumor is treated. For this reason, extensive lymph node dissection should be performed in a minimum range of that necessarily required by a patient.

The method used either to determine whether a patient requires extensive lymph node dissection or to determine the range of lymph node dissection is a surgery for determining sentinel lymph node biopsy. The sentinel lymph node refers to the first lymph node receiving lymphatic drainage from

the area of the primary tumor. The sentinel lymph node can represent the state of all the lymph nodes, that is, the metastasis of a tumor. Thus, when the sentinel lymph node is removed while minimizing damage to a living body (e.g., minimizing the size of the dissected site without unnecessarily extensively dissecting lymph nodes), whether extensive lymph node dissection is to be performed can be determined by examining only the sentinel lymph node. In order to find the sentinel lymph node, a diagnostic agent that is drained and accumulated in the sentinel lymph node is administered, the sentinel lymph node is found in various manners and dissected, and the dissected sentinel lymph node is pathologically examined.

Current methods of finding the sentinel lymph node by detecting a signal emitted from the sentinel lymph node that the diagnostic agent has reached are largely divided into two methods: a method employing a radioactive material; and a method employing a dye.

The radioactive material has very excellent in vivo permeability, and thus makes it very easy to detect a signal ex vivo. It can be imaged using a gamma-camera and can also be measured using a gamma-detector. However, the radioactive material requires special equipment such as a gamma-camera or detector, related equipment, related approval and related manpower, and thus is difficult to use in a small-scale hospital or a hospital in which radioactivity-related equipment is not provided.

The method employing the dye is a very convenient method, because it does not require special equipment, related equipment, related approval and related manpower. However, the method is poor at detecting the sentinel lymph node, and thus is used in combination with the method employing the radioactive material rather used in isolation. In this case, the above-mentioned problems arise.

(B) Regarding target-guided surgery

Generally, surgery aims to dissect a lesion while minimizing damage to normal tissue and minimizing damage to function. To achieve this goal, it is most important to accurately determine the location of the lesion and to more clearly distinguish the boundary between the lesion and the surrounding tissue.

However, in some cases, it is not easy to distinguish the boundary between the lesion and the surrounding tissue, particularly when the lesion is small in size. Meanwhile, with the development of diagnostic technology, it frequently happens that a fine lesion is located but it is not always easy to accurately dissect the fine lesion found during diagnosis. In many cases, even though the fine lesion was located by a diagnostic device, it is invisible to the surgeon s eye or a surgical microscope, and particularly, it is difficult to reliably find the boundary between the lesion and the surrounding tissue.

Because of this background and for this reason, surgical treatment of inducing visibility of the location of a fine lesion (that is, guiding a target) during surgery in various ways and dissecting only the fine lesion has received attention. This method of surgical treatment is called target-guided surgery . In methods for targeting a lesion, similar to the method of finding the sentinel lymph node, a diagnostic agent that is selectively accumulated in the lesion is administered, and the lesion is dissected while confirming the location of the agent during surgery using a device allowing the distribution of the agent to be confirmed. By doing so, it is possible to reliably dissect a fine lesion which is invisible to the eyes or a surgical microscope. The method employing the sentinel lymph node as described above is also a kind of target-guided surgery. This method is also being widely used, because the method employing the radioactive material is very advantageous. However, this method also suffers from the above-described problems, because it uses a radioactive material.

Accordingly, there is a need for an apparatus and method capable of more effectively and conveniently guiding a target without having to use a radioactive material.

It is an object of the present invention to provide a fluorescence sensing probe capable of detecting a substance, which has been bound thereto or contains therein a fluorescent dye that is distributed in a human or animal body, at high sensitivity when contacting tissue or in proximity to tissue.

Another object of the present invention is to provide a fluorescence detection method capable of detecting a substance which has been bound thereto or contains therein a fluorescent dye that is distributed in a human or animal body, using said fluorescence-sensing probe.

Still another object of the present invention is to provide a method for dissecting a sentinel lymph node, which can more effectively detect the sentinel lymph node using said fluorescence detection method.

Still another object of the present invention is to provide a target-guided surgery method capable of more effectively examining and dissecting a specific fine lesion using said fluorescence detection method.

To achieve the above objects, in one aspect, the present invention provides a fluorescence-sensing probe including: a light source that emits light of specific wavelength; an excitation filter that selectively transmits, among the light emitted from the light source, only excitation light exciting a fluorescent substance included in vivo; an emission filter that selectively transmits only fluorescence emitted from the fluorescent substance; and a light-sensing element capable of detecting fluorescence passed through the emission filter.

As used herein, the term "probe" is one used to bring an input terminal such as a vacuum tube voltmeter (VTVM) or an oscilloscope into contact with a measurement point. The constituent element of the probe varies depending on a signal to be measured. Typical signals include temperature, pressure, pH, etc., and a typical probe for medical applications is an ultrasonic probe. The input may be a one-dimensional signal, and may, if necessary, be a high-dimensional signal, such as a two-dimensional signal (image, etc.) or a three-dimensional signal. The probe described herein is a device for medical or biological applications and usually has an elongated shape which is easily used in contact with a specific biological site such as a wound or a surgical site.

Preferably, the probe serves to detect ex vivo a fluorescent substance in the visible light or near-infrared region in proximity to or in contact with the fluorescent substance or to detect the fluorescent substance in contact with a surgical site.

As used herein, the term "near-infrared region" generally refers to the near-infrared region known in the art. The use of a wavelength in the near-infrared region has an advantage in that the location of a sentinel lymph node can be found without cutting the skin.

As used herein, the term "visible-light region" refers to a region having a visible wavelength range in the light spectrum and generally has a wavelength range of 380-770 nm. In the visible-light region, the change in properties according to wavelength appears as a color, and wavelengths get progressively shorter from red to violet. Monochromatic light appears red at 700-610 nm, orange at 610-590 nm, yellow at 590-570 nm, green at 570-500 nm, blue at 500-450 nm, and violet at 450-400 nm.

Preferably, the excitation filter is connected with one end of a first optical fiber, and the emission filter is connected with one end of a second optical fiber, in which the other ends of the first and second optical fibers are received in the same housing.

Preferably, one of the excitation filter and the emission filter is connected with one end of an optical fiber, and the other end of the optical fiber is received in the same housing.

Preferably, the housing comes in contact with the surface of a living body to excite a fluorescent material included in the living body and to detect fluorescence emitted from the fluorescent material.

Preferably, the housing can sense emitted fluorescence even when contacting or in proximity to the surface of the living body.

Preferably, the housing can concentrate and sense efficiently emitted light that passes or scattered the living body, by contacting or in proximity to the surface of the living body.

Preferably, the housing can temporarily reduce the thickness of the tissue or temporarily reduce blood within tissue between our sensing unit and fluorescent material of interest by giving pressure to the tissue by contacting or in proximity to the surface of the living body. Blood is the main material for absorbing fluorescence in the tissue.

Preferably, the housing can avoid a tissue in which it is difficult to pass the fluorescence, by changing the direction of the light receiving part or by contacting or in proximity to the surface of the living body.

Preferably, the light source is any one or more of a laser, a laser diode, an LED, a halogen lamp, and a xenon lamp.

Preferably, the light-sensing element is any one or more of CCD, CMOS, and a photodiode.

Preferably, the fluorescence-sensing probe may further include an amplifier capable of amplifying the detected fluorescence.

Preferably, the fluorescence-sensing probe may further include a display unit capable of displaying the detected fluorescence.

Preferably, the fluorescence-sensing probe may further include a display unit capable of displaying the detected fluorescence, in which the display unit is a light-emitting device that is provided on the housing.

Preferably, the excitation filter and the emission filter are provided in plurality or configured such that they can be selectively replaced.

Also, the fluorescence detection method according to the present invention includes the steps of: (a) injecting into a subject a substance which has been bound thereto or contains therein a fluorescent dye; and (b) detecting fluorescence in the subject using the fluorescence-sensing probe.

Preferably, the substance which has been bound thereto or contains therein the fluorescent dye is indocyanine green-labeled human serum albumin.

Preferably, the subject includes a human or animal body.

As used herein, the term "indocyanine green" refers to a widely used near-infrared dye for fluorescence imaging, which is degraded or excreted in the urine and feces about 1 hour after having been injected into the body of a human or an animal. Thus, it is a fluorescent dye that can be used in a human or animal body and is advantageous for clinical applications.

In another aspect, the present invention provides a sentinel lymph node biopsy method including the steps of: (a) injecting into a living body a substance which has been bound thereto or contains therein a fluorescent dye; (b) placing the above-described fluorescence-sensing probe in proximity to the living body to detect a sentinel lymph node present in the living body; and (c) dissecting the detected sentinel lymph node.

In still another aspect, the present invention provides a target-guided surgery method including the steps of: (a) injecting into a living body a substance which has been bound thereto or contains therein a fluorescent dye and is selectively accumulated in a specific lesion; (b) placing the above-described fluorescence-sensing probe in proximity to the living body to confirm the location of the specific lesion present in the living body; and (c) dissecting the confirmed specific lesion.

According to the present invention, a substance which has been bound thereto or contains therein a fluorescent dye that is distributed in a human or animal body can be detected at high sensitivity in proximity to tissue.

Also, according to the present invention, a sentinel lymph node can be more effectively dissected by applying fluorescence detection when performing dissection thereof. Furthermore, by applying the fluorescence detection method to target-guided surgery, a specific fine lesion can be more effectively examined and dissected.

The fluorescence-sensing probe according to the present invention has an advantage in sensing weak emitted fluorescence by detecting emitted fluorescence by contacting the surface of the skin unlike the existing camera method. Further, because the emitted fluorescence can be sensed by contacting the body, the thickness of the region where fluorescence exists can be reduced or the fluorescence absorbing materials can be avoided using a method of pushing the body or changing the direction, and thus the emitted fluorescence incident upon the light receiving unit is strengthened and it becomes easy to sense weak emitted fluorescence. Thus, it is clinically useful and can more effectively detect a fluorescent target material distributed in vivo, compared to a conventional method employing a fluorescence camera. Accordingly, if it is used in place of or in combination with the fluorescence camera method, the clinical utility of fluorescence can be significantly improved. Further, fluorescence can be applied to sentinel lymphadencetomy or target-guided operation which has been difficult due to low tissue permeability of fluorescent materials.

Also, a fluorescent substance has no risk of radioactive radiation or antipathy, unlike a radioactive marker. It allows the application of a multiplexing imaging method that uses two or more fluorescence wavelengths simultaneously, such that the locations of several fluorescent markers can be detected at one time. Thus, it may find new clinical applications.

Furthermore, the fluorescence-sensing probe according to the present invention can reduce the possibility of false-negative diagnosis which is high when a fluorescence camera is used to detect fluorescence in vivo. Also, it can reduce the accuracy of targeted surgery.

In addition, since the fluorescence-sensing probe according to the present invention employs a small number of elements and fewer people compared to a conventional method, it can facilitate the use of sentinel lymph node dissection in hospitals and assist in applying sentinel lymph node dissection to various types of cancer. Also, it can be widely used in various applications, including endoscopy, laparoscopy and robot-assisted surgery.

The above and other objects, features and advantages of the present invention will be more clearly understood from the following detailed description taken in conjunction with the accompanying drawing, in which:

FIG. 1 shows a schematic configuration of a fluorescence-sensing probe 100 according to one embodiment of the present invention;

FIG. 2 shows an entire system including the fluorescence-sensing probe 100 according to one embodiment of the present invention;

FIG. 3 is a set of photographs showing whether fluorescence was detected, in which FIG. 3(a) is a photograph showing that no fluorescence was detected in the left foot pad and left inguinal region of a mouse, and FIG. 3(b) is a photograph showing that fluorescence was detected in the right foot pad and right inguinal region of a mouse; and

FIG. 4(a) is a photograph showing that a sentinel lymph node dissection is performed using the fluorescence-sensing probe 100 according to the present invention; and FIG. 4(b) is a photograph showing that the sentinel lymph node was suitably dissected by dissecting a site thought to be the sentinel lymph node and detecting fluorescence from the dissected sentinel lymph node using the fluorescence-sensing probe 100 according to the present invention.

Hereinafter, preferred embodiments of a fluorescence-sensing probe according to the present invention and a method for detecting fluorescence using the fluorescence-sensing probe will be described with reference to the accompanying drawings. Herein, the thickness of lines or the dimensions of components constituting the present invention may be exaggerated in the drawings for clearness and convenience. The terms are defined in consideration of the functions of the components in the present invention, and can be different depending on the intentions and usual practices of users and operators. Therefore, definition of these terms must be determined in the context of the overall contents of the specification.

<Embodiments>

FIG. 1 shows a schematic configuration of a fluorescence-sensing probe 100 according to one embodiment of the present invention, and FIG. 2 shows an entire system including the fluorescence-sensing probe 100 according to one embodiment of the present invention.

The fluorescence-sensing probe 100 according to one embodiment of the present invention will now be described with reference to FIGS. 1 and 2.

The fluorescence-sensing probe 100 comprises a light source 110, an excitation filter 120, an emission filter 130 and a light-sensing element 140. Also, the probe 100 may additionally comprise an amplifier 150 and a display unit 160.

The light source 110 serves to emit light of a specific wavelength. This light source 110 may be any one of a laser, a laser diode, an LED, a halogen lamp, a xenon lamp, and a combination of two or more thereof. Namely, the type of light source 100 used may vary depending on the type of fluorescent substance used.

The excitation filter 120 serves to selectively transmit, from among the light emitted from the light source 110, only excitation light that excites a fluorescent substance distributed in an animal or human body. The type of excitation filter 120 used may also vary depending on the type of fluorescent substance used. Further, the excitation filter 120 can be omitted in the case where the excitation light is selectively diffused from the light source.

The emission filter 130 serves to selectively transmit only light emitted from the fluorescent substance by the excitation light. The type of emission filter 130 used may also vary depending on the type of fluorescent substance used.

The light-sensing element 140 serves to detect fluorescence that has passed through the emission filter 130. This light-sensing element 140 may be any one of CCD, CMOS, a photodiode, and a combination of two or more thereof. Namely, the type of light-sensing element 140 is not specifically limited, as long as it can accurately detect the amount of fluorescence emitted from the fluorescent substance distributed in the human or animal body.

Meanwhile, as shown in FIGS. 1 and 2, the excitation filter 120 is connected with one end of a first optical fiber 120a, and the emission filter 130 is connected with one side of a second optical fiber 130a. The other end of each of the first optical fiber 120a and the second optical fiber 130a may be received in the same housing.

Although the first optical fiber 120a and the second optical fiber 130a are illustrated separately herein, it should be understood that one optical fiber may also serve as both the first optical fiber (120a) and the second optical fiber 130a.

Namely, it is to be understood that one optical fiber having excellent filtering performance may be used to perform both the excitation and emission of light.

This housing 170 serves as an actual probe and has an elongated shape such that it is conveniently used in contact with a specific biological site such as a wound or a surgical site. Namely, a user or a surgeon may manually bring this housing 170 into contact with a specific biological site and may non-invasively measure ex vivo the distributed location and amount of fluorescence.

Namely, the other end of each of the first optical fiber 120a and the second optical fiber 130a is received in the same housing, and the tip of the housing 170 is placed in proximity to or comes in contact with the surface of the living body, whereby only excitation light can be emitted through the tip of the same housing 170, thereby exciting the fluorescent substance included in the living body and detecting fluorescence emitted from the fluorescent substance.

Meanwhile, in this embodiment, the light source 110 and the light-sensing element 140 are connected with the first optical fiber 120a and the second optical fiber 130a, respectively, so as to emit excitation light from the tip of the housing 170 or sense fluorescence, but it is to be understood that the light source 110 and the light-sensing element 140 are not connected with the optical fibers, and may be placed directly within in the housing 170 so as to directly emit excitation light or to detect fluorescence. In this case, the fluorescence-sensing probe 100 may function as one module such that it can operate in a wireless manner without using any optical fiber.

The amplifier 150 serves to increase the sensitivity of fluorescence by amplifying detected fluorescence. This amplifier 150 may be any one of a PM tube, a semiconductor, a vacuum tube, and a combination of two or more thereof. Namely, it should be understood that various amplifiers may be used.

Meanwhile, it is to be understood that this amplifier 150 is an optional element and may be included or excluded according to the intention of a user.

The display unit 160 serves to display the intensity of in vivo fluorescent substance detected by the light-sensing element 140. Thus, the display unit 160 can display the distribution of the fluorescent substance in the surface of the living body with which the fluorescence-sensing probe 100 comes into contact, such that the distribution of the fluorescent substance can be quantitatively determined ex vivo. The configuration of this display unit 160 is known in the art, and thus the detailed description thereof will be omitted herefrom.

Although the display unit 160 illustrated herein is shown as a separate monitor that is located separated apart from the housing 170 by a distance, it is to be understood that the type of display unit 160 does not need to be limited to the illustrated configuration.

Namely, in another embodiment, the display unit 160 may also be a light-emitting device that is attached directly to the housing 170 or provided on the housing 170. For example, if the display unit 160 is a light-emitting device that is attached directly to the housing or provided on the housing 170, it may be configured such that either the degree of light emission increases in proportion to the intensity of fluorescence of a specific wavelength or it emits light when the intensity of fluorescence detected is higher than a specific threshold value.

According to this configuration, a user or a surgeon can more easily determine whether fluorescence was detected, using the fluorescence-sensing probe 100, without turning the head or directing the eye toward the sides.

Meanwhile, the excitation filter 120 and the emission filter 130 may be provided in plurality or be configured such that they can be selectively replaced. Thus, it is possible to perform a multispectral analysis in which excitation light of various wavelengths is received at one time and separately detected according to wavelength. Also, it is possible to perform multiple fluorescence detection for two or more fluorescent substances.

It is to be understood that this fluorescence-sensing probe 100 may be used together with optical therapeutic systems such as a laparoscopy device, a laparoscopy device or a surgery robot. Also, this fluorescence-sensing probe 100 may be used together with an existing imaging system such as an ultrasonic system.

According to this configuration, the fluorescence-sensing probe 100 of the present invention can sensitively detect a fluorescent dye-containing material, distributed in the body, for example, by bringing the probe into direct contact with the body surface or a surgical site, and applying pressure to the contacted probe. Also, it can accurately confirm the spatial location of the distributed site of the fluorescent substance by detecting only the fluorescent substance in a local area surrounding the site with which the probe is in direct contact.

The principle and advantages of the fluorescence-sensing probe 100 according to the present invention will now be described again.

The fluorescence-sensing probe 100 according to the present invention is not a device for imaging the spatial distribution of fluorescence like a camera, but is a device for measuring only the amount of fluorescence around a site with which the fluorescence-sensing probe 100 comes in direct contact or to which the probe is placed in proximity. Namely, the fluorescence-sensing probe 100 measures only fluorescence which is near the probe while existing within a narrow arrow ahead of the probe, and thus the probe can accurately detect the position of the fluorescence. Namely, if measurements are carried out while the probe is moved along the surface of a living body, the spatial distribution of the fluorescence can be deduced and the site of the fluorescence can be accurately determined. Because the probe is used in this way, it is suitable for surgery or animal studies.

Also, because the fluorescence-sensing probe 100 is used in such a manner that it comes in contact with the site in a human or animal body, it can receive a significantly strong signal compared to a method that uses a fluorescence camera in a fluorescence emission area, and thus it can accurately find a weak or small lesion which the fluorescence camera has difficulty sensing.

The probe is advantageous in that it can be positioned at an appropriate angle in tissue while being kept away from a site blocking the movement of fluorescence, for example, bone or a site with thick muscle. Also, the probe comes in close contact with a living body, and this is advantageous in that the possibility of intervention with the surrounding light or other variables causing error is low.

Also, because the fluorescence-sensing probe according to the present invention operates in direct contact with the surface of a living body, it can employ strong incident excitation light and also effectively receive emitted fluorescent light, unlike a method that uses a fluorescence camera. Accordingly, the probe has increased sensitivity, and thus can overcome the disadvantage of the method that uses a fluorescent substance or a fluorescence camera.

Furthermore, it does not obtain a two-dimensional or three-dimensional fluorescence image using emitted fluorescent light, but measures only the intensity of the sum of all fluorescent signals, such that the fluorescent signals can be more effectively used. Also, it is connected to the amplifier, such that fluorescence having different sensitivities can be more effectively detected. Further, it is more efficient because diffused emitted light, which is not used in the fluorescence camera method, is also used.

Moreover, the fluorescence-sensing probe can be manually handled by a surgeon during surgery, and thus can be very skillfully handled by the surgeon.

Also, it can simultaneously measure fluorescent substances of various wavelengths by adjusting incident excitation light and fluorescence emission light, thus making multiple fluorescence detection possible. As a fluorescent substance to be detected by the fluorescence-sensing probe of the present invention, any fluorescent substance may be used without limitation, as long as the excitation and emission wavelengths thereof sufficiently differ from each other. For the human body, a fluorescent dye of near-infrared wavelength which has low attenuation in the human body is advantageous.

Hereinafter, the fluorescence detection method according to the present invention will be described in detail with reference to the experimental examples. The fluorescence detection method according to the present invention comprises the steps of: (a) injecting into a subject a substance which contains therein or has been bound to a fluorescent dye; (b) detecting fluorescence using a fluorescence-sensing probe.

Preparation of experimental materials

As a light source, a 780-nm LD laser (Coherent) was used. Specifically, it had a wavelength of 784.1 nm and was used at a power of 31.54 mW and a current of 118 mA.

As an optical fiber cable, a dual branch light guide (NT54-202) optical fiber cable (Edmunds Optics) with a diameter of about 1/4 inches was used.

As a light-sensing element, a WAT-902H2 supreme fluorescence camera (WATEC) was used. To the camera lens was attached a long pass filter (Thorlabs) serving as an emission filter that passes only wavelengths higher than 820 nm.

A received image signal was transmitted to an LCD monitor to detect fluorescence.

As a fluorescent substance, indocyanine green-labeled albumin was used.

Mice used in the experiment were purchased from Central Lab. Animal Inc. (Korea).

Experimental Example: Experiment on detection of mouse sentinel lymph node

The left hind foot pad of the mice was subcutaneously injected with 10 ㎕ of albumin, and the right hind foot pad was subcutaneously injected with 10 ㎕ of indocyanine green-labeled albumin.

30 minutes after the injection, the fluorescence-sensing probe 100 according to the present invention was brought into contact with both the pads and both the inguinal regions of the mice to detect fluorescence. Then, a sentinel lymph node was detected using the fluorescence-sensing probe 100 and dissected using a surgical instrument. Then, the detection of fluorescence from the sentinel lymph node was confirmed.

Discussion of results

FIG. 3 is a set of photographs showing whether fluorescence was detected, in which FIG. 3(a) is a photograph showing that no fluorescence was detected in the left foot pad and left inguinal region of a mouse, and FIG. 3(b) is a photograph showing that fluorescence was detected in the right foot pad and right inguinal region of a mouse.

Referring to FIG. 3, it could be seen that, by the fluorescence-sensing probe 100 according to the present invention, no fluorescence was detected in the left foot pad and left inguinal region injected with albumin, but strong fluorescence was detected in the right foot pad and right inguinal region injected with indocyanine green-labeled albumin.

FIG. 4(a) is a photograph showing that sentinel lymph node dissection is performed using the fluorescence-sensing probe 100 according to the present invention; and FIG. 4(b) is a photograph showing that the sentinel lymph node was suitably dissected by dissecting a site thought to be the sentinel lymph node and detecting fluorescence from the dissected sentinel lymph node using the fluorescence-sensing probe 100 according to the present invention.

Referring to FIG. 4, it can be seen that, when the dissected tissue was examined using the fluorescence-sensing probe 100 of the present invention, strong fluorescence was detected and fluorescence in the dissected site was significantly reduced. Also, the dissected tissue was pathologically examined and, as a result, it was found to be lymph node tissue. Namely, it could be seen that a sentinel lymph node could be very accurately dissected using the fluorescence-sensing probe 100 of the present invention.

Hereinafter, the effects of surgery that can be performed in the human body using the fluorescence-sensing probe 100 of the present invention will be examined.

An example of using the fluorescence-sensing probe 100 of the present invention to detect a sentinel lymph node will now be described. When a fluorescent dye-conjugated sentinel lymph node marker is injected into a tumor tissue site, the fluorescent dye-conjugated sentinel lymph node marker (hereinafter referred to as a fluorescence marker ) flows into a lymphatic vessel and moves along the lymphatic vessel to a lymph node, and then the movement speed thereof in the lymph node becomes slower or it is adsorbed onto the lymph node. The lymph node that the fluorescence marker first meets is the sentinel lymph node. This lymph node contains a large amount of the fluorescence marker and is distributed mainly in the armpit in the case of breast cancer. It is distributed mainly in the adipose tissue (subcutaneous soft tissue) in the armpit. Because the sentinel lymph node is located below the soft tissue at a depth of a few cm in the case of the human body, light for obtaining fluorescence is inaccessible to the sentinel lymph node, because the soft tissue absorbs the light. Thus, if light that is relatively less absorbed is used and strong light such as a laser is irradiated or if incidence of the surrounding light is prevented, the sentinel lymph node will be weakly visible as an image by a fluorescence camera.

However, in this method, the process of obtaining an image is complicated, it is inconvenient to examine the image while performing a surgical operation, and also sensitivity is low for the above-described reasons. A radioactive isotope lymph node marker which is currently widely used makes it possible to obtain an image with a camera, but in actual applications, a radioactive probe is used to find a lymph node, and this method has been clinically established for the diagnosis of patients.

Thus, in the case of the fluorescence marker, it is believed that the contact-type method that uses the fluorescence-sensing probe 100 of the present invention is clinically more convenient and has high sensitivity compared to the method that uses the fluorescence camera, and thus it is proposed as a solution for the above-described problems.

Also, the fluorescence-sensing probe 100 may be widely used not only in sentinel lymph node surgery, but also in other types of surgery and experimentation, which use a fluorescent marker. Typically, it may be widely used to examine the distribution of various fluorescence markers that bind to various biological targets. For example, if it is required to inject a fluorescent marker that highly binds to a specific cancer and to find and dissect the specific cancer in which the fluorescent marker is distributed, the cancer can be dissected by surgery while more accurately examining the distribution of the cancer with the fluorescent probe in proximity to the cancer site. Also, in cases such as very small tumors that are difficult to visually examine during surgery, the fine tumor site can be labeled with a fluorescent substance prior to surgery and examined with the fluorescence sensing probe during surgery, and fluorescence in the lesion dissected by surgery can be measured to confirm whether the site to be removed was dissected well. As described above, the fluorescence-sensing probe can be widely used in various applications.

As described above, according to the present invention, a substance, which has been bound to or contains therein a fluorescent dye that is distributed in a human or animal body, can be detected at high sensitivity in proximity to tissue. Thus, sentinel lymph node dissection and target-guided surgery can be more effectively performed using fluorescence detection. In addition, the present invention may be applied to various fields, including endoscopy, laparoscopy and robot-assisted surgery.

Although the preferred embodiments of the present invention have been described for illustrative purposes, those skilled in the art will appreciate that various modifications, additions and substitutions are possible, without departing from the scope and spirit of the invention as disclosed in the accompanying claims.

Claims

A fluorescence-sensing probe comprising:

a light source that emits light of specific wavelength;

an excitation filter that selectively transmits, from among the light emitted from the light source, only excitation light exciting a fluorescent substance included in vivo;

an emission filter that selectively transmits only fluorescence emitted from the fluorescent substance; and

a light-sensing element capable of detecting fluorescence that has passed through the emission filter.
The fluorescence-sensing probe of claim 1, wherein the excitation filter is connected with one end of a first optical fiber, and the emission filter is connected with one end of a second optical fiber, in which the other ends of the first and second optical fibers are received in the same housing.
The fluorescence-sensing probe of claim 1, wherein the excitation filter and the emission filter are connected with one end of an optical fiber, and the other end of the optical fiber is received in the same housing.
The fluorescence-sensing probe of any one of claims 1 to 3, wherein the housing serves to come in contact with the surface of a living body to excite a fluorescent substance included in the living body and to detect fluorescence emitted from the fluorescent substance.
The fluorescence-sensing probe of any one of claims 1 to 3, wherein the light source is any one or more of a laser, a laser diode, an LED, a halogen lamp, and a xenon lamp.
The fluorescence-sensing probe of any one of claims 1 to 3, wherein the light-sensing element is any one or more of CCD, CMOS, and a photodiode.
The fluorescence-sensing probe of any one of claims 1 to 3, wherein the fluorescence-sensing probe further comprises an amplifier capable of amplifying the detected fluorescence.
The fluorescence-sensing probe of any one of claims 1 to 3, wherein the fluorescence-sensing probe further comprises a display unit capable of displaying the detected fluorescence.
The fluorescence-sensing probe of any one of claim 2 or 3, wherein the fluorescence-sensing probe further comprises a display unit capable of displaying the detected fluorescence, in which the display unit is provided on the housing.
The fluorescence-sensing probe of claim 1, wherein the excitation filter and the emission filter are provided in plurality or configured such that they can be selectively replaced.
A fluorescence-sensing probe comprising:

a light source that emits light of the wavelength that excites the fluorescent substance included in vivo;

an emission filter that selectively transmits only fluorescence emitted from the fluorescent substance; and

a light-sensing element capable of detecting fluorescence that has passed through the emission filter.
A fluorescence detection method comprising the steps of:

(a) injecting into a subject a substance which has been bound thereto or contains therein a fluorescent dye; and

(b) detecting fluorescence in the subject using the fluorescence-sensing probe of any one of claims 1 to 11.
A sentinel lymph node dissection method comprising the steps of:

(a) injecting into a living body a substance which has been bound thereto or contains therein a fluorescent dye;

(b) placing the fluorescence-sensing probe of any one of claims 1 to 11 in proximity to the living body to detect a sentinel lymph node present in the living body; and

(c) dissecting the detected sentinel lymph node.
A target-guided surgery method comprising the steps of:

(a) injecting into a living body a substance which has been bound thereto or contains therein a fluorescent dye and is selectively accumulated in a specific lesion;

(b) placing the fluorescence-sensing probe of any one of claims 1 to 10 in proximity to the living body to confirm the location of the specific lesion present in the living body; and

(c) dissecting the confirmed specific lesion.