CN208872669U - A kind of device using tumour cell in stimlated Raman spectrum detection blood - Google Patents
A kind of device using tumour cell in stimlated Raman spectrum detection blood Download PDFInfo
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- CN208872669U CN208872669U CN201821485131.3U CN201821485131U CN208872669U CN 208872669 U CN208872669 U CN 208872669U CN 201821485131 U CN201821485131 U CN 201821485131U CN 208872669 U CN208872669 U CN 208872669U
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Abstract
A kind of device using tumour cell in stimlated Raman spectrum detection blood, including a glass capillary, the both ends of glass capillary are respectively arranged with the first optical window and the second optical window；Further include a pump laser source, the second collimation lens, beam splitter, a pair are disposed in the optical path of pump laser source to axicon mirror and reflecting mirror, an object lens are provided between reflecting mirror and the second optical window；Reflecting mirror reflects the light beam of pump laser source into glass capillary；Further include a Stokes laser source, first collimation lens is provided between beam splitter and Stokes laser source；On the laser refraction that beam splitter issues Stokes laser source to the optical path of pump laser source；Collecting lens, filter and photodetector are disposed in the side of the first optical window.The utility model is circulating tumor cell in the excited Raman projection Real-time Imaging blood based on bessel beam, realizes and detects to the high sensitivity of different tumour cells.
Description
Technical field
The invention belongs to field of bioinformatics, are related to the detection method of tumour cell, specifically it is a kind of using by
Swash the device of tumour cell in Raman spectrum detection blood.
Background technique
Malignant tumour DISTANT METASTASES IN be clinically solid malignant treatment failure or recurrence the main reason for one of.And it follows
The presence of ring tumour cell is exactly the root of solid malignant DISTANT METASTASES IN.Circulating tumor cell is to be present in blood to have
The antigenicity of former tumour and the tumour cell of molecular genetic properties.Circulating tumor cell derives from primary tumo(u)r or metastatic tumor, when
The matrix environment of suitable organ, tissue is encountered, will settle down herein, grow i.e. tumorigenic transfer.However enter circulation
Most of tumour cell can all lose activity, only remote organ is reached less than 0.01%, by migration, adherency, mutually interpolymerized
Collection forms small cancer embolus, and under the conditions of suitable microenvironment, new tumour is generated, so as to cause the generation of metastases.Cause
This circulating tumor cell quantity is very rare and is hidden in tens billion of blood cells, is directed to the detection of circulating tumor cell at present
Great challenge.
U.S. National Institutes (NIH) are just pointed out in annual cancer research report in 2011, pass through Analysis of New Technology
Circulating tumor cell is epoch-making achievement in cancer research and clinical treatment history, can be effectively reduced the diagnosis and treatment of cancer at
This.It is pre- that circulating tumor cell is detected as curative effects, the judgements such as the relapse and metastasis of early detection tumour, assessment operation, radiotherapy, chemistry
Afterwards, determine that tumor cells feature, the suitable individualized treatment of selection etc. provide an important basis, it will be to cancer early stage
Investigation and personalized medicine generate revolutionary impact.
In the past ten years, circulating tumor cell detection technique has attracted the very big concern of industry.It is different according to mechanism,
These methods can be sorted out at present are as follows: immunocytochemical technique, RT-polymerase chain reaction, fluorescence in situ hybridization technique and
Flow cytometry etc..
However, current method often rely on fluorescent marker for chemistry targeting, this may upset in life system
Biological function.In addition blood sampling volume is relatively large, and screening efficiency is low, and detection means is single and quality is low, each to circulating tumor cell
The identification and analysis sensitivity of class molecular composition is limited.
Summary of the invention
It is described the purpose of the present invention is to provide a kind of device using tumour cell in stimlated Raman spectrum detection blood
This device for detecting tumour cell in blood to solve to detect the method sensitivity of tumour cell in the prior art not
High, blood sample handles cumbersome, the other reagents labels of needs and technical problem complicated for operation.
The present invention provides a kind of dresses using tumour cell in stimlated Raman spectrum detection blood for realizing the above method
It sets, the device includes a glass capillary, and the both ends of the glass capillary are respectively arranged with first optics
Window and second optical window；Further include a pump laser source, is successively set in the optical path of the pump laser source
The second collimation lens, beam splitter, a pair are equipped with to axicon mirror and reflecting mirror, in the reflecting mirror and second optics
An object lens are provided between window；The laser optical path that the beam splitter and the pump laser source issue is in 45 degree of folder
Angle, the reflecting mirror and the beam splitter are in 90 degree of angle, and the reflecting mirror reflects the light beam of pump laser source
Enter glass capillary；Further include a Stokes laser source, the beam splitter and the Stokes laser source it
Between be provided with first collimation lens；The laser refraction that the beam splitter issues Stokes laser source is to pumping laser
In the optical path in source, to reflect into glass capillary；A collection is disposed in the side of first optical window
The pumping laser is arranged in lens, filter and photodetector, the collecting lens, filter and photodetector
The light beam in source reflected after optical path on.
Further, be provided on the side wall of the glass capillary of the second optical window side sample into
Mouthful；A sample export is provided on the side wall of the glass capillary of the first optical window side.
Method using tumour cell in above-mentioned device detection blood includes the following steps:
1) for tumor cells specimens known to one group, the tumour cell sample is measured in the way of stimulated Raman scattering
Product obtain the stimlated Raman spectrum data of the chemical component of related substances in tumor cells specimens, are analyzed and swollen by data
The feature stimlated Raman spectrum of chemical component described in oncocyte sample, establishes database, and the database includes tumour
The excited Raman profile data collection of chemical component described in cell sample；
2) the excited Raman profile data for testing associated tumor cells characteristic chemical ingredient in sample to be tested, in this step
In, test sample enters one end of glass capillary under the action of a constant pressure, and test sample travels forward, and from institute
The side of the other end for the glass capillary stated is flowed out, and the optics closed windows of both ends end to end of glass capillary are placed in laser
Receive the measurement of stimlated Raman spectrum between photodetector；
3) excited Raman of associated tumor cells characteristic chemical ingredient in the sample to be tested for measuring acquisition in step 2 is special
Modal data is levied compared with the tumour cell characteristic chemical ingredient excited Raman data in the database established in step 1), determines and surveys
Whether contain tumour cell in test agent.
Further, in step 2, the Stokes light beam that Stokes laser source is issued is by beam splitter in sky
Between the upper pump beam issued with a pump laser source be overlapped, the light beam being collinearly overlapped is sent to a pair of of axicon mirror, use
In generating annular beam, then annular beam guiding reflecting mirror is reflected, then is focused in glass capillary by object lens
In test sample, focus on light beam is in bessel beam distribution in the test sample, and the Bezier of transmission is collected by collecting lens
Then light beam is directed to photodetector, obtain measurement stimlated Raman spectrum.
Wherein, filter is fixed on before photodetector to remove Stokes light beam ingredient.
Invention demonstrates a method the excited Raman projection imagings based on bessel beam, are used for unmarked volume chemical imaging,
It can be with the chemical substance in fast quantification three-D volumes, to quick and precisely detect the tumour cell in blood.
Further, the bessel beam, by Gaussian beam by twin shaft pyramid mirror generate annular beam after by object
Mirror, which focuses, to be generated；
Further, the glass capillary, length is less than 20 millimeters, and diameter is less than 500 microns, so as to entire capillary glass
Blood sample carries out the excited Raman projection imaging based on bessel beam in glass pipe；
The working principle of the invention is: blood sample is entered glass capillary, blood under the action of a constant pressure
Liquid is travelled forward with certain speed, and glass tube is vertically arranged between light source and photodetector.Stokes laser and pumping swash
Light beam of light source weight merga pass a pair of axicon mirror, generates annular beam and focuses on blood sample in glass capillary by object lens
On product, focus on light beam has bessel beam feature distribution, can keep focusing over long distances along capillary, almost covering is entire
Blood sample in capillary.It is long along the focusing of input bessel beam by the Raman signal that photodetector detects
The integral of entire blood sample stimulated Raman scattering intensity on degree is total chemical composition of 3D volume in entire focal zone,
Detection efficiency greatly improves.Simultaneously using the difference on the frequency between pumping incident laser and Stokes incident laser and with Raman
When the vibration frequency matching of active molecule, excitation generates resonant check, forms stimulated Raman scattering, substantially increases unimolecule
The sensitivity of detection provides high Raman signal light intensity or brightness, and it is special to have that chemical specificity is strong and the signal acquisition time is short etc.
Point.
The present invention is using the intrinsic molecular vibration raman spectral signal of tumour cell characteristic chemical ingredient as being excited to draw
The marker of graceful imaging projects real time imagery by the excited Raman based on bessel beam, and fast quantification measures entire capillary
The chemical substance of test sample in pipe selectively obtains the letter of tumour cell specified chemical ingredient in entire test sample
Breath.
The present invention is that the stimulated Raman scattering imaging technique based on bessel beam is that one kind is novel, is not necessarily to fluorescent marker
Imaging method.It is a kind of nonlinear coherent Raman scattering process, it inherits the molecular specificity of spontaneous Raman spectrum, can
Different kind organism molecule is imaged with the differentiation of selectivity；The extremely faint disadvantage of spontaneous Raman signal is overcome simultaneously, is greatly enhanced
Signal can realize quick living imaging, in terms of the unmarked imaging of the detection of tumour, DNA, small-molecule drug
Have wide application prospects.
The present invention is compared with prior art, its technical effect is that actively and apparent.The present invention is directed to circulating tumor cell
Clinical detection application demand, invented based on based on bessel beam excited Raman projection Real-time Imaging blood in follow
Ring tumour cell.And the multidisciplinary technology such as combination Raman spectroscopy, nonlinear optical technique and oncobiology is intersected and is melted
It closes, develops a kind of device using tumour cell in stimlated Raman spectrum detection blood, the circulating tumor in efficient detection blood
Cell establishes real-time diagnosis platform, realizes and detects to the high sensitivity of different tumour cells.Also turn simultaneously for diagnosing tumour clinic
It moves and treatment new strategy provides technical support of new generation.
Detailed description of the invention
Fig. 1 is a kind of structural schematic diagram that the device of tumour cell in blood is detected using stimlated Raman spectrum of the present invention.
Wherein, 1: photodetector；2: filter；3: collecting lens；4: the first optical windows；5: glass capillary；6: the second optics
Window；7: Stokes laser source；8: the first collimation lenses, 9: pump laser source；10: the second collimation lenses；11: beam splitter；
12: a pair of of axicon mirror；13: reflecting mirror；14: object lens；15: blood sample import；16: blood sample outlet.
Specific embodiment
Embodiment 1
As shown in Figure 1, the present invention provides a kind of device using tumour cell in stimlated Raman spectrum detection blood, packet
A glass capillary 5 is included, the both ends of the glass capillary 5 are respectively arranged with first optical window 4 and one
Two optical windows 6；
It further include a pump laser source 9, it is saturating to be disposed with the second collimation in the optical path of the pump laser source 9
Mirror 10, beam splitter 11, a pair are to axicon mirror 12 and reflecting mirror 13, in the reflecting mirror 13 and second optical window
An object lens 14 are provided between 6；It is in 45 between the laser optical path that the beam splitter 11 and the pump laser source 9 issue
The angle of degree, is in 90 degree of angle between the reflecting mirror 13 and the beam splitter 11, and the reflecting mirror 13 will pump
The light beam of laser source 9 is reflected into glass capillary 5；
It further include a Stokes laser source 7, between the beam splitter 11 and the Stokes laser source 7
It is provided with first collimation lens 8；The laser refraction that the beam splitter 11 issues Stokes laser source 7 swashs to pumping
In the optical path of light source, to reflect into glass capillary 5；
A collecting lens 3, filter 2 and photodetection are disposed in the side of first optical window 4
Device 1, after the light beam that the pump laser source 9 is arranged in the collecting lens 3, filter 2 and photodetector 1 is reflected
Optical path on (in the optical path of the length direction of the glass capillary 5).
Specifically, be provided on the side wall of the glass capillary 5 of 6 side of the second optical window a sample into
Mouthful；
A sample export is provided on the side wall of the glass capillary 5 of 4 side of the first optical window.
Embodiment 2
Using device shown in the embodiment of the present invention 1, the present invention carries out stimlated Raman spectrum analysis to the fat drips in blood.
Fat drips are made of the hydrophobic core that Lipid monolayer and neutral fats are constituted, and many albumen are distributed in surface.It is one " lazy
Property " cell inclusion.We take capillary glass length of tube 2mm, inside diameter: 100um.We take 5 milliliters of blood sample pumps
Enter glass capillary circulation, Raman diffused light is penetrated and is adjusted to 2,850 cm -1CH2Symmetric vibration, to be carried out to fat drips
Excited Raman imaging.Stokes laser source 7 generates two synchronous laser lights with pump laser source 9 with the repetition rate of 80MHz
Beam.Wherein, Stokes laser 7 has the fixed wave length of 1,040nm.Pumping laser 9 with 680 to 1,300nm can harmonic
It is long.Then by beam splitter 11, spatially light beam is combined.The light beam conversion of overlapping is circularized using two axicon mirrors 12
Light beam, then annular beam is directed to the reflection of reflecting mirror 13, is focused in glass capillary 5 on blood sample by object lens 14.
Focus on light beam is distributed in blood sample in bessel beam in entire glass capillary 5.After blood sample, we are used
60 times of water immersion objectives 3 collect the bessel beam of transmission.This high NA target ensures high RST collection efficiency, to prevent figure
As background carries out Cross-phase Modulation.Be filtered first by 2 pairs of transmitting light beams of filter, with remove Stokes light beam at
Point, it is then oriented to photodetector 1, photodetector 1 uses bulk silicon photodiode (S3994-01, Hamamatsu).
Total Raman image acquisition time is 1.2 seconds.
It is respectively by the pumping laser and Stokes laser power of the bessel beam center lobe of 10 times of object lens generation
0.6 and 25mW.By to 2,850 cm-1Locate excited Raman luminous intensity measurement, can clearly obtain total lipid content
0.12mM, the result demonstrate excited Raman micromole's sensitivity.
The present embodiment further highlights raw in the projection real time imagery quantization volume of the excited Raman based on bessel beam
The advantage of the speed of the global information of object molecule.
Notwithstanding the preferred embodiment of the present invention, it should be apparent to those skilled in the art that
Without departing from the spirit or scope of the present invention, it can carry out various modifications and change in the present invention.Therefore, this hair
It is bright to be intended to cover the modifications and variations of the invention come within the scope of the appended claims and their.
Claims (2)
1. a kind of device using tumour cell in stimlated Raman spectrum detection blood, it is characterised in that: including a capillary glass
Glass pipe, the both ends of the glass capillary are respectively arranged with first optical window and second optical window；Also wrap
A pump laser source is included, the second collimation lens, beam splitter, a pair are disposed in the optical path of the pump laser source
To axicon mirror and reflecting mirror, an object lens are provided between the reflecting mirror and second optical window；It is described
Beam splitter and the laser optical path that issues of the pump laser source be in 45 degree of angle, the reflecting mirror and the beam splitting
Mirror is in 90 degree of angle, and the reflecting mirror reflects the light beam of pump laser source into glass capillary；It further include this support
Gram this laser source, is provided with first collimation lens between the beam splitter and the Stokes laser source；Institute
On the laser refraction that the beam splitter stated issues Stokes laser source to the optical path of pump laser source, to reflect into capillary glass
Glass pipe；It is disposed with a collecting lens, filter and photodetector in the side of first optical window, it is described
Collecting lens, filter and photodetector be arranged in the pump laser source light beam reflected after optical path on.
2. a kind of device using tumour cell in stimlated Raman spectrum detection blood according to claim 1, feature
It is to include: to be provided with a sample inlet on the side wall of the glass capillary of the second optical window side；Institute
A sample export is provided on the side wall of the glass capillary for the first optical window side stated.
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| CN201821485131.3U CN208872669U (en) | 2018-09-12 | 2018-09-12 | A kind of device using tumour cell in stimlated Raman spectrum detection blood |
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| CN201821485131.3U CN208872669U (en) | 2018-09-12 | 2018-09-12 | A kind of device using tumour cell in stimlated Raman spectrum detection blood |
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Effective date of registration: 20221010 Address after: Room 4292, building 24, No.2 Xincheng Road, Pudong New Area, Shanghai, 201306 Patentee after: SHANGHAI LELEE LASER TECHNOLOGY Co.,Ltd. Address before: 519031 room 105-52680, No.6 Baohua Road, Hengqin New District, Zhuhai City, Guangdong Province (centralized office area) Patentee before: ZHUHAI HEJING TECHNOLOGY DEVELOPMENT Co.,Ltd. |
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