CN109374511A - A kind of optical path adjustment device of the flow cytometer without fluid path situation - Google Patents

A kind of optical path adjustment device of the flow cytometer without fluid path situation Download PDF

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Publication number
CN109374511A
CN109374511A CN201811177492.6A CN201811177492A CN109374511A CN 109374511 A CN109374511 A CN 109374511A CN 201811177492 A CN201811177492 A CN 201811177492A CN 109374511 A CN109374511 A CN 109374511A
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optical path
forward scattering
microballoon
light
detection circuit
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CN109374511B (en
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祝连庆
娄小平
董明利
郭阳宽
刘超
孟晓辰
张文昌
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Beijing Information Science and Technology University
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Beijing Information Science and Technology University
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    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10Investigating individual particles
    • G01N15/14Optical investigation techniques, e.g. flow cytometry
    • G01N15/1434Optical arrangements

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  • Dispersion Chemistry (AREA)
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  • Biochemistry (AREA)
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  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
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  • Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)

Abstract

A kind of optical path adjustment device the present invention provides flow cytometer without fluid path situation, described device includes: radiation source, irradiation spot shaping optical path, standard microballoon rotating device, non-forward scattering light beam shaping optical path, non-forward scattering optical detection circuit, forward scattering light and fluorescent light beam shaping optical path, forward scattering optical detection circuit, multicolor fluorescence light splitting optical path and multichannel fluorescence detection circuit, wherein, the radiation source provides excitation light source for fluorescence excitation;The irradiation spot shaping optical path is used for the illumination spot of light beam of light source boil down to certain size;The standard microballoon rotating device simulates individual cells for rotating the disk for being mounted with standard microballoon one by one by irradiation hot spot;The non-forward scattering light beam shaping optical path is for being focused a certain range of non-forward scattering light light beam;The non-forward scattering optical detection circuit is used to carry out photoelectric conversion to non-forward scattering light and realizes parameter extraction to the electric impulse signal of generation.

Description

A kind of optical path adjustment device of the flow cytometer without fluid path situation
The application is that application No. is 201610896195.1, applying date 2016.10.14, a kind of entitled streamings The divisional application of optical path adjustment device and method of the cell instrument without fluid path situation
Technical field
The present invention relates to the adjustment of the optical path of flow cytometer and Microsphere field, especially a kind of light of no fluid path situation Road adjusts device.
Background technique
In flow cytometer optical path adjustment system, including irradiating the adjustment of hot spot and the light splitting collection of optical excitation signal. Since the detection zone of microballoon in flow cytometer is located at the point of intersection of irradiation hot spot and liquid flow path direction, and liquid channel system is not Stablize to will cause microballoon and enter the relative position of detection zone and have differences, swashs so as to cause the irradiation of microballoon in the detection area Hair degree is not identical, causes forward scattering light, non-forward scattering light and each channel fluorescence difference in signal strength anisotropic big.Subsequent inspection Slowdown monitoring circuit obtains pulse parameter information inaccuracy.Existing optical adjustment system places one's entire reliance upon high-precision fluid path control system System, independence are poor.Fluid path control system complex is cumbersome, and the verification method for controlling precision and laminar flow effect has no standard Index.
A kind of optical path therefore, it is necessary to flow cytometer that can effectively solve the above problem in no fluid path situation adjusts dress It sets.
Summary of the invention
According to an aspect of the invention, there is provided a kind of optical path adjustment device of flow cytometer without fluid path situation, institute Stating device includes: radiation source, irradiation spot shaping optical path, standard microballoon rotating device, non-forward scattering light beam shaping light Road, non-forward scattering optical detection circuit, forward scattering light and fluorescent light beam shaping optical path, forward scattering optical detection circuit, polychrome Fluorescence spectrophotometer optical path and multichannel fluorescence detection circuit, wherein
The radiation source provides excitation light source for fluorescence excitation;
The irradiation spot shaping optical path is used for the illumination spot of light beam of light source boil down to certain size;
The standard microballoon rotating device is for rotating the disk for being mounted with standard microballoon to which simulation is single thin Born of the same parents are one by one by irradiation hot spot;
The non-forward scattering light beam shaping optical path is for gathering a certain range of non-forward scattering light light beam It is burnt;
The non-forward scattering optical detection circuit is used to carry out non-forward scattering light photoelectric conversion and to the electric arteries and veins of generation It rushes signal and realizes parameter extraction;
The forward scattering light and fluorescent light beam shaping optical path are used for a certain range of forward scattering light and fluorescence light Beam is focused;
The forward scattering optical detection circuit is used to carry out photoelectric conversion to forward scattering light and believe the electric pulse of generation Number realize parameter extraction;
The multicolor fluorescence light splitting optical path is corresponding for being separated and being transferred to by the fluorescence signal of each wave-length coverage Detection circuit, the multichannel fluorescence detection circuit are used to carry out each channel fluorescence signal photoelectric conversion and to the electric pulse of generation Signal realizes parameter extraction.
Preferably, using the laser of certain power and wavelength as radiation source, and pass through irradiation spot shaping optical path Compression shaping is carried out to laser transmitting light beam, which moves according to microballoon in standard microballoon rotating device The spot size compression of two orthogonal directions is realized in direction, while making to move microballoon in hot spot coke by adjusting spot shaping optical path It is irradiated at point.
Preferably, which includes one and is mounted with the turntable of standard microballoon, turntable driving dress It sets and a motor motion control circuit, the turntable for being mounted with standard microballoon passes through rotating blade drive and motor motion control Circuit is connected, and the control to rotary speed is realized by adjusting motor speed, to change microballoon by hot spot irradiation time and phase The illuminated time interval of adjacent microballoon.
Preferably, the turntable for being mounted with standard microballoon is made of bottom and top layer two parts, wherein having in one layer for putting The pit for setting standard microballoon, standard microballoon, which is sealed in pit, by the bonding of top layer and bottom prevents microballoon in rotary course In fall off.The turntable rotary course Plays microballoon for being mounted with standard microballoon passes through irradiation spot area one by one and forms lateral dissipate Penetrate light, forward scattering light and assorted fluorescence signal.
Preferably, which is placed in non-forward-scattering angle detection zone, to certain angle Non- forward scattering light in degree range is focused shaping, and the beam Propagation after shaping is electric to non-forward scattering light detection Road.The non-forward scattering optical detection circuit is realized at the photoelectric conversion of non-forward-scattering signal and the conditioning of electric impulse signal Reason, and realize that the electric pulse parameter to characterization microballoon characteristic extracts.
Preferably, the forward scattering light and fluorescent light beam shaping optical path are placed in illumination beam direction, to certain angle model Forward scattering light and multichannel fluorescent light beam in enclosing are focused shaping, and by the beam Propagation after shaping to forward scattering light Detection circuit and multicolor fluorescence light splitting optical path.The forward scattering optical detection circuit realizes the photoelectric conversion to forward-scattering signal And the conditioning processing of electric impulse signal, and realize that the electric pulse parameter to characterization microballoon characteristic extracts.
Preferably, which separates multichannel fluorescence signal by a series of Optical devices, And the assorted fluorescent light beam after separation is transferred to corresponding multichannel fluorescence detection circuit.The multichannel fluorescence detection circuit is real Now the conditioning of the photoelectric conversion of fluorescence signal and electric impulse signal is handled, and realizes the electric pulse parameter to characterization microballoon characteristic It extracts.
According to another aspect of the present invention, a kind of optical path adjusting method of the flow cytometer without fluid path situation is provided, is wrapped Include step:
Irradiation spot shaping optical path carries out the processing such as compressing to the light beam that radiation source generates, and obtains the irradiation of certain size Hot spot, and adjusting hot spot focal length passes through microballoon from focal point;
Standard microballoon rotating device adjusts the movement velocity of microballoon, changes microballoon detection frequency and microballoon irradiation time, from And change corresponding light pulse and electrical impulse frequency and duration;
Non- forward scattering light beam shaping optical path carries out the processing such as shaping to non-forward scattering light, and adjustment optical path focal length makes non- Detector in forward scattering optical detection circuit is in focal position;
Non- forward scattering optical detection circuit realizes the photoelectric conversion to non-forward scattering light, electric pulse conditioning, analog/digital conversion And the processing such as parameter extraction, identify the starting point and end point of electric pulse;
Forward scattering light and fluorescent light beam shaping optical path carry out the processing such as shaping to forward scattering light and fluorescent light beam, and will Beam Propagation after focusing is to multicolor fluorescence light splitting optical path;
Forward scattering light and each channel fluorescence light beam are carried out light-splitting processing according to wavelength information by multicolor fluorescence light splitting optical path, And by the beam Propagation after light splitting to forward scattering optical detection circuit and corresponding multichannel fluorescence detection circuit;
Forward scattering optical detection circuit realizes the photoelectric conversion to forward scattering light, electric pulse conditioning, analog/digital conversion and ginseng The processing such as number extraction, identify the starting point and end point of electric pulse;
Multichannel fluorescence detection circuit realizes that the photoelectric conversion to each fluorescence, electric pulse improve, analog/digital conversion and parameter mention It the processing such as takes, identifies the starting point and end point of electric pulse.
It should be appreciated that aforementioned description substantially and subsequent detailed description are exemplary illustration and explanation, it should not As the limitation to the claimed content of the present invention.
Detailed description of the invention
With reference to the attached drawing of accompanying, the more purposes of the present invention, function and advantage are by the as follows of embodiment through the invention Description is illustrated, in which:
Fig. 1 is the embodiment schematic diagram of optical path adjustment device of the flow cytometer of the present invention without fluid path situation;
Fig. 2 is that flow cytometer of the invention forms connection schematic diagram.
Fig. 3 a-3c is radiation source shown in Fig. 2 and a kind of exemplary structure schematic diagram for irradiating spot shaping optical path.Wherein Fig. 3 a shows change in shape of the laser in the hot spot after shaping lens of radiation source sending.Fig. 3 b-3c is shown in Fig. 3 a Hot spot light distribution schematic diagram is irradiated after shaping.
Fig. 4 a-4b shows an implementation diagram of standard microballoon rotating device and standard microballoon.
Fig. 5 shows a specific optical path example structure figure of schematical non-forward scattering light beam shaping optical path.
Fig. 6 shows the physical circuit module map of non-forward scattering optical detection circuit.
Fig. 7 shows the physical circuit module map of forward scattering optical detection circuit.
Fig. 8 shows the specific optical path example structure figure of schematical multicolor fluorescence light splitting optical path.
Fig. 9 shows the specific optical path example structure figure of another schematical multicolor fluorescence light splitting optical path.
Figure 10 shows a physical circuit module map of multichannel fluorescence detection circuit.
Specific embodiment
Hereinafter, the embodiment of the present invention will be described with reference to the drawings.In the accompanying drawings, identical appended drawing reference represents identical Or similar component or same or like step.
By reference to exemplary embodiment, the purpose of the present invention and function and the side for realizing these purposes and function Method will be illustrated.However, the present invention is not limited to exemplary embodiment as disclosed below;Can by different form come It is realized.The essence of specification is only to aid in those skilled in the relevant arts' Integrated Understanding detail of the invention.
It is described in detail for combination schematic diagram of the present invention, when describing the embodiments of the present invention, for purposes of illustration only, indicating The sectional view of device architecture can disobey general proportion and make partial enlargement, and the schematic diagram is example, should not be limited herein The scope of protection of the invention processed.In addition, the three-dimensional space of length, width and depth should be included in actual fabrication.
A kind of optical path adjustment device the present invention provides flow cytometer without fluid path situation, Fig. 1 are that streaming of the present invention is thin The system block diagram schematic diagram of optical path adjustment device of born of the same parents' instrument without fluid path situation, as shown in Figure 1, described device includes: radiation source 101, irradiate spot shaping optical path 102, standard microballoon rotating device 103, non-forward scattering light beam shaping optical path 104, it is non-before To scattering optical detection circuit 105, forward scattering light and fluorescent light beam shaping optical path 106, forward scattering optical detection circuit 107, more Color fluorescence spectrophotometer optical path 108 and multichannel fluorescence detection circuit 109, wherein
The radiation source 101 provides excitation light source for fluorescence excitation;
The irradiation spot shaping optical path 102 is used for the illumination spot of light beam of light source boil down to certain size;
The standard microballoon rotating device 103 is for rotating the disk for being mounted with standard microballoon to which simulation is single Cell is one by one by irradiation hot spot;
The non-forward scattering light beam shaping optical path 104 is used to carry out a certain range of non-forward scattering light light beam It focuses;
The non-forward scattering optical detection circuit 105 is used to carry out non-forward scattering light photoelectric conversion and to the electricity of generation Pulse signal realizes parameter extraction;
The forward scattering light and fluorescent light beam shaping optical path 106 are used for a certain range of forward scattering light and fluorescence Light beam is focused;
The forward scattering optical detection circuit 107 is used to carry out forward scattering light photoelectric conversion and to the electric pulse of generation Signal realizes parameter extraction;
The multicolor fluorescence light splitting optical path 108 is corresponding for being separated and being transferred to by the fluorescence signal of each wave-length coverage Detection circuit;
The multichannel fluorescence detection circuit 109 is used to carry out each channel fluorescence signal photoelectric conversion and to the electricity of generation Pulse signal realizes parameter extraction.
Fig. 2 is that a better embodiment of optical path adjustment device of the flow cytometer of the present invention without fluid path situation is illustrated Figure.Which schematically illustrates radiation source 101, standard microballoon rotating device 103, non-forward scattering light beam shaping optical paths 104 and forward scattering light and the several parts of fluorescent light beam shaping optical path 106.
Preferably, using the laser of certain power and wavelength as radiation source 101, and pass through irradiation spot shaping light Road 102 carries out compression shaping to laser transmitting light beam, and the irradiation spot shaping optical path 102 is according to standard microballoon rotating device The microballoon direction of motion realizes the spot size compression of two orthogonal directions in 103, while being made by irradiating spot shaping optical path 102 Movement microballoon is irradiated at light spot focus.Fig. 3 a-3c is radiation source shown in Fig. 2 and the one kind for irradiating spot shaping optical path Exemplary structure schematic diagram.Wherein Fig. 3 a shows shape of the laser in the hot spot after shaping lens of the sending of radiation source 101 Shape variation.Fig. 3 b-3c is irradiation hot spot light distribution schematic diagram after shaping shown in Fig. 3 a.
Preferably, Fig. 4 a-4b shows an implementation diagram of standard microballoon rotating device 103 and standard microballoon.It should Standard microballoon rotating device 103 includes one and is mounted with the turntable of standard microballoon, a rotating blade drive and a motor fortune Dynamic control circuit, the turntable for being mounted with standard microballoon are connected with motor motion control circuit by rotating blade drive, are passed through Control of the motor speed realization to rotary speed is adjusted, so that it is illuminated by hot spot irradiation time and adjacent microballoon to change microballoon Time interval.
Preferably, the turntable for being mounted with standard microballoon is made of bottom and top layer two parts, wherein having in one layer for putting The pit for setting standard microballoon, standard microballoon, which is sealed in pit, by the bonding of top layer and bottom prevents microballoon in rotary course In fall off.The turntable rotary course Plays microballoon for being mounted with standard microballoon passes through irradiation spot area one by one and forms lateral dissipate Penetrate light, forward scattering light and assorted fluorescence signal.
Fig. 5 shows a specific optical path example structure of schematical non-forward scattering light beam shaping optical path 104 Figure.Preferably, which is placed in non-forward-scattering angle detection zone, to certain angle range Interior non-forward scattering light is focused shaping, as shown in figure 5, collecting non-forward scattering light by collecting lens, then passes through After the filtering of barrier filter, then by condenser lens, by the beam Propagation after shaping to non-forward scattering optical detection circuit 105。
Non- forward scattering optical detection circuit 105 realizes the photoelectric conversion of non-forward-scattering signal and the tune of electric impulse signal Reason processing, and realize that the electric pulse parameter to characterization microballoon characteristic extracts.Fig. 6 shows non-forward scattering optical detection circuit 105 physical circuit module map.As shown in fig. 6, non-forward scattering optical detection circuit 105 includes sensor, for what will be collected Optical signal is converted into electric signal, the electric impulse signal of sensor output is amplified, filter etc. handles, and by the electricity after conditioning Pulse signal transmission is converted into digital signal to subsequent analog-to-digital conversion module, then is input to electric pulse parameter extraction module use Electric signal parameter needed for extracting.
Preferably, the forward scattering light and fluorescent light beam shaping optical path 106 are placed in illumination beam direction, to certain angle Forward scattering light and multichannel fluorescent light beam in range are focused shaping, are transferred to forward scattering optical detection circuit 107.It should Forward scattering optical detection circuit 107, which is realized, handles the photoelectric conversion of forward-scattering signal and the conditioning of electric impulse signal, and Realize that the electric pulse parameter to characterization microballoon characteristic extracts.
Fig. 7 shows the physical circuit module map of forward scattering optical detection circuit 107.As shown in fig. 7, forward scattering light is examined Slowdown monitoring circuit 107 include sensor, for the optical signal of collection to be converted into electric signal, to sensor output electric impulse signal into The processing such as row amplification, filtering, and the electric impulse signal after conditioning is transferred to subsequent analog-to-digital conversion module to be converted into number Signal, then electric pulse parameter extraction module is input to for extracting required electric signal parameter.
Fig. 8 and Fig. 9 respectively illustrates the specific optical path example structure figure of schematical multicolor fluorescence light splitting optical path 108. As shown in Figure 8 and Figure 9, forward scattering light is collected by collecting lens, is then divided by multiple dichroism spectroscopes, After bandpass filter filters, signal multiplication collection is carried out by multiple photomultiplier tubes (PMT1-6).Preferably, the polychrome is glimmering Light light splitting optical path 108 separates multichannel fluorescence signal by a series of Optical devices, and by the assorted fluorescence after separation Beam Propagation is to corresponding multichannel fluorescence detection circuit 109.
The multichannel fluorescence detection circuit 109 is realized to the photoelectric conversion of fluorescence signal and the conditioning of electric impulse signal Reason, and realize that the electric pulse parameter to characterization microballoon characteristic extracts.Figure 10 shows multichannel fluorescence detection circuit 109 One physical circuit module map.As shown in Figure 10, multichannel fluorescence detection circuit 109 includes multichannel fluorescence detection circuit, each Road includes sensor, for the optical signal of collection to be converted into electric signal, the electric impulse signal of sensor output is amplified, The processing such as filtering, and the electric impulse signal after conditioning is transferred to subsequent analog-to-digital conversion module to be converted into digital signal, then Electric pulse parameter extraction module is input to for extracting required electric signal parameter.
Another aspect of the present invention provides a kind of optical path adjusting method of the flow cytometer without fluid path situation, including step It is rapid:
The light beam that irradiation spot shaping optical path 102 generates radiation source 101 carries out the processing such as compressing, and obtains certain size Irradiation hot spot, and adjust hot spot focal length make microballoon from focal point pass through;
Standard microballoon rotating device adjusts the movement velocity of microballoon, changes microballoon detection frequency and microballoon irradiation time, from And change corresponding light pulse and electrical impulse frequency and duration;
Non- forward scattering light beam shaping optical path carries out the processing such as shaping to non-forward scattering light, and adjustment optical path focal length makes non- Detector in forward scattering optical detection circuit is in focal position;
Non- forward scattering optical detection circuit realizes the photoelectric conversion to non-forward scattering light, electric pulse conditioning, analog/digital conversion And the processing such as parameter extraction, identify the starting point and end point of electric pulse;
Forward scattering light and fluorescent light beam shaping optical path carry out the processing such as shaping to forward scattering light and fluorescent light beam, and will Beam Propagation after focusing is to multicolor fluorescence light splitting optical path;
Forward scattering light and each channel fluorescence light beam are carried out light-splitting processing according to wavelength information by multicolor fluorescence light splitting optical path, And by the beam Propagation after light splitting to forward scattering optical detection circuit and corresponding multichannel fluorescence detection circuit;
Forward scattering optical detection circuit realizes the photoelectric conversion to forward scattering light, electric pulse conditioning, analog/digital conversion and ginseng The processing such as number extraction, identify the starting point and end point of electric pulse;
Multichannel fluorescence detection circuit realizes that the photoelectric conversion to each fluorescence, electric pulse improve, analog/digital conversion and parameter mention It the processing such as takes, identifies the starting point and end point of electric pulse.
In conjunction with the explanation and practice of the invention disclosed here, the other embodiment of the present invention is for those skilled in the art It all will be readily apparent and understand.Illustrate and embodiment is regarded only as being exemplary, true scope of the invention and purport are equal It is defined in the claims.

Claims (3)

1. a kind of optical path adjustment device of flow cytometer without fluid path situation, which is characterized in that described device includes: irradiation light Source, irradiation spot shaping optical path, standard microballoon rotating device, non-forward scattering light beam shaping optical path, the inspection of non-forward scattering light Slowdown monitoring circuit, forward scattering light and fluorescent light beam shaping optical path, forward scattering optical detection circuit, multicolor fluorescence light splitting optical path and multi-pass Road fluorescence detection circuit, wherein
The radiation source provides excitation light source for fluorescence excitation;
The irradiation spot shaping optical path is used for the illumination spot of light beam of light source boil down to certain size;
The standard microballoon rotating device includes one and is mounted with the turntable of standard microballoon, a rotating blade drive and an electricity Machine motion control circuit, the turntable for being mounted with standard microballoon are connected by rotating blade drive with motor motion control circuit, The control to rotary speed is realized by adjusting motor speed, is shone to change microballoon by hot spot irradiation time and adjacent microballoon The time interval penetrated;
Standard microballoon rotating device is simulated individual cells for rotating the disk for being mounted with standard microballoon and is passed through one by one Cross irradiation hot spot;
The non-forward scattering light beam shaping optical path is placed in non-forward-scattering angle detection zone, within the scope of certain angle Non- forward scattering light is focused shaping, and by the beam Propagation after shaping to non-forward scattering optical detection circuit;
Non- forward scattering light beam shaping optical path is for being focused a certain range of non-forward scattering light light beam;
The non-forward scattering optical detection circuit is used to carry out photoelectric conversion to non-forward scattering light and believe the electric pulse of generation Number realize parameter extraction, realize non-forward-scattering signal photoelectric conversion and electric impulse signal conditioning processing, and realize pair The electric pulse parameter of characterization microballoon characteristic extracts;
The forward scattering light and fluorescent light beam shaping optical path be used for a certain range of forward scattering light and fluorescent light beam into Line focusing;
The forward scattering optical detection circuit is used to carry out forward scattering light photoelectric conversion and to the electric impulse signal of generation reality Existing parameter extraction;
The multicolor fluorescence light splitting optical path by the fluorescence signal of each wave-length coverage for being separated and being transferred to corresponding detection Circuit, the multichannel fluorescence detection circuit are used to carry out each channel fluorescence signal photoelectric conversion and to the electric impulse signal of generation Realize parameter extraction.
2. optical path adjustment device according to claim 1, it is characterised in that: made using the laser of certain power and wavelength For radiation source, and light beam is emitted to laser by irradiation spot shaping optical path and carries out compression shaping, the irradiation spot shaping Optical path realizes that the spot size of two orthogonal directions is compressed according to the microballoon direction of motion in standard microballoon rotating device, passes through simultaneously Adjustment spot shaping optical path is irradiated movement microballoon at light spot focus.
3. optical path adjustment device according to claim 1, it is characterised in that: be mounted with the turntable of standard microballoon by bottom and Top layer two parts composition, wherein there is the pit for placing standard microballoon in one layer, by the bonding of top layer and bottom by standard Microballoon, which is sealed in pit, prevents microballoon from falling off in rotary course.This is mounted with the turntable rotary course Plays of standard microballoon Microballoon passes through irradiation spot area one by one and forms side scattered light, forward scattering light and assorted fluorescence signal.
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