CN113218846A - ICP device for single-cell mass spectrometry flow analysis - Google Patents
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Abstract
The invention provides an ICP device for single-cell mass spectrometry flow analysis, which mainly comprises: the system comprises a monochromator, a radio frequency generator, a sample introducing system, a photoelectric conversion device, a control system, a data processing system and analysis operation software; the invention provides a use method of an ICP device for single-cell mass spectrometry flow analysis, which mainly comprises the following steps: generating plasma, adding a sample, converting signals, processing data and the like. Compared with the conventional detection mode, the device provided by the application can detect the surface of the cell membrane and the intracellular protein at the same time: the cell surface markers, the cell internal markers and the cell nucleus can be simultaneously dyed, the number of detection channels can reach 100, and the requirement on comprehensive analysis of biological samples can be met; no interference exists between channels, calculation and compensation are not needed, and the background is extremely low; and the diversified data processing mode realizes the in-depth analysis of the sample.
Description
Technical Field
The invention relates to the field of mass flow analysis, in particular to an ICP device for single-cell mass flow analysis.
Background
Cytometry is the measurement of different properties of cells. The measured parameter may be the size, shape, quantity, etc. of the cell, or may be an event corresponding to a certain process. Cytometry is derived from the counting of blood cells as a means of observing and initially measuring blood-related pathologies. The apparatus used to carry out the cytometry is known as a cytometer.
The flow cytometry method is characterized in that the cells are physically limited (such as reducing the aggregation degree of the cells in a solution, controlling the size of a cell channel and the like), so that the arrangement of the cells tends to be in a single cell state rather than an aggregated state, and measurement and counting of a certain character by taking the cells as a unit are realized. This approach made flow cytometry one of the first single cell detection instruments.
Over the course of a half century of development, cytometers have been developed to count cells by expressing them through various technologies, including fluorescent cytometers and mass spectrometers (mass cytometers) emerging in the last decade.
Traditional fluorescence flow-based techniques are mainly based on the detection of fluorescence emission spectra, but fluorophore emission spectra are generally broad and often overlap.
Firstly, it can bring about a serious cross color problem, signals of many channels need complex compensation calculation to deduct 'leakage' optical signals among the channels, and the compensation is an information deduction process, which not only can cause information loss, but also can introduce more interference to reduce the accuracy of data;
second, the compensation calculation is very complex and as the colors used increase, the compensation becomes more complex, which limits the number of channels that the flow can increase, and it is currently difficult for fluorescent flows to exceed 20 colors.
To solve these problems, mass spectrometry has been developed. Compared with the traditional fluorescence flow cytometry technology, the method has two main improvements:
firstly, different from the label system, the fluorescence flow technology mainly uses various fluorophores as labels of antibodies, and the latter uses various metal elements as labels;
second, the difference of the detection system is that the former uses a laser and a photomultiplier as a detection means, and the latter uses an ICP mass spectrometry technique as a detection means. Because the cells are marked by the metal isotope and detected by mass spectrometry, the color cross problem of the traditional fluorescence flow can be avoided, no interference exists among detection channels, and compensation calculation is not needed. Therefore, the mass spectrum flow type can detect more channels than the fluorescence flow type, and detection of more parameters is skillfully realized.
Compared with the traditional flow cytometry technology, the ICP-based mass spectrometry has the following characteristics:
first, the number of channels increases to hundreds. An ICP mass spectrometer in a mass cytometer has a very wide atomic weight detection range (88-210 Da), so that hundreds of different parameters can be detected for a single cell at the same time.
Second, there is no interference between channels and no need to calculate compensation. The ICP mass spectrometry has an ultra-high resolving power, and can completely distinguish various elements for labeling. Experimental data show that the interference between adjacent channels is less than 0.3%, which can be basically ignored and no compensation needs to be calculated. Therefore, the experimental process is simplified, and the samples and the reagents are saved.
Third, the number of metal tags is high and has a very low background. The content of the metal element used as the label in the cell is extremely low, and the nonspecific binding capacity of the metal label and the cell component is extremely low, so the background of the signal is extremely low. There are more than 30 kinds of metal labels currently used for labeling antibodies, and as the technology advances, more elements can be used as labels, and the variety will further increase.
Fourthly, the deep analysis of the sample is realized by diversified data processing modes. The proliferation of the number of channels in mass cytometry has led to a doubling of the amount of information. The traditional flow analysis method can not completely meet the requirement, so that various dimension reduction processing needs to be carried out on the data to extract useful biological information contained in the data. Common analytical methods are: SPADE, PCA, visNE, and Gemstone, among others.
The measurement by mass to charge ratio extends the number of different traits of cells detected simultaneously. Third generation mass spectrometerHaving 135 detection channels, e.g. commercially available Helios (TM), asystem, clearly surpassing the dozen detection channel comparison of the classical fluorescence cell instrument, and the expansion improves the possibility of researching the connection and change between various different characters of cells.
However, there has been little and no research on the mass spectrometer itself. Over the course of decades, mass cytometry has not achieved the functionality that scientists have sought. Although some subject groups in China are usedBut there are few published articles on the design of mass cytometry itself and its design is prone to analysis of cellular material.
To commercializeA mass cytometer is an example, and the mass cytometer expresses cell properties by the mass-to-charge ratio of a tag metal to realize cell counting.The device consists of a cell channel, an inductive coupled plasma (inductively coupled plasma) ionization component and a time of flight (flight) mass analysis component. Wherein ICP (inductively coupled plasma) is also called as a plasma reflectometer and is a mass cytometryThe key components of (a).
The invention aims to solve the problems that the mass spectrum flow type analysis device formed by the ICP plasma reflection device can meet various requirements which cannot be met by the conventional fluorescence flow type detection device, and relatively speaking, the accuracy, the stability and the overall detection efficiency of a detection result are influenced by the problems that the fluorescence flow type detection device is low in detection speed, narrow in measurement range, easy to receive background influence of an analysis result and the like.
At present, an ICP plasma reflecting device based on single-cell mass spectrometry flow analysis is not available in domestic markets. Therefore, it is a problem to be solved by those skilled in the art how to provide an ICP apparatus for single-cell mass spectrometry.
Disclosure of Invention
At present, how to provide an ICP apparatus for single-cell mass spectrometry flow analysis, which is applied to the field of cell detection, has solved the problems of slow detection speed, narrow measurement range, easy background influence on analysis results, and the like of the conventional fluorescence flow detection apparatus, and is a problem to be solved urgently for those skilled in the art.
In order to solve the technical problem, the invention provides an ICP apparatus for single-cell mass spectrometry, the ICP apparatus is an ICP inductively coupled plasma reflecting apparatus, and the ICP inductively coupled plasma reflecting apparatus comprises: the device comprises a monochromator, a radio frequency generator, a sample introducing system, a photoelectric conversion device, a control system, a data processing system and analysis operation software.
The operating principle of the ICP inductively coupled plasma reflecting device is as follows: the emitter is based on the characteristic radiation generated by exciting the atoms or ions of the elements to be measured in the light source, and the qualitative and quantitative analysis is carried out on each element by judging the existence of the characteristic radiation and the intensity of the characteristic radiation.
Furthermore, the radio frequency generator is an ICP instrument reaching international similar instruments, and can realize functions of automatic ignition, automatic control and the like.
Furthermore, the circuit type of the radio frequency generator is an inductive feedback self-excited oscillation circuit, coaxial cable output, matching tuning and power feedback signal acquisition, and closed-loop automatic control is carried out.
Further, the operating frequency of the radio frequency generator is: 40.68 MHz.
Further, the frequency stability of the radio frequency generator: less than or equal to 0.1 percent.
Further, the output power of the rf generator: 750 ~ 1600W are adjustable.
Further, the output power stability of the rf generator: less than or equal to 0.1 percent.
Further, the electromagnetic field leakage intensity of the radio frequency generator: distance 20cm from fuselage, electric field intensity E: is less than 2V/m.
Furthermore, an output working coil of the sample feeding device is a red copper load coil, and cooling water is introduced into the sample feeding device with the inner diameter of 25mm and 3 turns.
Furthermore, a quartz torch tube with three concentric types and an outer diameter of 20mm is adopted as a torch tube of the sample injection device.
Further, the external diameter of the coaxial sprayer of the sample injection device is 6 mm.
Further, the outer diameter of the double-cylinder type fog chamber of the sample injection device is 35 mm.
Furthermore, the observation position of the sample feeding device can be longitudinally observed before and after the height and the transverse observation, and can be intuitively adjusted to be optimal through software.
Furthermore, the sample introduction device is provided with a control flow meter.
Further, the optical path of the monochromator is of the Czerny-Turner type.
Further, the focal length of the monochromator is 1000 mm.
Further, the grating of the monochromator adopts an ion etching holographic grating, the groove density is 3600 lines/mm, and the groove area is as follows: 80X 110 mm.
Further, the groove density of the grating of the monochromator is 2400 lines/mm, and the groove area: 80X 110 mm.
Further, the reciprocal of the linear dispersion ratio of the monochromator was 0.26 nm.
Further, the resolution of the monochromator: < 0.007nm (3600 lines/mm grating was chosen).
Further, the scanning wavelength range of the monochromator: 3600 lines/mm scanning wavelength range: 180-530 nm, 2400 lines/mm scanning wavelength range: 180-800 nm.
Further, the stepper motor of the monochromator drives the minimum step: 0.0003 nm.
Further, the exit and entrance slits of the monochromator were 20 μm and 25 μm, respectively.
Further, the mirror specification of the monochromator is 80 × 110 × 16 mm.
Further, the lens of the monochromator is phi 30,1:1 imaging.
Further, the reflector of the monochromator is a concave reflector.
Furthermore, the monochromator is also provided with a monochromator heat preservation device, the optimal working temperature of the monochromator is 25 +/-3 ℃, and the monochromator heat preservation device is used for achieving the auxiliary function that the temperature of the monochromator is increased according with the optimal working temperature range of the instrument when the room temperature is lower than 20 ℃ in winter. After the function is started, the temperature can be automatically controlled in the optimal temperature working range of the instrument.
Further, the photomultiplier of the photometric device is charged with a high voltage: the temperature can be automatically adjusted at 0-1000V, and the stability is less than 0.03%.
Furthermore, the current measuring range of the photomultiplier of the photometric device is 10 < -4 > to 10 < -9 > A.
Further, the signal acquisition of the photometric device is V/F exchange, wherein 1mV corresponds to 100 Hz.
Furthermore, the sampling circuit of the photometric device uses a high-precision operational amplifier, so that the sensitivity and the accuracy of the photometric device are greatly improved.
Furthermore, the instrument data acquisition obtained by the photometric device is connected with a computer through a serial port, so that the circuit is simplified, and the connection is more convenient.
Furthermore, the light measuring mode of the light measuring device is single-element and multi-element sequential measurement.
Further, the requirements of the data processing system and the analysis operating software are:
(1) operating the system: chinese and English operation software under Windows7, Windows8 and Windows10 operation platforms;
(2) measuring the number of wavelengths: optionally selecting a spectral line;
(3) a database: more than 11 ten thousand spectral line banks;
(4) multi-window: after the result is measured for one time, the next sample can be measured while the last result is kept in the display window;
(5) there are standard addition methods: the software has a standard adding method so as to be used under different conditions and applications;
(6) analysis mode: the software defaults to an optimal analysis mode, and has multiple functions of instrument diagnosis, spectrogram analysis, several measurement modes, several integral modes, unit optional selection and the like;
(7) a history database: the analysis results are automatically saved, and the batch analysis results which are required to be printed can be selectively printed in the historical database;
(8) output data request: the software supports the laser printer to print the analysis results.
The invention provides a detection method of the ICP inductively coupled plasma reflection device, which mainly comprises the following steps:
step 2, forming aerosol from the aqueous solution to be detected through a sprayer and allowing the aerosol to enter a central channel of the quartz torch tube;
and 3, ionizing the atoms under the action of external energy, wherein the atoms in the excited state are unstable, and release huge energy when transitioning from a higher energy level to a ground state, and the energy is radiated in the form of electromagnetic waves with certain wavelengths. Different elements produce different characteristic spectra. The characteristic spectra are emitted to a grating in the optical splitter through a lens, the grating is calculated by controlling a stepping motor to rotate, and the transmission mechanism accurately positions the light intensity of the characteristic spectral line of the element to be measured after light splitting at an outlet slit;
step 4, converting the light intensity of the spectral line into current through a photomultiplier of the photoelectric conversion device, and then forming an electric signal after processing and V/F conversion through a circuit operational amplifier;
and 5, processing the data of the electric signals through a computer, and finally printing an analysis result by a printer.
The invention has the following effects:
(1) cell membrane surface and intracellular proteins can be detected simultaneously: simultaneous staining of cell surface markers, cell internal markers and cell nuclei can be achieved.
(2) The channel can be detected simultaneously: the number of detection channels can reach more than 100 internationally, and the requirement of comprehensive analysis on biological samples can be met.
(3) No interference exists between channels, no calculation compensation is needed, and the background is extremely low: the method can adopt a time-of-flight mass spectrometry (TOF) technology, has extremely high signal resolution capability, and can completely separate signals of various isotope labels. In addition, the content of the metal element as a label in the cell is extremely low, and the nonspecific binding capacity with the cell component is also extremely low, so that the signal background is extremely low.
(4) The diversified data processing mode realizes the in-depth analysis of the sample: the proliferation of the number of channels in a mass cytometer has multiplied the amount of information, and various dimensionality reduction processes need to be performed on the data to extract useful biological information contained therein.
Drawings
FIG. 1 is a schematic diagram comparing conventional fluorescence flow and mass flow techniques;
FIG. 2 is a schematic diagram of the principle of mass flow technology.
Detailed Description
The present invention is described in further detail below with reference to specific examples and with reference to the data. It will be understood that these examples are intended to illustrate the invention and are not intended to limit the scope of the invention in any way.
Example 1
An ICP device for single-cell mass spectrometry flow analysis, the ICP device is an ICP inductively coupled plasma reflection device, and the ICP inductively coupled plasma reflection device comprises: the device comprises a monochromator, a radio frequency generator, a sample introducing system, a photoelectric conversion device, a control system, a data processing system and analysis operation software.
The operating principle of the ICP inductively coupled plasma reflecting device is as follows: the emitter is based on the characteristic radiation generated by exciting the atoms or ions of the elements to be measured in the light source, and the qualitative and quantitative analysis is carried out on each element by judging the existence of the characteristic radiation and the intensity of the characteristic radiation.
The radio frequency generator is an ICP instrument reaching international similar instruments and can realize functions of automatic ignition, automatic control and the like. The circuit type of the radio frequency generator is an inductive feedback self-excited oscillation circuit, a coaxial cable output, matching tuning and power feedback signal acquisition, and closed-loop automatic control is carried out. Operating frequency of the radio frequency generator: 40.68 MHz.
Frequency stability of the radio frequency generator: less than or equal to 0.1 percent. Output power of the radio frequency generator: 750 ~ 1600W are adjustable. Output power stability of the rf generator: less than or equal to 0.1 percent. Electromagnetic field leakage intensity of the radio frequency generator: distance 20cm from fuselage, electric field intensity E: is less than 2V/m.
The output working coil of the sample feeding device is a red copper load coil, and cooling water is introduced into the sample feeding device with the inner diameter of 25mm and 3 turns. The torch tube of the sample injection device adopts a three-concentric quartz torch tube with the outer diameter of 20 mm. The outer diameter of the coaxial sprayer of the sample injection device is 6 mm. The external diameter of the double-cylinder type fog chamber of the sample injection device is 35 mm. The observation position of the sample feeding device can be longitudinally observed before and after the height and the transverse observation, and can be intuitively adjusted to be optimal through software. The sample introduction device is provided with a control flowmeter.
The optical path of the monochromator is of the Czerny-Turner type. The focal length of the monochromator is 1000 mm. The grating of the monochromator adopts an ion etching holographic grating, the groove density is 3600 lines/mm, and the groove area is as follows: 80X 110 mm. Reticle density of the grating of the monochromator 2400 lines/mm, reticle area: 80X 110 mm. The reciprocal of the linear dispersion ratio of the monochromator was 0.26 nm. Resolution of monochromator: < 0.007nm (3600 lines/mm grating was chosen). Scanning wavelength range of monochromator: 3600 lines/mm scanning wavelength range: 180-530 nm, 2400 lines/mm scanning wavelength range: 180-800 nm. Step motor drive minimum step pitch of monochromator: 0.0003 nm. The exit and entrance slits of the monochromator are 20 μm and 25 μm, respectively. The mirror specification of the monochromator is 80X 110X 16 mm. The lens of the monochromator is phi 30,1:1 imaging. The reflector of the monochromator is a concave reflector.
The monochromator is also provided with a monochromator heat preservation device, the optimal working temperature of the monochromator is 25 +/-3 ℃, and the monochromator heat preservation device is used for achieving the auxiliary function that the temperature of the monochromator can be increased according with the optimal working temperature range of the instrument when the room temperature is lower than 20 ℃ in winter. After the function is started, the temperature can be automatically controlled in the optimal temperature working range of the instrument.
Negative high voltage of photomultiplier of photometry device: the temperature can be automatically adjusted at 0-1000V, and the stability is less than 0.03%. The current measuring range of the photomultiplier of the photometric device is 10 < -4 > to 10 < -9 > A. The signal acquisition of the photometric device is V/F exchange, 1mV corresponding to 100 Hz. The sampling circuit of the photometric device uses a high-precision operational amplifier, so that the sensitivity and the accuracy of the photometric device are greatly improved. The instrument data acquisition that the photometric device obtained is connected with the computer through the serial ports to simplified the circuit and made the connection more convenient. The light measuring mode of the light measuring device is single-element and multi-element sequential measurement.
In addition, the ICP device is also provided with a cymbal copper spring and shielding glass for improving good grounding performance and reducing ultraviolet radiation.
The requirements of the data processing system and the analysis operating software are:
(1) operating the system: chinese and English operation software under Windows7, Windows8 and Windows10 operation platforms;
(2) measuring the number of wavelengths: optionally selecting a spectral line;
(3) a database: more than 11 ten thousand spectral line banks;
(4) multi-window: after the result is measured for one time, the next sample can be measured while the last result is kept in the display window;
(5) there are standard addition methods: the software has a standard adding method so as to be used under different conditions and applications;
(6) analysis mode: the software defaults to an optimal analysis mode, and has multiple functions of instrument diagnosis, spectrogram analysis, several measurement modes, several integral modes, unit optional selection and the like;
(7) a history database: the analysis results are automatically saved, and the batch analysis results which are required to be printed can be selectively printed in the historical database;
(8) output data request: the software supports the laser printer to print the analysis results.
The technical quality standard of the ICP device for single-cell mass spectrometry flow analysis is as follows:
1. wavelength indicating error: the repeatability of +/-0.02 nm is less than or equal to 0.003nm and is superior to the A-level national standard;
2. scanning wavelength range: (180 nm-530 nm)/3600 line/mm, (180 nm-800 nm)/2400 line/mm (grating configuration 3600 line/mm or 2400 line/mm);
3. measurement resolution: less than or equal to 0.007nm (whole wave band);
4. repeatability of the instrument: the relative standard deviation RSD is less than or equal to 1.5 percent;
5. stability of the apparatus: the relative standard deviation RSD is less than or equal to 2.0 percent.
Single cell mass spectrometry-oriented ICP device working environment and facility requirements
1. The normal working environment of the instrument:
temperature: 20-28 deg.C (temperature change rate is less than 2 deg.C/h);
and (3) the other: indoor cleanness, dryness, no dust, no corrosive gas, no vibration and the like, and the door and window has good sealing property to prevent sand wind and moisture from entering the room.
2. Laboratory essential conditions:
2.1 Room
A room with about 15 square meters, a reagent cabinet for placing the marking liquid, a reagent rack, a computer desk, a computer chair and the like, and is provided with more than 2 air conditioners.
2.2 Power supply requirements
2.2.1 Power, Voltage, frequency requirements of the Power supply
(1) A host computer: 220V + -10%, 50Hz, single-phase AC,
(2) Microcomputer, printer: single phase AC 220V + -10%, 50Hz, 400W
(3) Intelligent temperature control cooling circulation water tank: single phase AC 220V + -10%, 50Hz, 1.5KW
(4) Air exhaust equipment: single phase AC 220V + -10%, 50Hz, 15W
2.2.2 Wiring mode
(1) When the laboratory is powered, the power supply is preferably independently pulled from the power distribution room and separated from the high-power equipment phase line.
(2) The phase line for the ICP of the mainframe is preferably separate from the microcomputer and the intelligent temperature controlled cooling circulation tank and other instrumentation phase lines.
(3) The power supply connected into the ICP chamber is connected with a 60A breaker firstly, then connected with a 32A breaker, a 20A breaker and a 10A breaker respectively, so as to supply different matched instruments.
2.2.3 grounding
In order to ensure the safe and stable operation of the instrument, the instrument must be grounded separately, a red copper plate with the length of no less than 300mm, the width of 300mm and the thickness of 3mm is buried in the depth of 1.5m underground, a red copper strip with the width of 30mm and the thickness of 0.5mm is used for welding and is connected to an indoor instrument connection position, more than 1kg of salt is added at the position where the red copper plate is buried, and a proper amount of water is added, so that the resistance is less than 4 omega.
2.2.4 requirement for exhaust
(1) When ICP is used for analysis and test, the exhausted gas is mainly argon, but part of metal vapor and generated various gaseous substances exist, so that an exhaust device is arranged above an ICP sampling system.
(2) The air suction opening is arranged above the main machine ICP plasma chamber, and the air exhaust equipment consists ofThe tube is a tube of a flexible material,the bend head is provided with a bend head,a three-way pipe is arranged on the upper end of the main body,the tube is a tube of a flexible material,andthe PVC reducing connecting pipe and the air exhaust fan.
(3) The air discharge amount is preferably that a piece of paper is stuck at the air exhaust opening and can be lightly sucked. Flame stability is too much affected; too small to play a role in air exhaust. The concrete condition is determined according to the length, thickness and trend of the exhaust pipeline.
(4) The air exhaust duct is suggested to useThe PVC pipe is 4 m long,1 to 2 of the elbows are provided,the pipe clamp is provided with one pipe clamp,0.5m of PVC pipe,Andthe PVC reducing connecting pipe is connected with an air exhaust fan.
(5) The air discharge amount is preferably that a piece of paper is stuck at the air exhaust opening and can be lightly sucked. Flame stability is too much affected; too small to play a role in air exhaust. The concrete condition is determined according to the length, thickness and trend of the exhaust pipeline.
(6) Suggested use of exhaust ductThe PVC pipe is 4 meters long and is provided with a plurality of PVC pipes,1-2 PVC elbows are provided,the PVC tee joints are 1 to be connected with an exhaust fan.
2.2.5 gas Source requirement
Argon gas: a steel bottle was used to fill at least two bottles with 99.99% argon. And (3) adjusting the pressure of the oxygen distribution and pressure reduction meter by two meters, wherein the inlet pressure is 2.5MPa, and the outlet pressure is adjusted within the range of 0-1 MPa. When in use, the outlet pressure is adjusted to be 0.2 MPa.
2.2.6 Cooling Water requirements: cooling water for ICP operation
(1) The water pressure reaches 0.1MPa, the flow rate is more than 5 liters/minute, and the resistance of the cooling water is more than 1 MOmega.
(2) Water source outlet should be installed and its outer diameterThe plastic pipe is matched with the joint.
(3) An intelligent temperature control cooling circulation water tank and 35 liters of distilled water.
2.3 laboratory other requirements
2.3.1 this instrument is a large precision inorganic analyzer, equipped with a microcomputer, requiring a clean and dustless environment, not subject to drastic changes in room temperature, and the ICP chamber must be separated from the chemical chamber to prevent acids, alkalis and other corrosive gases from attacking the instrument.
2.3.2 for analyzing ultra-trace elements (especially Ca Mg Si Zn P) and elements which are easy to be polluted by environment by ICP, the indoor environment and vessels are very careful, and the used deionized water or quartz secondary distilled water is qualified.
The CP inductively coupled plasma reflection apparatus of the present embodiment has the following features:
(1) automatic matching and tuning of output power and power parameter program setting;
(2) the excellent optical system and the advanced control system ensure accurate peak position positioning and excellent signal-to-back ratio;
(3) the method has the advantages that the method has extremely small matrix effect, has higher spectral line resolution, can separate out Hg313.154 and 313.183nm double-line spectral lines, and can separate out iron quadruple peaks;
(4) the measurement range is wide, the ultramicro-to-constant analysis is realized, and the dynamic linear range is 5-6 orders of magnitude; can realize qualitative and quantitative analysis
(5) The output power range of Rf is 750-1600W, the stability of the output power is less than 0.1%, the detection limit is low, and the detection limit of most elements can reach ppb level;
(6) the negative high voltage of the photomultiplier can be independently adjusted within the range of 0-1000 v, conditions can be independently set according to different spectral lines of different elements, and the photomultiplier has a better detection limit compared with a full spectrum instrument;
(7) the measurement precision is good, the stability is less than or equal to 1.5 percent relative to the standard deviation RSD, and the stability is superior to the national A-level standard;
(8) the powerful and friendly human-computer interface analysis software can process data, compile a method and analyze results in the determination process, and is real multi-task working software; the software has powerful data processing function, provides various methods such as internal standard correction, IECS and QC monitoring functions and the like, and can obtain the optimal background subtraction point to eliminate interference; the output data can be directly printed or a result report in an Excel format can be automatically generated, and the data can be upgraded for free for life;
(9) cymbal brass leaf springs and specially treated shielding glass are used to absorb ultraviolet rays while causing the instrument to radiate less than 2V/m. The safety of an operator is ensured by adopting high shielding and good grounding;
(10) and the water path and the air path are automatically protected.
2. The technical indexes of the whole ICP inductively coupled plasma reflection device are as follows:
(1) the analysis speed is fast: one minute analysis scans more than 15 elements fastest;
(2) scanning range: the scanning range of 3600 lines/mm is 180-530 nm, the resolution of the whole wave band is less than 0.007nm,
the 2400 line/mm scanning range is 180-800 nm, and the full-wave-band resolution is less than or equal to 0.008 nm. The mode is a sine rod which is driven by a pulse motor controlled by a computer, and the minimum scanning step distance is 0.0003 nm;
(3) wavelength registration error and repeatability: wavelength indicating error: the repeatability of +/-0.02 nm is less than or equal to 0.003 nm;
(4) the correlation coefficient is more than or equal to 0.9998 percent;
(5) repeatability: the relative standard deviation RSD is less than or equal to 1.5 percent;
(6) stability: the relative standard deviation RSD is less than or equal to 2.0 percent;
(7) measurement range: ultra trace to constant;
(8) the detection limit is low: ppb (ug/L) level;
(9) the analysis elements are more: the quantitative or qualitative analysis can be carried out on 70 metal elements and partial nonmetal elements (such as B, P, Si, Se and Te);
(10) the measurement mode is as follows: sequentially measuring;
(11) power: 750W-1600W adjustable;
(12) the operation is convenient and fast: the third-generation multi-window upgrading Chinese and English analysis software running under brand-new Windows7, Windows8 and Windows10 has higher speed and more complete functions, and multi-window and multi-task execution is carried out simultaneously.
Example 2
A detection method of an ICP device (inductively coupled plasma) reflecting device for single-cell mass spectrometry flow analysis mainly comprises the following steps:
step 2, forming aerosol from the aqueous solution to be detected through a sprayer and allowing the aerosol to enter a central channel of the quartz torch tube;
and 3, ionizing the atoms under the action of external energy, wherein the atoms in the excited state are unstable, and release huge energy when transitioning from a higher energy level to a ground state, and the energy is radiated in the form of electromagnetic waves with certain wavelengths. Different elements produce different characteristic spectra. The characteristic spectra are emitted to a grating in the optical splitter through a lens, the grating is calculated by controlling a stepping motor to rotate, and the transmission mechanism accurately positions the light intensity of the characteristic spectral line of the element to be measured after light splitting at an outlet slit;
step 4, converting the light intensity of the spectral line into current through a photomultiplier of the photoelectric conversion device, and then forming an electric signal after processing and V/F conversion through a circuit operational amplifier;
and 5, processing the data of the electric signals through a computer, and finally printing an analysis result by a printer.
Although the present invention has been described with reference to the preferred embodiments, it should be understood that various changes and modifications can be made therein by those skilled in the art without departing from the spirit and scope of the invention as defined in the appended claims.
Claims (10)
1. An ICP apparatus for single cell mass spectrometry flow analysis, wherein the ICP apparatus is an ICP inductively coupled plasma reflector apparatus, the ICP inductively coupled plasma reflector apparatus comprising: the device comprises a monochromator, a radio frequency generator, a sample introducing system, a photoelectric conversion device, a control system, a data processing system and analysis operation software.
2. The ICP apparatus for single-cell mass spectrometry flow analysis as claimed in claim 1, wherein the radio frequency generator employs an inductive feedback self-excited oscillation circuit, coaxial cable output, matching tuning, power feedback signal extraction for closed loop automatic control.
3. The ICP apparatus for single-cell mass spectrometry flow analysis according to claim 2, wherein the operating frequency of the rf generator is 40.68MHz, and the output power of the rf generator is: 750-1600W, frequency stability of the radio frequency generator: not more than 0.1%, the output power stability of the radio frequency generator: less than or equal to 0.1 percent.
4. The ICP apparatus for single-cell mass spectrometry flow analysis of claim 1, wherein the sample introduction system comprises an output work coil, a torch, a coaxial type nebulizer, a dual barrel type mist chamber, a control flow meter, and a viewing position.
5. The ICP apparatus for single-cell mass spectrometry flow analysis according to claim 4, wherein the output work coil is a 3-turn output work coil, and the inner diameter of the output work coil is 25 mm.
6. The ICP apparatus for single-cell mass spectrometry flow analysis of claim 4, wherein the torch is a three-concentric quartz torch with an outer diameter of 20 mm.
7. An ICP apparatus for single cell mass spectrometry flow analysis according to claim 4, wherein the coaxial type atomizer has an outer diameter of 6mm and the double barrel type mist chamber has an outer diameter of 35 mm.
8. The ICP apparatus for single cell mass spectrometry flow analysis according to claim 1, wherein the optical path of the monochromator is of the Czerny-Turner type, the monochromator has a focal length of 1000mm, the monochromator has an exit slit of 20 μm and the monochromator has an entrance slit of 25 μm, the monochromator is provided with a temperature maintenance device, and the temperature maintenance device is automatically controlled by an automatic control device when the temperature is lower than 20 ℃.
9. The ICP apparatus for single-cell mass spectrometry flow analysis according to claim 1, wherein the photomultiplier of the photoelectric conversion device has a negative high voltage: the photoelectric sensor is automatically adjusted at 0-1000V, the stability is less than 0.03%, the current measurement range of the photomultiplier is 10 < -4 > -10 < -9 > A, the signal acquisition of the photomultiplier is that the V/F exchange is 1mV corresponding to 100Hz, and the light measurement mode of the photomultiplier adopts single-element and multi-element sequential measurement.
10. The ICP apparatus for single-cell mass spectrometry flow analysis according to claim 9, wherein a sampling circuit of the photoelectric conversion device employs an operational amplifier, and data collected by the photoelectric conversion device is connected to a computer through a serial port.
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Citations (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
DE69316807D1 (en) * | 1992-09-11 | 1998-03-12 | At & T Corp | Apparatus containing a mass spectrometer |
JP2001013074A (en) * | 1999-07-01 | 2001-01-19 | Nkk Corp | Laser icp analyzing method of high temperature sample |
JP2004198144A (en) * | 2002-12-16 | 2004-07-15 | Kobe Steel Ltd | Method for analyzing composition and/or particle size of nonmetallic inclusion in metal sample |
CN2697644Y (en) * | 2003-08-09 | 2005-05-04 | 吉林大学 | Inductive coupling plasma spectrometer |
CN105190829A (en) * | 2013-04-17 | 2015-12-23 | 富鲁达加拿大公司 | Sample analysis for mass cytometry |
CN107255722A (en) * | 2017-04-26 | 2017-10-17 | 马鞍山易廷生物科技有限公司 | Streaming combination ICP MS single cell protein detection method is marked based on metal isotope |
CN107796748A (en) * | 2017-09-28 | 2018-03-13 | 上海交通大学 | A kind of detection method for unicellular mass spectrum flow cytometry |
-
2021
- 2021-03-18 CN CN202110291239.9A patent/CN113218846A/en active Pending
Patent Citations (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
DE69316807D1 (en) * | 1992-09-11 | 1998-03-12 | At & T Corp | Apparatus containing a mass spectrometer |
JP2001013074A (en) * | 1999-07-01 | 2001-01-19 | Nkk Corp | Laser icp analyzing method of high temperature sample |
JP2004198144A (en) * | 2002-12-16 | 2004-07-15 | Kobe Steel Ltd | Method for analyzing composition and/or particle size of nonmetallic inclusion in metal sample |
CN2697644Y (en) * | 2003-08-09 | 2005-05-04 | 吉林大学 | Inductive coupling plasma spectrometer |
CN105190829A (en) * | 2013-04-17 | 2015-12-23 | 富鲁达加拿大公司 | Sample analysis for mass cytometry |
CN107255722A (en) * | 2017-04-26 | 2017-10-17 | 马鞍山易廷生物科技有限公司 | Streaming combination ICP MS single cell protein detection method is marked based on metal isotope |
CN107796748A (en) * | 2017-09-28 | 2018-03-13 | 上海交通大学 | A kind of detection method for unicellular mass spectrum flow cytometry |
Non-Patent Citations (3)
Title |
---|
吴蔓莉: "《普通高等教育"十三五"环境工程类专业基础课规划教材 环境分析化学实验》", 31 March 2018, 西安交通大学出版社 * |
裴世鑫: "《光电子技术原理与应用》", 31 August 2016, 国防工业出版社 * |
陈少杰: "宽波段叶阶梯光栅光谱仪设计与标定方法研究", 《中国优秀博硕士学位论文全文数据库(博士)工程科技Ⅱ辑》 * |
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