CN109813687A - A kind of imaging method of up-conversion fluorescence imaging system and up-conversion fluorescence - Google Patents
A kind of imaging method of up-conversion fluorescence imaging system and up-conversion fluorescence Download PDFInfo
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Abstract
The invention discloses a kind of up-conversion fluorescence imaging systems, the wave-length coverage of the incident light of the light source transmitting of the system is 1000nm to 2500nm, the wave-length coverage is in the area near-infrared II and the area near-infrared III, and the penetration depth of the infrared light of the wave band in the sample is deep；By the incident light excitation in the area near-infrared II or the area near-infrared III, the upper conversion fluorescent nano particle being marked in the sample in advance can issue the emergent light near infrared region, and the penetration depth of the emergent light in the sample is equally relatively deep；And since the wavelength of the incident light of light source sending includes the area near-infrared II and the area near-infrared III, which is very unlikely to cause the autofluorescence phenomenon of sample, so as to effectively increase fluorescence imaging depth and sensitivity；The invention also discloses a kind of imaging methods of up-conversion fluorescence, equally have above-mentioned beneficial effect.
Description
Technical field
The present invention relates to fluorescence imaging field, more particularly to a kind of up-conversion fluorescence imaging system and up-conversion fluorescence
Imaging method.
Background technique
With the continuous progress of science and technology in recent years, Imaging-PAM is applied increasingly wider in people's daily life
It is general.
Imaging-PAM is a series of with unionized low-energy radiation, non-intruding, hypersensitivity, high sensitivity etc. due to it
Advantage is widely used in field of biomedicine.Fluorescence can be real to the progress such as biomolecule, cell, tissue, organ simultaneously
Visible is applied, is usually used to carries out early diagnosis of tumor, external biological label analyte detection, drug in vivo at this stage
Metabolic pathway and distribution situation etc..
Among a variety of fluorescent materials, upper conversion fluorescent nano particle (Upconversion Nanoparticls, referred to as:
UCNPs luminescence mechanism) is to issue the photon of a high-energy by absorbing the photon of two or more low energy, i.e.,
Upper conversion fluorescent nano particle can issue the shorter light of wavelength by the long light of absorbing wavelength.It is glimmering compared to other
Luminescent material, upper conversion fluorescent nano particle have Stokes displacement big, and imaging signal to noise ratio is high, and emission spectrum is narrow, fluorescence lifetime is long
The features such as.
In the prior art, 808nm or 980nm laser excitation is usually used, acquisition 540nm, 660nm or 800nm's
Fluorescence signal realizes up-conversion fluorescence imaging.But in the prior art, using the upper conversion of 808nm or 980nm laser excitation
The fluorescence imaging depth of fluorescence imaging is usually lower.
Summary of the invention
The object of the present invention is to provide a kind of up-conversion fluorescence imaging systems, can effectively increase fluorescence imaging depth；This
The another object of invention is to provide a kind of imaging method of up-conversion fluorescence, can effectively increase fluorescence imaging depth.
In order to solve the above technical problems, the present invention provides a kind of up-conversion fluorescence imaging system, the up-conversion fluorescence at
As system includes light source, sample stage, optical signal receiver and processor；
The wave-length coverage of the incident light of the light source transmitting are as follows: 1000nm to 2500nm, including endpoint value；
The sample stage is used to be placed through the sample of upper conversion fluorescent nano particle label, and the sample is for absorbing institute
It states incident light and launches and penetrate light；
The optical signal receiver connects the processor, and the optical signal receiver is used to receive the emergent light, and
The emergent light is converted into image data, to be sent to the processor；
The processor is used to generate fluorescent image according to described image data.
Optionally, the upper conversion fluorescent nano particle is AReF4(Re3+, n%) and@mAReF4Or Re2O3(Re3+, n%)；
Wherein, A includes Li, Na, K；Re includes Sc, Y, La, Gd, Lu；Re3+Including Pr3+、Nd3+、Sm3+、Dy3+、Ho3+、Er3+、Tm3+；
The value range of the n are as follows: 0 to 100, including right end point value；The value range of the m are as follows: 0 to 30, including endpoint value.
Optionally, the light source is laser light source, is provided with laser beam between the laser light source and the sample stage
Adjust lens group.
Optionally, the laser beam adjust lens group include the collimation lens being sequentially distributed along optical axis, extender lens and
Shaping lens.
Optionally, bandpass filter, the bandpass filter are provided between the sample stage and the optical signal receiver
Piece is for choosing the monochromatic emergent light for presetting wavelength.
Optionally, it is provided with fluorescent light beam between the bandpass filter and the optical signal receiver and adjusts lens group.
Optionally, the fluorescent light beam adjust lens group include the collimation lens being sequentially distributed along optical axis, extender lens and
Shaping lens.
Optionally, the optical signal receiver is CCD detector.
The present invention also provides a kind of imaging methods of up-conversion fluorescence, which comprises
The sample of sample stage is arranged in light source transmitting incident light irradiation, wherein the wave-length coverage of the incident light are as follows:
1000nm to 2500nm, including endpoint value, the sample are the sample marked by upper conversion fluorescent nano particle；
The sample absorbs the incident light, and launches and penetrate light；
Optical signal receiver receives the emergent light, and the emergent light is converted into image data, to be sent to processing
Device；
The processor generates fluorescent image according to described image data.
Optionally, the upper conversion fluorescent nano particle is AReF4(Re3+, n%) and@mAReF4Or Re2O3(Re3+, n%)；
Wherein, A includes Li, Na, K；Re includes Sc, Y, La, Gd, Lu；Re3+Including Pr3+、Nd3+、Sm3+、Dy3+、Ho3+、Er3+、Tm3+；
The value range of the n are as follows: 0 to 100, including right end point value；The value range of the m are as follows: 0 to 30, including endpoint value.
A kind of up-conversion fluorescence imaging system provided by the present invention, the wavelength model of the incident light of the light source transmitting of the system
It encloses for 1000nm to 2500nm, which is in the area near-infrared II and the area near-infrared III, and the infrared light of the wave band is in sample
In penetration depth it is deep；By the incident light excitation in the area near-infrared II or the area near-infrared III, in advance in the sample into
The upper conversion fluorescent nano particle of line flag can issue the emergent light near infrared region, the emergent light penetrating in the sample
Depth is equally relatively deep；And since the wavelength of the incident light of light source sending includes the area near-infrared II and the area near-infrared III, the incidence
Optrode is not easy to cause the autofluorescence phenomenon of sample, so as to effectively increase fluorescence imaging depth and sensitivity.
The present invention also provides a kind of imaging methods of up-conversion fluorescence, equally have above-mentioned beneficial effect, herein no longer
It is repeated.
Detailed description of the invention
It, below will be to embodiment or existing for the clearer technical solution for illustrating the embodiment of the present invention or the prior art
Attached drawing needed in technical description is briefly described, it should be apparent that, the accompanying drawings in the following description is only this hair
Bright some embodiments for those of ordinary skill in the art without creative efforts, can be with root
Other attached drawings are obtained according to these attached drawings.
Fig. 1 is a kind of structural schematic diagram of up-conversion fluorescence imaging system provided by the embodiment of the present invention；
Fig. 2 is the structural schematic diagram of another kind up-conversion fluorescence imaging system provided by the embodiment of the present invention；
Fig. 3 is a kind of flow chart of the imaging method of up-conversion fluorescence provided by the embodiment of the present invention.
In figure: 10. light sources, 20. laser beams adjusting lens group, 21. collimation lenses, 22. extender lenses, 23. shapings are saturating
Mirror, 30. sample stages, 31. samples, 32. biopsy samples, 40. bandpass filters, 50. fluorescent light beams adjust lens group, 60. fluorescence
Signal acquisition object lens, 70. optical signal receivers, 80. processors, 90. reflecting mirrors, 91. camera bellows, 92. automatic lifting control units.
Specific embodiment
Core of the invention is to provide a kind of up-conversion fluorescence imaging system.In the prior art, 808nm is usually used
Or 980nm laser excitation, the fluorescence signal of acquisition 540nm, 660nm or 800nm realize up-conversion fluorescence imaging.Due to existing
The wave-length coverage of incident light that light source issues in technology is in the area near-infrared I, and wavelength is relatively short, in the sample penetrate depth
Degree is than shallower；Simultaneously by being excited by the incident light in the area near-infrared I, the emergent light that upper conversion fluorescent nano particle issues usually exists
Visible-range, the outgoing light wavelength is equally relatively short, and penetration depth in the sample is than shallower；Simultaneously because in existing skill
The wavelength for the incident light that light source issues in art is in the area near-infrared I, may cause the autofluorescence phenomenon of sample, to cause
Fluorescence imaging depth is shallower.
And a kind of up-conversion fluorescence imaging system provided by the present invention, the wavelength of the incident light of the light source transmitting of the system
Range is 1000nm to 2500nm, which is in the area near-infrared II and the area near-infrared III, and the infrared light of the wave band is in sample
Penetration depth in product is deep；By the incident light excitation in the area near-infrared II or the area near-infrared III, in advance in the sample
The upper conversion fluorescent nano particle being marked can issue the emergent light near infrared region, the emergent light wearing in the sample
Saturating depth is equally relatively deep；And since the wavelength of the incident light of light source sending includes the area near-infrared II and the area near-infrared III, this enters
It penetrates optrode to be not easy to cause the autofluorescence phenomenon of sample, so as to effectively increase fluorescence imaging depth and sensitivity.
In order to enable those skilled in the art to better understand the solution of the present invention, with reference to the accompanying drawings and detailed description
The present invention is described in further detail.Obviously, described embodiments are only a part of the embodiments of the present invention, rather than
Whole embodiments.Based on the embodiments of the present invention, those of ordinary skill in the art are not making creative work premise
Under every other embodiment obtained, shall fall within the protection scope of the present invention.
Referring to FIG. 1, Fig. 1 is a kind of structural representation of up-conversion fluorescence imaging system provided by the embodiment of the present invention
Figure.
Referring to Fig. 1, in embodiments of the present invention, the up-conversion fluorescence imaging system includes light source 10, sample stage 30, light
Signal receiver 70 and processor 80.
The wave-length coverage for the incident light that above-mentioned light source 10 emits are as follows: 1000nm to 2500nm, including endpoint value, i.e. light source 10
The wavelength of the incident light of sending includes 1000nm to 2500nm, while including 1000nm and 2500nm.Above-mentioned sample stage 30 is used for
It is placed through the sample 31 of upper conversion fluorescent nano particle label, the sample 31 is penetrated for absorbing the incident light and launching
Light.
In embodiments of the present invention, it is added to upper conversion fluorescent nano particle in the sample 31 placed on sample stage 30,
The upper conversion fluorescent nano particle is AReF4(Re3+, n%) and@mAReF4Or Re2O3(Re3+, n%)；Wherein, A include Li, Na,
K；Re includes Sc, Y, La, Gd, Lu；Re3+Including Pr3+、Nd3+、Sm3+、Dy3+、Ho3+、Er3+、Tm3+；The value range of the n
Are as follows: 0 to 100, including right end point value；The value range of the m are as follows: 0 to 30, including endpoint value.In above-mentioned AReF4(Re3+,
N%)@mAReF4In ,@indicates that core-shell structure, i.e., the described upper conversion fluorescent nano particle are clad structure.Above-mentioned m indicates AReF4
Shell number, if m value is 0, for each upper conversion fluorescent nano particle, only one layer of AReF from inside to outside4；If m value
It is 5, then for each upper conversion fluorescent nano particle, coats the AReF that is of five storeys altogether from inside to outside4, i.e., have altogether from inside to outside
There are 6 layers of AReF4.In embodiments of the present invention, the AReF4(Re3+, n%) and@mAReF4Shell number can be from 0 to 30 appoint
Meaning number, including 0 and 30.
Above-mentioned AReF4(Re3+, n%) and@mAReF4In, A represents alkali metal, including tri- kinds of elements of Li, Na, K；Above-mentioned AReF4
(Re3+, n%) and@mAReF4With Re2O3(Re3+, n%) in, Re is to be suitble to do rare earth in the 5 of upper conversion fluorescent nano particle matrix
Element, respectively Sc, Y, La, Gd, Lu.Other rare earth elements are not appropriate for the base for doing upper conversion fluorescent nano particle
Matter.Above-mentioned Re3+For the rare earth ion added in the matrix of upper conversion fluorescent nano particle, add in embodiments of the present invention
Rare earth ion is usually to be present in the matrix in the form of+trivalent.Above-mentioned Re3+Including Pr3+、Nd3+、Sm3+、Dy3+、Ho3 +、Er3+、Tm3+；Above-mentioned AReF4With Re2O3The matrix of respectively two kinds upper conversion fluorescent nano particles.For adulterating variety classes
Rare earth ion, upper conversion fluorescent nano particle has different absorption peaks, also has different emission peaks accordingly；On simultaneously
The matrix of conversion fluorescent nano particle also will affect its absorption peak and emission peak.For example, if AReF4(Re3+, n%) and@mAReF4In
The Re of doping3+For Tm3+, then the absorption peak of entire upper conversion fluorescent nano particle is 1630nm, corresponding up-conversion fluorescence is received
The emission peak of rice grain includes 465nm, 684nm, 785nm, 1190nm and 1800nm；If Re2O3(Re3+, n%) in adulterate Re3 +For Tm3+The absorption peak of so entire upper conversion fluorescent nano particle includes 800nm and 1200nm, and corresponding up-conversion fluorescence is received
The emission peak of rice grain is 1800nm.Since in embodiments of the present invention, upper conversion fluorescent nano particle can adulterate a variety of Re3 +, but a kind of Re is usually only adulterated for each upper conversion fluorescent nano particle3+.It certainly, in use can be simultaneously
Using a variety of doped with different Re3+Upper conversion fluorescent nano particle, can also simultaneously use a variety of matrix up-conversion fluorescence
Nano particle.Above-mentioned a variety of Re3+Corresponding absorption peak is distributed in the area near-infrared II and the near-infrared III of 1000nm to 2500nm
Area.Certainly, for a certain Re of addition3+For, it will usually there are multiple emission peaks.For example, if AReF4(Re3+, n%) and@
mAReF4The Re of middle doping3+For Dy3+, then the absorption peak of entire upper conversion fluorescent nano particle is 1680nm, corresponding upper turn
The emission peak for changing fluorescent nano particle includes 483nm, 582nm, 910nm, 1080nm and 1290nm, and above-mentioned emission peak includes from can
It is light-exposed until the biggish range of this span of near infrared light, but due to the characteristic of upper conversion fluorescent nano particle, to Mr. Yu
A kind of Re3+For, the wavelength of absorption peak is all larger than the wavelength of emission peak.Due to the Re of doping3+It, can with multiple emission peaks
To realize that multicolor fluorescence is imaged, detailed content will be described in detail in subsequent paragraph.
Certainly, for any one Re3+Absorption peak and emission peak for, all have corresponding peak width and half-peak breadth, institute
It is only wavelength corresponding to the peak value of absorption peak or emission peak with specific value used in above-mentioned illustration, in practical feelings
The wavelength of the emergent light of the wavelength and transmitting for the incident light that upper conversion fluorescent nano particle can be absorbed in condition is a range.
In embodiments of the present invention, the rare earth ion of doping is not restricted to above-mentioned 7 kinds of rare earth ions, simultaneously for the rare earth of doping
The valence state of ion is also not limited only to+trivalent, as long as the absorption peak in the rare earth ion of doping is in 1000nm between 2500nm
, the valence state for the rare earth ion of doping and ionic species and it is not specifically limited in embodiments of the present invention.
In above-mentioned expression formula, n represents doping with rare-earth ions Re3+Concentration.In embodiments of the present invention, rare earth ion
Doping concentration can from 0 to 100, including right end point value, the i.e. doping concentration of rare earth ion include 100%.For adulterating not
With the upper conversion fluorescent nano particle of rare earth ion, the doping concentration of rare earth ion only influences whether up-conversion fluorescence nanometer
The intensity for the transmitting light that grain issues, will not influence the position of absorption peak.
In embodiments of the present invention, the range of the absorption peak of the upper conversion fluorescent nano particle due to label in sample 31
It is the wave-length coverage for the incident light that corresponding above-mentioned light source 10 emits from 1000nm to 2500nm are as follows: 1000nm to 2500nm, packet
Include endpoint value.Under normal conditions, since laser has the features such as good monochromaticjty, directionality and brightness are high, in the present invention
It is the usually laser light source of light source 10 used in example.Since the monochromaticjty of the incident light of laser light source sending is stronger, wave crest
It is relatively narrow, wavelength and the above-mentioned upper conversion fluorescent nano particle of the incident light for needing laser light source to issue in the specific use process
Absorption peak is corresponding, if such as the absorption peak of the glimmering nano particle of upper conversion be located at 2000nm, then needing accordingly issue wave
The laser light source of the incident light of a length of 2000nm.At this stage, according to the rare earth ion adulterated in upper conversion fluorescent nano particle
Type it is different, can to select the wavelength of transmitting incident light be 1050nm, 1150nm, 1250nm, 1500nm, 1600nm,
The semiconductor laser of 1700nm or 1950nm is as light source 10.It certainly, is in example for the light source 10 in the present invention
The wavelength of the incident light of transmitting is simultaneously not specifically limited, as long as being located at 1000nm between 2500nm.
Due to the usual very little of irradiated area for the incident light that laser light source issues, it can be provided with laser beam under normal conditions
Lens group 20 is adjusted to adjust the shape of incident light, incident light is uniformly irradiated on sample 31.In general, described
The incident light that laser light source issues is usually to pass through optical fiber to be transferred to laser beam adjusting lens group 20, and the laser beam is adjusted
Lens group 20 includes collimation lens 21, extender lens 22 and shaping lens 23, these three lens are successively distributed along optical axis.That is institute
The incident light for stating the sending of light source 10 first passes through collimation lens 21 and carrys out collimated incident light, using extender lens 22 come to incident light into
Row expands, and finally the shape of incident light is adjusted by shaping lens 23.Certainly, the laser beam adjusts lens group
20 structure is not limited to above-mentioned three kinds of lens, can also include other lens, while being also not necessarily limited to have to using upper
Three kinds of lens are stated, above-mentioned three kinds of lens are also possible to other distribution modes along the distribution of optical axis.The related laser beam tune
The specific structure of section lens group 20 is not specifically limited in embodiments of the present invention.
In embodiments of the present invention, light source 10 can issue the sample 31 that sample stage 30 is arranged in incident light irradiation, the sample
Product 31 are the sample 31 marked by upper conversion fluorescent nano particle；The incident light can be absorbed in the sample 31, and emits
Emergent light, the emergent light, that is, fluorescence.It concretely, is that the upper conversion fluorescent nano particle added in sample 31 absorbs incidence
Light, and emit fluorescence.In embodiments of the present invention, due to integrally-built limitation, it can be provided with reflecting mirror 90 under normal conditions,
The incident light emitting that light source 10 is issued is to sample 31.Specific limit is not done simultaneously for the specific structure of the sample stage 30
It is fixed.The sample stage 30 is in addition to for that can also have function of auto-lift to adjust sample stage 30 other than placing sample 31
Highly；Also it can have temperature incubation function, to guarantee when sample 31 is biological sample, the activity of the biological sample.Certainly, institute
Other functions can also be had by stating sample stage 30, will not enumerate herein.
In embodiments of the present invention, the up-conversion fluorescence imaging system further includes having optical signal receiver 70 and processor
80, the optical signal receiver 70 connects the processor 80.The optical signal receiver 70 is used to receive the emergent light, and
The emergent light is converted into image data, to be sent to the processor 80.
After sample 31 emits emergent light, the optical signal receiver 70 can receive the emergent light, and go out by described in
It penetrates light and is converted into image data, to be sent to the processor 80.The effect of the i.e. described optical signal receiver 70 is will be received
Optical signal is converted into electric signal, is handled with the fluorescence signal that convenient processor 80 issues sample 31.Due in the present invention
In embodiment, the emergent light that the sample 31 added with upper conversion fluorescent nano particle issues generally includes the monochrome there are many wavelength
Emergent light, but since the wavelength of each monochromatic emergent light is different, penetration depth is also not quite similar in sample 31 accordingly.It is logical
In normal situation, above-mentioned optical signal receiver 70 only receives a kind of monochromatic emergent light, and the monochromatic emergent light that will be received every time
It is converted into corresponding image data and is sent to processor 80.Correspondingly, needing in the sample stage 30 and optical signal receiver
Bandpass filter 40 is provided between 70, the bandpass filter 40 is for choosing the monochromatic emergent light for presetting wavelength.
Bandpass filter 40 is one kind of optical filter, can isolate a certain wave band monochromatic light.The bandpass filter 40
Allow by the wave-length coverage of monochromatic emergent light need with the peak position of the emission peak of above-mentioned upper conversion fluorescent nano particle and partly
Peak width is corresponding.In embodiments of the present invention, it can be set there are many bandpass filter 40 and successively choose the list of different wave length
Color emits light, and the corresponding optical signal receiver 70 can also be sequentially received the monochromatic emission light of different wave length, and right
Corresponding image data can be generated in each monochromatic emission light, finally by multiple images data be sent to processor 80 into
Row processing.Since penetration depth depth of the monochromatic emergent light in sample 31 of different wave length is different, multiple bandpass filters are used
Piece 40 after allowing optical signal receiver 70 successively to obtain the monochromatic emergent light of different wave length, facilitates processor 80 described in the later period raw
At the multicolor fluorescence image with different imaging depths.
Certainly, the optical signal receiver 70 can receive a variety of monochromatic emergent lights simultaneously and be converted into image data, but
It is that this is unfavorable for post processor 80 and handles image data, while also influencing whether the fluorescent image ultimately generated
Effect.
Under normal conditions, it is provided with fluorescent light beam between bandpass filter 40 and optical signal receiver 70 and adjusts lens group
50, to adjust the shape of emergent light, so that optical signal receiver 70 may finally obtain clearly image, to realize for outgoing
The acquisition of light.Similar with laser beam adjusting lens group 20, it generally also includes along optical axis that the fluorescent light beam, which adjusts lens group 50,
Collimation lens, extender lens and the shaping lens being sequentially distributed.Detailed content please refers to above-mentioned related laser beam and adjusts lens
The specific introduction of group 20, is no longer repeated herein.The specific structure for adjusting lens group 50 in relation to the fluorescent light beam exists
It is equally not specifically limited in the embodiment of the present invention, as long as being adjustable the shape of the emergent light.
Between above-mentioned fluorescent light beam lens group and optical signal receiver 70, it usually can also be provided with fluorescence signal acquisition object
Mirror 60, for collecting above-mentioned emergent light.In embodiments of the present invention, the operating wavelength range of fluorescence signal acquisition object lens 60 is usual
In 200nm between 2500nm.According to the needs of practical application, multiple and different amplification factors and digit punch can be specifically set
The fluorescence signal acquisition object lens 60 of diameter, in relation to the specific amplification factor of fluorescence signal acquisition object lens 60 and digital aperture numerical value at this
It is not specifically limited in inventive embodiments.
In embodiments of the present invention, the optical signal receiver 70 is usually CCD (Charge-coupled Device) inspection
Survey device.Since the emission peak of above-mentioned upper conversion fluorescent nano particle is usually in ultraviolet light wave band, visible light wave range and near-infrared wave
Section is all distributed, so usually requiring to include at least in the CCD detector there are two types of fluorescent probe, one is be used to detect
The focal plane array detector of ultraviolet light and visible light wave range fluorescence signal, another kind are for detecting near infrared light wave band fluorescence
The focal plane array detector of signal.CCD detector, which may be implemented, by above two focal plane array detector can receive
From ultraviolet light wave band, visible light wave range is arrived, then arrives the fluorescence signal of infrared band, and the fluorescence signal is converted into picture number
According to.The fluorescence signal corresponds to emergent light above-mentioned in the embodiment of the present invention.
In embodiments of the present invention, optical signal receiver 70 can receive above-mentioned emergent light, and the emergent light is converted
At image data, to be sent to processor 80；The processor 80 generates fluorescent image according to described image data.
Above-mentioned processor 80 is mainly used for being calculated to the image data received, analyzing, handle and rebuild etc., with
Finally fluorescent image is generated according to described image data.In embodiments of the present invention, above-mentioned sample stage 30 can further be set
It is equipped with stepper motor, the processor 80 can connect stepper motor, and the processor 80 can be by automatically controlling stepping electricity
Machine works to realize the lifting of sample stage 30, is imaged in high precision with this to realize.The processor 80 usually can also be real simultaneously
The now focusing in above-mentioned imaging process, the switching of bandpass filter 40, the selection of sample imaging position etc..Related processor 80 has
The function that body may be implemented, is not specifically limited in embodiments of the present invention, as long as fluorescence can be generated according to described image data
Image.
In embodiments of the present invention, visible light source can be further added in up-conversion fluorescence imaging system, be used for
Realize light field imaging.Visible light is emitted to irradiate the sample 31 by visible light source, the face of sample 31 is observed with this
Color, shape etc. appearance information.Under normal conditions, select sodium lamp as visible light source.It is of course also possible to select others
Device does visible light source, is not specifically limited in embodiments of the present invention.
A kind of up-conversion fluorescence imaging system provided by the present invention, the wavelength for the incident light that the light source 10 of the system emits
Range is 1000nm to 2500nm, which is in the area near-infrared II and the area near-infrared III, and the infrared light of the wave band is in sample
Penetration depth in product 31 is deep；By the incident light excitation in the area near-infrared II or the area near-infrared III, in advance in sample
The upper conversion fluorescent nano particle being marked in 31 can issue the emergent light near infrared region, and the emergent light is in sample 31
In penetration depth it is equally relatively deep；And since the wavelength of the incident light of the sending of light source 10 includes the area near-infrared II and near-infrared
The area III, the incident light are very unlikely to cause the autofluorescence phenomenon of sample 31, so as to effectively increase fluorescence imaging depth
Degree.
In the present invention, when sample 31 is biopsy samples, it usually needs biopsy samples are put into camera bellows, are needed simultaneously
The more uniform irradiation biopsy samples of the incident light that light source 10 issues.Detailed content please refers to following inventive embodiments.
Referring to FIG. 2, Fig. 2 is the structural representation of another kind up-conversion fluorescence imaging system provided by the embodiment of the present invention
Figure.
Referring to fig. 2, up-conversion fluorescence imaging system provided by the embodiment of the present invention is provided with foregoing invention embodiment
Up-conversion fluorescence imaging system be substantially the same, difference place is in the embodiment of the present invention entire up-conversion fluorescence imaging system
Most of component is in camera bellows 91, and the light source 10 can issue multichannel incident light to irradiate biopsy samples 32.
In embodiments of the present invention, it is provided with multiple laser beams in the camera bellows 91 and adjusts lens group 20, it is multiple described
Laser beam adjusts lens group 20 and is distributed generally uniformly in the inner wall of the camera bellows 91.The incident light meeting that the light source 10 issues
Be transmitted separately to multiple laser beams by multiple optical paths and adjust lens groups 20, under normal conditions the incident light be by mostly with
Optical fiber is transmitted to multiple laser beams and adjusts lens group 20.The laser beam adjusts lens group 20 in the shape to incident light
, can be with direct irradiation on biopsy samples 32 after shape is adjusted, or biopsy samples 32 are radiated at by reflecting mirror 90
On.In embodiments of the present invention, 32 region of biopsy samples is imaging region, and the embodiment of the present invention will be realized described
Incident light uniform irradiation will guarantee that the light intensity deviation of difference in imaging region is less than in the imaging region under normal conditions
5.0%.
The sample stage 30 for placing the biopsy samples 32 usually requires have temperature incubation function, i.e., the described sample stage 30 is provided with
Heating and constant temperature component, the vital sign to guarantee the biopsy samples 32 in imaging process are normal.The following table of the sample stage 30
Face is usually connected with automatic lifting control unit 92, and the processor 80 usually connects the automatic lifting control unit 92, place
Reason device 80 can adjust the height of sample stage 30 by automatic lifting control unit 92, be focused with this to realize, it is ensured that final
The clarity of the fluorescent image of formation.
In embodiments of the present invention, the upper conversion fluorescent nano particle marked in biopsy samples 32 is usually AReF4(Tm3+,
N%)@mAReF4, i.e., the rare earth ion adulterated in upper conversion fluorescent nano particle is Tm3+.Due to AReF4(Tm3+, n%) and@
mAReF4Absorption peak be located at 1630nm, emission peak includes 465nm, 684nm, 785nm, 1190nm and 1800nm, absorption peak with
The wave band of emission peak includes the area near-infrared II and the area near-infrared III, and the light wave of the two wave bands all has in biopsy samples 32
Good penetration depth uses AReF accordingly4(Tm3+, n%) and@mAReF4Carrying out fluorescence imaging can greatly increase finally
The imaging depth of the fluorescent image of generation.Certainly, it can also be adulterated in the upper conversion fluorescent nano particle other kinds of dilute
Native ion, equally may be implemented absorption peak and emission peak is near infrared region simultaneously.The related upper conversion fluorescent nano particle
The type of middle doping with rare-earth ions in embodiments of the present invention and is not specifically limited.
If using AReF4(Tm3+, n%) and@mAReF4As upper conversion fluorescent nano particle, the corresponding light source 10 is excellent
Be selected as can issuing the laser light source that wavelength is 1630nm, the subsequent bandpass filter 40 used be preferably allow by light wave
Wavelength is 1190nm.
After the biopsy samples 32 issue emergent light, up-conversion fluorescence imaging system provided by the embodiment of the present invention
Preset monochromatic emergent light usually can be also chosen by corresponding bandpass filter 40, and is adjusted thoroughly by fluorescent light beam
The shape of 50 pairs of microscope group monochromatic emergent lights is adjusted, and is transmitted to monochromatic emergent light eventually by fluorescence signal acquisition object lens 60
Optical signal receiver 70.The optical signal receiver 70 can will scheme after the monochromatic emergent light is converted into image data
As data are sent to processor 80, finally there is processor 80 to generate fluorescent image according to described image data.
Remaining content of up-conversion fluorescence imaging system described in the embodiment of the present invention is in foregoing invention embodiment
In be described in detail, concrete condition please refers to foregoing invention embodiment, is no longer repeated herein.
A kind of up-conversion fluorescence imaging system provided by the present invention can adjust lens group 20 by multiple laser beams
Come realize the incident light uniform irradiation for issuing light source 10 on biopsy samples 32, the matter for the fluorescent image that Lai Tigao is ultimately generated
Amount.And sample stage 30 has heating and constant temperature component, it is ensured that the vital sign of biopsy samples 32 is being just in imaging process
Often.
Up-conversion fluorescence imaging system provided by the present invention can further be applied in fluorescence operation guiding system,
Detailed content please refers to following inventive embodiments.
Upper conversion provided by up-conversion fluorescence imaging system provided by the embodiment of the present invention and foregoing invention embodiment
Fluoroscopic imaging systems are substantially the same, and difference place is: in embodiments of the present invention, the incident light that light source 10 issues is through too drastic
After light light beam regulation lens group 20, irradiated area can be formed not less than 200cm2Incident light irradiate the focal zone of patient
Domain.
Before the surgery, patient can inject the AReF that surface modification has target molecule4(Er3+, n%) and@mAReF4, i.e., upper conversion
The rare earth ion adulterated in fluorescent nano particle is Er3+, simultaneously because upper conversion fluorescent nano particle surface modification has target molecule,
The upper conversion fluorescent nano particle selectively can largely be gathered in the pathological tissue of patient.Due to AReF4(Er3+,
N%)@mAReF4Absorption peak be located at 1500nm, emission peak respectively in 540nm, 660nm, 800nm, 980nm and 1730nm, by
In AReF4(Er3+, n%) and@mAReF4With multiple absorption peaks, multicolor fluorescence imaging may be implemented, use AReF accordingly4(Er3 +, n%) and@mAReF4Carry out the information for the entire pathological tissue of acquisition that multicolor fluorescence imaging can have levels.Certainly, described upper turn
Other kinds of rare earth ion can also be adulterated by changing in fluorescent nano particle, equally can have simultaneously multiple absorption peaks to realize
Multicolor fluorescence imaging.Type in relation to doping with rare-earth ions in the upper conversion fluorescent nano particle is in embodiments of the present invention simultaneously
It is not specifically limited.
If using AReF4(Er3+, n%) and@mAReF4As upper conversion fluorescent nano particle, the corresponding light source 10 is excellent
Be selected as can issuing the laser light source that wavelength is 1500nm, the subsequent multiple bandpass filters 40 used be preferably allow by
Optical wavelength is respectively 540nm, 660nm, 800nm, 980nm and 1730nm.
After optical signal receiver 70 gets multiple monochromatic emergent lights by multiple bandpass filters 40, it can be generated
Multiple images data, and image data is sent to processor 80.In embodiments of the present invention, three-dimensional is stored in processor 80
Algorithm for reconstructing can reconstruct the three of pathological tissue based on the monochromatic emergent light of the different wave length of the acquisition of optical signal receiver 70
Dimension boundary and volume instruct the controllable precise of surgical operation to implement, significantly improve hand to provide precise information for surgical operation
The quality of art treatment.
Remaining content of up-conversion fluorescence imaging system described in the embodiment of the present invention is real in above-mentioned two invention
It applies in example and is described in detail, concrete condition please refers to foregoing invention embodiment, is no longer repeated herein.
A kind of up-conversion fluorescence imaging system provided by the present invention, can on the basis of generating multicolor fluorescence image,
The three-dimensional boundaries and volume for reconstructing patient's pathological tissue instruct surgical operation to provide precise information for surgical operation
Controllable precise is implemented, and the quality of operative treatment is significantly improved.
A kind of imaging method of up-conversion fluorescence provided in an embodiment of the present invention is introduced below, it is described below at
Image space method can correspond to each other reference with above-described up-conversion fluorescence imaging system.
Fig. 3 is a kind of flow chart of the imaging method of up-conversion fluorescence provided by the embodiment of the present invention, referring to Fig. 3, institute
Stating imaging method may include:
S101: the sample of sample stage is arranged in light source transmitting incident light irradiation.
In embodiments of the present invention, the wave-length coverage of the incident light are as follows: 1000nm to 2500nm, including endpoint value, institute
Stating sample is the sample marked by upper conversion fluorescent nano particle.
S102: sample absorbs incident light, and launches and penetrate light.
S103: optical signal receiver receives emergent light, and emergent light is converted into image data, to be sent to processor.
S104: processor generates fluorescent image according to image data.
In embodiments of the present invention, the upper conversion fluorescent nano particle is specially AReF4(Re3+, n%) and@mAReF4Or
Re2O3(Re3+, n%)；Wherein, A includes Li, Na, K；Re includes Sc, Y, La, Gd, Lu；Re3+Including Pr3+、Nd3+、Sm3+、Dy3+、
Ho3+、Er3+、Tm3+；The value range of the n are as follows: 0 to 100, including right end point value；The value range of the m are as follows: 0 to 30, packet
Include endpoint value.
The imaging method of the up-conversion fluorescence of the present embodiment is used for using up-conversion fluorescence imaging system above-mentioned, therefore on
The embodiment part of the visible up-conversion fluorescence imaging system hereinbefore of specific embodiment in the imaging method of conversion fluorescence,
So specific embodiment is referred to the description of corresponding various pieces embodiment, details are not described herein.
Each embodiment in this specification is described in a progressive manner, the highlights of each of the examples are with it is other
The difference of embodiment, same or similar part may refer to each other between each embodiment.For being filled disclosed in embodiment
For setting, since it is corresponded to the methods disclosed in the examples, so being described relatively simple, related place is referring to method part
Explanation.
Professional further appreciates that, unit described in conjunction with the examples disclosed in the embodiments of the present disclosure
And algorithm steps, can be realized with electronic hardware, computer software, or a combination of the two, in order to clearly demonstrate hardware and
The interchangeability of software generally describes each exemplary composition and step according to function in the above description.These
Function is implemented in hardware or software actually, the specific application and design constraint depending on technical solution.Profession
Technical staff can use different methods to achieve the described function each specific application, but this realization is not answered
Think beyond the scope of this invention.
The step of method described in conjunction with the examples disclosed in this document or algorithm, can directly be held with hardware, processor
The combination of capable software module or the two is implemented.Software module can be placed in random access memory (RAM), memory, read-only deposit
Reservoir (ROM), electrically programmable ROM, electrically erasable ROM, register, hard disk, moveable magnetic disc, CD-ROM or technology
In any other form of storage medium well known in field.
The imaging method of a kind of up-conversion fluorescence imaging system provided by the present invention and up-conversion fluorescence is carried out above
It is discussed in detail.Used herein a specific example illustrates the principle and implementation of the invention, above embodiments
Explanation be merely used to help understand method and its core concept of the invention.It should be pointed out that for the common of the art
, without departing from the principle of the present invention, can be with several improvements and modifications are made to the present invention for technical staff, these
Improvement and modification are also fallen within the protection scope of the claims of the present invention.
Claims (10)
1. a kind of up-conversion fluorescence imaging system, which is characterized in that the up-conversion fluorescence imaging system includes light source, sample
Platform, optical signal receiver and processor；
The wave-length coverage of the incident light of the light source transmitting are as follows: 1000nm to 2500nm, including endpoint value；
The sample stage is used to be placed through the sample of upper conversion fluorescent nano particle label, the sample for absorb it is described enter
It penetrates light and launches and penetrate light；
The optical signal receiver connects the processor, and the optical signal receiver is for receiving the emergent light, and by institute
It states emergent light and is converted into image data, to be sent to the processor；
The processor is used to generate fluorescent image according to described image data.
2. system according to claim 1, which is characterized in that the upper conversion fluorescent nano particle is AReF4(Re3+,
N%)@mAReF4Or Re2O3(Re3+, n%)；Wherein, A includes Li, Na, K；Re includes Sc, Y, La, Gd, Lu；Re3+Including Pr3+、
Nd3+、Sm3+、Dy3+、Ho3+、Er3+、Tm3+；The value range of the n are as follows: 0 to 100, including right end point value；The value model of the m
It encloses are as follows: 0 to 30, including endpoint value.
3. system according to claim 2, which is characterized in that the light source is laser light source, the laser light source and institute
It states and is provided with laser beam adjusting lens group between sample stage.
4. system according to claim 3, which is characterized in that the laser beam adjust lens group include along optical axis successively
Collimation lens, extender lens and the shaping lens of distribution.
5. system according to claim 2, which is characterized in that be arranged between the sample stage and the optical signal receiver
There is bandpass filter, the bandpass filter is for choosing the monochromatic emergent light for presetting wavelength.
6. system according to claim 5, which is characterized in that between the bandpass filter and the optical signal receiver
It is provided with fluorescent light beam and adjusts lens group.
7. system according to claim 6, which is characterized in that the fluorescent light beam adjust lens group include along optical axis successively
Collimation lens, extender lens and the shaping lens of distribution.
8. system according to claim 2, which is characterized in that the optical signal receiver is CCD detector.
9. a kind of imaging method of up-conversion fluorescence, which is characterized in that the described method includes:
The sample of sample stage is arranged in light source transmitting incident light irradiation, wherein the wave-length coverage of the incident light are as follows: 1000nm is extremely
2500nm, including endpoint value, the sample are the sample marked by upper conversion fluorescent nano particle；
The sample absorbs the incident light, and launches and penetrate light；
Optical signal receiver receives the emergent light, and the emergent light is converted into image data, to be sent to processor；
The processor generates fluorescent image according to described image data.
10. according to the method described in claim 9, it is characterized in that, the upper conversion fluorescent nano particle is AReF4(Re3+,
N%)@mAReF4Or Re2O3(Re3+, n%)；Wherein, A includes Li, Na, K；Re includes Sc, Y, La, Gd, Lu；Re3+Including Pr3+、
Nd3+、Sm3+、Dy3+、Ho3+、Er3+、Tm3+；The value range of the n are as follows: 0 to 100, including right end point value；The value model of the m
It encloses are as follows: 0 to 30, including endpoint value.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN201394004Y (en) * | 2009-05-07 | 2010-02-03 | 复旦大学 | Up-conversion luminescence in-vivo rat imaging system of steady-state laser pumping |
| CN107286924A (en) * | 2017-06-13 | 2017-10-24 | 复旦大学 | Upper conversion nano crystalline material of the infrared ray excited orthogonal fluorescent emission of redgreenblue and preparation method thereof |
-
2017
- 2017-11-22 CN CN201711174200.9A patent/CN109813687A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN201394004Y (en) * | 2009-05-07 | 2010-02-03 | 复旦大学 | Up-conversion luminescence in-vivo rat imaging system of steady-state laser pumping |
| CN107286924A (en) * | 2017-06-13 | 2017-10-24 | 复旦大学 | Upper conversion nano crystalline material of the infrared ray excited orthogonal fluorescent emission of redgreenblue and preparation method thereof |
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