CN112444479B - Single cell mass spectrometry system and method based on parallel processing technology - Google Patents
Single cell mass spectrometry system and method based on parallel processing technology Download PDFInfo
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- 238000004949 mass spectrometry Methods 0.000 title claims abstract description 49
- 238000005516 engineering process Methods 0.000 title claims abstract description 29
- 238000000034 method Methods 0.000 title claims abstract description 20
- 238000004140 cleaning Methods 0.000 claims abstract description 42
- 238000001819 mass spectrum Methods 0.000 claims description 25
- 238000000605 extraction Methods 0.000 claims description 21
- 239000000284 extract Substances 0.000 claims description 13
- 238000003384 imaging method Methods 0.000 claims description 8
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- 238000004458 analytical method Methods 0.000 abstract description 11
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- 239000007924 injection Substances 0.000 description 6
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- 230000008569 process Effects 0.000 description 3
- 238000011160 research Methods 0.000 description 3
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
- G01N15/10—Investigating individual particles
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- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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Abstract
The invention provides a single-cell mass spectrometry system and a method based on a parallel processing technology, wherein the single-cell mass spectrometry system comprises a cell bearing unit, a cleaning unit and a mass spectrometer; the base is provided with a first rotating shaft; the bracket is connected with the first rotating shaft and rotates around the first rotating shaft; m groups of working units, wherein M is an integer not less than 1, each working unit comprises a mechanical arm and a capillary needle, and the capillary needles are arranged on the mechanical arms; in the rotation of the bracket, when a first working unit in any group of working units is in a first working position, a second working unit is in a second working position, and a third working unit is in a third working position; the cell bearing unit is arranged at each first working position, the mass spectrometer is arranged at each second working position, and the cleaning unit is arranged at each third working position. The invention has the advantages of high working efficiency, accurate analysis result and the like.
Description
Technical Field
The invention relates to cell analysis, in particular to a single-cell mass spectrometry system and a single-cell mass spectrometry method based on a parallel processing technology.
Background
The cells are basic composition units of the morphological structure and the life activities of organisms, the life activities are realized in the cells, and the growth, the differentiation, the metabolic propagation and the like of the cells are related to the overall state of the cells. Most of the previous researches are to analyze population cells to obtain an average result, with the continuous improvement of analysis technologies, people find that different cell individuals have differences, and the differences among single cells have important significance in the research of the life activities and the physiological and pathological processes of the cells. However, the analysis of the population cells cannot reflect the individual difference between the cells, and can mask the information of the single cells, so the single cell analysis is increasingly paid high attention.
At present, the single cell mass spectrometry technology can realize simultaneous detection of multi-component samples, obtain structural information of unknown components in a large number of samples, has high sensitivity, can solve the problem that the single cell samples are difficult to detect below nL level, and provides effective technical support for research of single cell metabonomics, proteomics and the like. The defects of the single-cell mass spectrometry technology are as follows:
the analysis efficiency is low, all the functional modules are executed in sequence, and the time is consumed in the process waiting; in addition, most single cell mass spectrometry devices do not consider cleaning, and there is a potential for cross contamination.
Disclosure of Invention
In order to overcome the defects in the prior art, the invention provides the single-cell mass spectrometry system based on the parallel processing technology, which has high working efficiency, good accuracy and low use cost.
The purpose of the invention is realized by the following technical scheme:
the single-cell mass spectrometry system based on the parallel processing technology comprises a cell bearing unit, a cleaning unit and a mass spectrometer, wherein the mass spectrometer is provided with a sample inlet; the parallel processing technology-based single-cell mass spectrometry system further comprises:
a base having a first rotational axis;
the bracket is connected with the first rotating shaft and rotates around the first rotating shaft;
m groups of working units, wherein M is an integer not less than 1, each group of working units comprises P working units, and P is an integer not less than 3; the working unit is arranged on the bracket and comprises a mechanical arm and a capillary needle, and the capillary needle is arranged on the mechanical arm; in the rotation of the bracket, when the first working unit in any group of working units is in the first working position, the second working unit is in the second working position, and the third working unit is in the third working position; the cell bearing unit is arranged at each first working position, the mass spectrometer is arranged at each second working position, and the cleaning unit is arranged at each third working position.
The invention also aims to provide a mass spectrometry method for single cells of the single cell mass spectrometry system based on the parallel processing technology, and the aim of the invention is realized by the following technical scheme:
the single cell mass spectrometry method of the single cell mass spectrometry system based on the parallel processing technology is applied, and comprises the following steps:
the bracket rotates, so that when the first working unit in any group of working units is positioned at the first working position to extract single cells, the second working unit which extracts the single cells is positioned at the second working position to realize mass spectrum sample introduction, and the third working unit after the mass spectrum sample introduction is positioned at the third working position to realize cleaning;
through rotation, the same working unit is in a single cell extraction state, a mass spectrum sample introduction state and a cleaning state in sequence.
Compared with the prior art, the invention has the beneficial effects that:
1. the working efficiency is high;
through the design of the working units, the first working unit in the same group of working units is positioned at the first working position to extract single cells, the second working unit which extracts the single cells is positioned at the second working position to realize mass spectrum sample introduction, and the third working unit after the mass spectrum sample introduction is positioned at the third working position to realize cleaning, namely, the single cells are extracted, the mass spectrum sample introduction and the cleaning corresponding to the same group of working units are simultaneously carried out, namely, the parallel processing is carried out, the time consumption of the waiting process of each link is greatly reduced, and the working efficiency is remarkably improved;
the M groups of working units work simultaneously (a single-circle one-time working cycle and multiple working cycles) so that the working efficiency is further improved;
2. the analysis result is accurate;
the working units are required to be cleaned at a third working position before the next extraction, so that cross contamination is avoided, and the final single-cell mass spectrometry data is more accurate and reliable;
by utilizing the suction pump, the cleaning fluid is repeatedly sucked into and discharged from the capillary needle, the cleaning effect is good, and the accuracy of the single-cell mass spectrometry data is ensured;
analyzing mass spectrum signals in real time and carrying out closed-loop control on the single-cell extraction process, so that the quality of the single-cell mass spectrum signals is ensured, and the accuracy of single-cell analysis is also ensured;
2. the use cost is low;
the capillary needle can be repeatedly used and put into use again after being cleaned, so that blockage is avoided.
Drawings
The disclosure of the present invention will become more readily understood with reference to the accompanying drawings. As is readily understood by those skilled in the art: these drawings are only for illustrating the technical solutions of the present invention and are not intended to limit the scope of the present invention. In the figure:
FIG. 1 is a schematic diagram of a single-cell mass spectrometry system based on parallel processing techniques according to an embodiment of the present invention;
FIG. 2 is a schematic structural diagram of a single-cell mass spectrometry system based on a parallel processing technique according to embodiment 3 of the present invention;
FIG. 3 is a schematic structural diagram of a single-cell mass spectrometry system based on parallel processing technology according to embodiment 4 of the present invention.
Detailed Description
Fig. 1-3 and the following description depict alternative embodiments of the invention to teach those skilled in the art how to make and use the invention. Some conventional aspects have been simplified or omitted for the purpose of explaining the technical solution of the present invention. Those skilled in the art will appreciate that variations or substitutions from these embodiments will be within the scope of the invention. Those skilled in the art will appreciate that the features described below can be combined in various ways to form multiple variations of the invention. Thus, the present invention is not limited to the following alternative embodiments, but is only limited by the claims and their equivalents.
Example 1:
fig. 1 is a schematic structural diagram of a single-cell mass spectrometry system based on a parallel processing technology according to an embodiment of the present invention, and as shown in fig. 1, the single-cell mass spectrometry system based on a parallel processing technology includes:
the cell loading unit 11, the cleaning unit 13 and the mass spectrometer 12, wherein the mass spectrometer 12 is provided with a sample inlet;
a base having a first rotational axis;
a bracket 41, wherein the bracket 41 is connected with the first rotating shaft and rotates around the first rotating shaft;
m groups of working units, wherein M is an integer not less than 1, each group of working units comprises P working units, and P is an integer not less than 3; the working unit is arranged on the bracket and comprises a mechanical arm and a capillary needle, and the capillary needle is arranged on the mechanical arm; in the rotation of the bracket 41, when the first working unit 21 in any group of working units is in the first working position, the second working unit 22 is in the second working position, and the third working unit 23 is in the third working position; the cell bearing unit 11 is positioned at each first working position, the mass spectrometer 12 is positioned at each second working position, and the cleaning unit 13 is positioned at each third working position; when the bracket 41 rotates clockwise or counterclockwise, the same working unit is sequentially in the first working position, the second working position and the third working position, that is, in the single cell extraction state, the mass spectrometry sample injection state and the cleaning state.
In order to adjust the angle of the capillary needle relative to the support 41, so that the same capillary needle is in a single cell extraction state, a mass spectrum sample injection state and a cleaning state at different times, further, the mechanical arm is provided with a second rotating shaft, and the capillary needle is connected with the second rotating shaft; the second rotation axis of each mechanical arm in any group of working units forms the same included angle with the radial direction of the support 41.
In order to deliver the cell extract and realize the function of cleaning the capillary needle, further, the single-cell mass spectrometry system further comprises:
and the suction pump is communicated with the inside of the capillary needle through the gas channel.
In order to deliver the cell extract and to perform the functions of washing the capillary needle and ionizing, further, the single-cell mass spectrometry system further comprises:
a fixed tube, the capillary needle being disposed within the fixed tube; the gas channel passes through the fixed pipe;
an electrode disposed within the capillary needle.
In order to reduce the structural complexity and the system operation reliability, further, the single-cell mass spectrometry system further comprises:
the driving unit drives the cell bearing unit 11 to translate two-dimensionally on a horizontal plane, so that the capillary needle selectively extracts single cells on the cell bearing unit 11, and the cell bearing unit 11 comprises cell bearing positions 111 which are distributed in a matrix form and are isolated from each other;
an imaging unit for imaging the cell-bearing sites 111 and the capillary needles on their upper side such that the capillary needles extract single cells of a selected cell-bearing site 111.
In order to achieve the washing function of the capillary needle, further, the washing unit 13 includes:
a first container having an outlet at a bottom thereof; when the capillary needle rotates, the bottom end opening of the capillary needle is positioned at the lower side of the outlet;
and the opening of the second container faces upwards and is arranged at the lower sides of the first container and the bottom end of the capillary needle.
The single-cell mass spectrometry method of the embodiment of the present invention, that is, the working method of the single-cell mass spectrometry system based on the parallel processing technology of the embodiment of the present invention, includes:
the bracket 41 rotates around the first rotating shaft, so that when the first working unit 21 in any group of working units is positioned at the first working position to extract single cells, the second working unit 22 which extracts the single cells is positioned at the second working position to realize mass spectrum sample introduction, the third working unit 23 after the mass spectrum sample introduction is positioned at the third working position to realize cleaning, the single cells are extracted, the mass spectrum sample introduction and the cleaning are simultaneously carried out, and the working efficiency is obviously improved;
in the process, all the working units in the M groups of working units work simultaneously, wherein the working units in any group of working units are respectively in a single cell extraction state, a mass spectrum sample introduction state and a cleaning state; along with the rotation of the bracket 41, the same working unit is in a single cell extraction state, a mass spectrum sample introduction state and a cleaning state in sequence.
Example 2:
the application example of the single-cell mass spectrometry system and the method based on the parallel processing technology in the embodiment 1 of the invention.
In the present application example, as shown in fig. 1, a group of working units is disposed on the bracket 41, the only group of working units includes three working units, the three working units (the first working unit 21, the second working unit 22, and the third working unit 23) are uniformly disposed on the entire circumference of the bracket 41, and the central angle corresponding to the adjacent working units is 120 degrees;
the cell bearing unit 11 is arranged at a first working position, the mass spectrometer 12 is arranged at a second working position, and the cleaning unit 13 is arranged at a third working position; the cell bearing unit 11 is provided with cell bearing positions 111 and a third container 112 which are distributed in a matrix form and are isolated from each other, and cell extraction liquid is filled in the third container 112; the cell bearing unit 11 is driven by the driving unit to translate, so that the capillary needles in the working units at the first working position extract the single cells selected on the cell bearing unit 11; in order to confirm that the capillary needle extracts the selected single cell, a microscopic imaging unit is arranged at the lower side of the cell bearing unit 11, and whether the capillary needle extracts the selected single cell is judged through the imaging of the cell bearing position 111 and the capillary needle; the mechanical arm is provided with a second rotating shaft, and the capillary needle is connected with the second rotating shaft; the included angle between the second rotating shaft of each mechanical arm in any group of working units and the radial direction of the support 41 is the same; the bottom end of the capillary needle is designed to be a pointed end, and the upper end of the capillary needle is arranged in the fixed tube and is sealed; the needle-shaped electrode is arranged in the capillary needle and is connected with an external power supply; the fixed tube is provided with an opening, and a suction pump is communicated with the inside of the capillary needle through the opening; the cleaning unit 13 includes a first container having an opening at the bottom thereof, a valve being provided at the opening, and a second container provided at the lower side of the first container;
the single cell mass spectrometry method of the embodiment of the invention specifically comprises the following steps:
the bracket 41 rotates around the first rotating shaft in the forward direction, so that the first working unit 21 is in the first working position, meanwhile, the second working unit 22 is in the second working position, and the third working unit 23 is in the third working position;
the first working unit 21 is in the first working position: under the action of the driving unit and the rotation of the mechanical arm, the first capillary needle 24 of the first working unit 21 extends into the third container 112, the suction pump works, and the cell extract in the third container 112 is sucked into the first capillary needle 24;
under the action of the driving unit and the rotation of the mechanical arm, and under the imaging and judgment of the microscopic imaging unit, the bottom end of the first capillary needle 24 is positioned at the upper side of the single cell on the selected cell bearing position 111, moves downwards, the suction pump works, the cell extraction liquid in the first capillary needle 24 is discharged out of the first capillary needle 24 to cover the selected single cell, and then the first capillary needle 24 moves upwards;
after the extraction is completed, the first capillary needle 24 moves downwards, and the suction pump works at the same time, so that the single-cell extraction substance enters the first capillary needle 24;
the second working unit 22 is in the second working position: the second capillary needle 25 of the second working unit 22, which has previously completed cell extraction, has therein a cell extraction substance; under the action of the mechanical arm, the bottom outlet of the second capillary needle 25 corresponds to the sample inlet of the mass spectrometer 12, the electrode discharges, the cell extraction substance is ionized and is ejected out of the bottom outlet to enter the sample inlet, and the single cell information is obtained after mass spectrometry;
the third working unit 23 is in the third working position: the third capillary 26 of the third working unit 23 that has completed the mass spectrometry sample injection is under the action of the mechanical arm, the bottom outlet is at the lower side of the outlet of the first container, the valve is opened, and at the same time, the suction pump works, so that the cleaning solution at the bottom outlet is sucked into the third capillary 26; then, under the action of the suction pump, the cleaning liquid in the third capillary needle 26 is discharged and dropped into the second container on the lower side; the cleaning solution is repeatedly sucked and discharged, so that the third capillary needle 26 is cleaned without contacting the cleaning solution in the first container, cross contamination is prevented, and the accuracy of single cell analysis is ensured;
the bracket 41 positively rotates for 120 degrees around the first rotating shaft, the first capillary needle 24 rotates to a second working position, the cell extraction substance in the first capillary needle 24 is ionized and sent to the mass spectrometer 12 for analysis, and the mode is the same as the mass spectrum sample introduction of the second capillary needle 25; meanwhile, the second capillary needle 25 rotates to a third working position, the second capillary needle 25 is cleaned in the same manner as the third capillary needle 26, the cleaned third capillary needle 26 rotates to the first working position, single cells are selected and extracted, and the specific manner is the same as that of the first capillary needle 24;
the bracket 41 continuously rotates forwards for 120 degrees around the first rotating shaft, the first capillary needle 24 rotates to a third working position, the first capillary needle 24 is cleaned in the same way as the third capillary needle 26, meanwhile, the cleaned second capillary needle 25 rotates to the first working position, single cells are selected and extracted, and the specific way is the same as that of the first capillary needle 24; the third capillary needle 26 rotates to the second working position, the cell extraction substance is ionized and sent to the mass spectrometer 12 for analysis, and the mode is the same as the mass spectrum sample introduction of the second capillary needle 25;
the holder 41 continues to rotate forward by 120 degrees about the first rotating shaft, the first capillary 24 rotates to the first working position, at the same time, the second capillary 25 rotates to the second working position, the third capillary 26 rotates to the third working position,
it can be seen that, for each rotation of the holder 41, the same capillary needle (the first capillary needle 24, the second capillary needle 25, or the third capillary needle 26) undergoes one working cycle, that is, in the first working position, the second working position, and the third working position in sequence; at the same time, only one capillary needle is in an extraction state, only one capillary needle is in a mass spectrum sample injection state, and only one capillary needle is in a cleaning state;
if the signal output by the mass spectrum is abnormal, the bracket 41 rotates in the reverse direction for 120 degrees, the capillary needle in the mass spectrum sampling is rotated to the first working position, the cell is extracted again, and then the capillary needle rotates in the forward direction and enters the second working position until the signal output by the mass spectrum is normal.
Example 3:
according to the application example of the single-cell mass spectrometry system and the method based on the parallel processing technology in the embodiment 1 of the invention, the difference from the embodiment 2 is that:
as shown in fig. 2, there are two groups of working units, each group of working units includes three working units, the first group of working units includes a first working unit 21, a second working unit 22 and a third working unit 23, the second group of working units includes a fourth working unit 27, a fifth working unit 28 and a sixth working unit 29, the six working units of the two groups of working units are uniformly arranged in the whole circumference of the bracket 41, and the corresponding central angles of the two adjacent working units are 60 degrees; corresponding to the two groups of working units, the first cell bearing unit 113 and the second cell bearing unit 141 are respectively positioned at a first working position, the first mass spectrometer 121 and the second mass spectrometer 151 are respectively positioned at a second working position, and the first cleaning unit 131 and the second cleaning unit 161 are respectively positioned at a third working position;
when the bracket 41 rotates one circle counterclockwise, the same working unit undergoes a secondary working cycle and is sequentially located at a first working position, a second working position, a third working position, a first working position, a second working position and a third working position; at the same time, two capillary needles are in an extraction state, two capillary needles are in a mass spectrum sample injection state, and two capillary needles are in a cleaning state; a first cell carrying unit 113 and a second cell carrying unit 141 are respectively disposed at each first working position, a first mass spectrometer 121 and a second mass spectrometer 151 are disposed at each second working position, and a first cleaning unit 131 and a second cleaning unit 161 are respectively disposed at each third working position; the working unit is changed to the working position every time the bracket 41 rotates 60 degrees counterclockwise.
Example 4:
according to the application example of the single-cell mass spectrometry system and method based on the parallel processing technology in the embodiment 1 of the invention, the difference from the embodiment 3 is that:
as shown in fig. 3, there are two groups of work units, each group of work units including three work units, the first group of work units including a first work unit 21, a second work unit 22, and a third work unit 23, and the second group of work units including a fourth work unit 27, a fifth work unit 28, and a sixth work unit 29; the three working units of each group of working units are uniformly arranged in the whole circumferential direction of the bracket 41, the central angle corresponding to two adjacent working units in each group is 120 degrees, that is, any working unit in the second group of working units is positioned between the adjacent working units of the first group of working units, and the central angle between the working units of the adjacent first group of working units and the working units of the second group of working units is less than 120 degrees, such as 30 degrees, 45 degrees, 60 degrees and 90 degrees; that is, the six working units are uniformly or non-uniformly arranged on the entire circumference of the holder 41;
when the support 41 rotates 120 degrees clockwise, the working units in each group of working units are changed to working positions, but the working units in each group are not changed, for example, the working units in the first group of working units are not changed to the working positions corresponding to the second group of working units.
It can be seen that, for each rotation of the support 41, the same capillary needle undergoes a working cycle, i.e. is in the first working position, the second working position and the third working position in sequence; at the same time, two capillary needles are in an extraction state, two capillary needles are in a mass spectrum sample injection state, and two capillary needles are in a cleaning state.
The above embodiment shows two working units, but there may be more working units, such as 3 or 4, and more cell carrying units, mass spectrometers and cleaning units are needed.
Claims (10)
1. The single-cell mass spectrometry system based on the parallel processing technology comprises a cell bearing unit, a cleaning unit and a mass spectrometer, wherein the mass spectrometer is provided with a sample inlet; the single-cell mass spectrometry system based on the parallel processing technology is characterized by further comprising:
a base having a first rotational axis;
the bracket is connected with the first rotating shaft and rotates around the first rotating shaft;
m groups of working units, wherein M is an integer not less than 1, each group of working units comprises P working units, and P is an integer not less than 3; the working unit is arranged on the bracket and comprises a mechanical arm and a capillary needle, and the capillary needle is arranged on the mechanical arm; in the rotation of the bracket, when the first working unit in any group of working units is in the first working position, the second working unit is in the second working position, and the third working unit is in the third working position; the cell bearing unit is arranged at each first working position, the mass spectrometer is arranged at each second working position, and the cleaning unit is arranged at each third working position.
2. The parallel processing technology-based single-cell mass spectrometry system of claim 1, wherein the individual working units in any group of working units are arranged uniformly throughout the circumference of the rack; the cell bearing unit, the mass spectrometer and the cleaning unit are arranged in the following mode: when the support rotates 360 degrees, any working unit only goes through one working cycle, and the working cycle comprises a first working position, a second working position and a third working position which are sequentially arranged.
3. The parallel processing technology-based single-cell mass spectrometry system of claim 1, wherein the groups of working units are sequentially and uniformly arranged throughout the circumference of the rack; the cell bearing unit, the mass spectrometer and the cleaning unit are arranged in the following mode: when the support rotates 360 degrees, any work unit sequentially goes through work cycles of N times, N = M, and N is greater than 1, the work cycles include being in first work position, second work position and third work position in proper order.
4. The parallel processing technology-based single-cell mass spectrometry system of claim 1, wherein the robotic arm has a second rotational axis, and a capillary needle is connected to the second rotational axis; and the second rotating shafts of the mechanical arms in any group of working units form the same included angle with the radial direction of the support.
5. The parallel processing technology-based single-cell mass spectrometry system of claim 1, further comprising:
and the suction pump is communicated with the inside of the capillary needle through the gas channel.
6. The parallel processing technology-based single-cell mass spectrometry system of claim 5, further comprising:
a fixed tube, the capillary needle being disposed within the fixed tube; the gas channel passes through the fixed pipe;
an electrode disposed within the capillary needle.
7. The parallel processing technology-based single-cell mass spectrometry system of claim 1, further comprising:
the driving unit drives the cell bearing unit to translate two-dimensionally on a horizontal plane, so that the capillary needle selectively extracts single cells on the cell bearing unit, and the cell bearing unit comprises cell bearing positions which are distributed in a matrix form and are isolated from each other;
an imaging unit for imaging the cell-bearing site and the upper side of the capillary needle.
8. The parallel processing technology-based single-cell mass spectrometry system of claim 1, wherein the cleaning unit comprises:
a first container having an outlet at a bottom thereof; when the capillary needle rotates, the bottom end opening of the capillary needle is positioned at the lower side of the outlet;
and the opening of the second container faces upwards and is arranged at the lower sides of the first container and the bottom end of the capillary needle.
9. A single-cell mass spectrometry method using the parallel processing technology-based single-cell mass spectrometry system of any one of claims 1 to 8, the single-cell mass spectrometry method comprising:
the bracket rotates, so that when the first working unit in any group of working units is positioned at the first working position to extract single cells, the second working unit which extracts the single cells is positioned at the second working position to realize mass spectrum sample introduction, and the third working unit after the mass spectrum sample introduction is positioned at the third working position to realize cleaning;
through rotation, the same working unit is in a single cell extraction state, a mass spectrum sample introduction state and a cleaning state in sequence.
10. The method of mass spectrometry of single cell as claimed in claim 9, wherein in the rotation of the support, M groups of working units work simultaneously, wherein M working units are in single cell extraction state, M working units are in mass spectrometry sample introduction state, M working units are in cleaning state, and M is an integer no less than 1.
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| CN114113443A (en) * | 2021-11-22 | 2022-03-01 | 杭州谱育科技发展有限公司 | System and method for on-line continuous monitoring of aldehyde ketone compounds in gas |
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