Single cell analysis device and method based on mass spectrometry

Abstract

The present invention belongs to the field of cell analysis, and specifically relates to a single cell analysis device and method based on mass spectrometry, wherein the single cell analysis device comprises a first flow cell and an optical detection unit, wherein the optical detection unit comprises a light source, a detector and an analysis module; the pretreatment unit comprises a second flow cell and a vibration module, wherein the vibration module is arranged against the outer wall of the second flow cell, so that the cells in the second flow cell are arranged and flow in sequence along the central axis of the second flow cell, and the outlet of the second flow cell is connected to the inlet of the first flow cell; the nebulizer is used to nebulize the output cells of the first flow cell and send them to the mass spectrometry analysis unit; the control unit is used to control the operation of the mass spectrometry analysis unit according to the detection result of the analysis module. The present invention has the advantages of no need for sheath fluid, etc., and can be applied to flow single cell analysis.

Description

Technical Field

[0001] The present invention relates to single cell analysis, and in particular to a single cell analysis device and method based on mass spectrometry. Background Art

[0002] In single-cell analysis, flow cytometry can help people understand and recognize the complex composition of samples by fluorescently staining antigens on cells and analyzing the parameters of a large number of cells in a short period of time. Medical diagnoses can be made based on flow cytometry results to assist in the treatment of diseases. However, as the number of target parameters increases, the defects of fluorescent staining also become apparent. Common problems such as spectral leakage and autofluorescence greatly limit the results of flow cytometry.

[0003] Mass spectrometry flow cytometry can introduce isotope labels, use ICP-MS to vaporize cells, and obtain flow analysis results through stable isotope labels. The mass spectrometry flow cytometry method solves the problem of spontaneous fluorescence in traditional flow cytometry, and the overall solution is simple. However, at the same time, mass spectrometry flow cytometry has a small flux, slow injection speed, high sample requirements, a limited number of labels, and cannot perform cell analysis through forward and side scatter signals, which is different from traditional flow cytometry results. At the same time, mass spectrometry flow cytometry generally analyzes single cells, and non-single cell events generated during the injection process are invalid time. This type of data processing takes up a lot of transmission space and computing space, limiting the efficiency of mass spectrometry flow cytometry.

[0004] The full-spectrum flow cytometry method also solves the compensation problem in traditional flow cytometry, avoids spectral loss, and can achieve multi-color analysis. Unlike mass spectrometry flow cytometry, full-spectrum flow cytometry relies on the stability of sample injection. Through fluid focusing, each cell passes through the optical detection point and excites the spectral signal for analysis. If the spectral mass spectrometry method is combined, the optical signal and metal isotope labeling can be used to analyze cells at the same time, greatly improving the flow analysis capability. However, spectral flow cytometry uses a large amount of high-speed flowing sheath fluid, which cannot be applied to mass spectrometry flow cytometry. Summary of the invention

[0005] In order to solve the deficiencies in the above-mentioned prior art solutions, the present invention provides a single cell analysis device based on mass spectrometry and spectroscopy.

[0006] The objective of the present invention is achieved through the following technical solutions: A single cell analysis device based on mass spectrometry and spectroscopy, comprising a first flow cell and an optical detection unit, wherein the optical detection unit comprises a light source, a detector and an analysis module; the single cell analysis device also comprises: A pretreatment unit, the pretreatment unit comprising a second flow cell and a vibration module, the vibration module being arranged close to the outer wall of the second flow cell so that the cells in the second flow cell are sequentially arranged and flow along the central axis of the second flow cell, and the outlet of the second flow cell is connected to the inlet of the first flow cell; A nebulizer and a mass spectrometry unit, wherein the nebulizer is used to nebulize the cells output from the first flow cell and send them to the mass spectrometry unit; A control unit is used to control the operation of the mass spectrometry analysis unit according to the detection result of the analysis module.

[0007] The present invention also aims to provide a single cell analysis method using the above-mentioned single cell analysis device, and the present invention aims to be achieved through the following technical solutions: The single cell analysis method includes the following steps: (A1) The liquid containing cells enters the second flow cell; (A2) the vibration module emits mechanical waves to drive the cells in the second flow cell to move to the central axis of the second flow cell, to be arranged one by one and to move forward; (A3) The cells enter the first flow cell in sequence, the excitation light emitted by the light source is incident on the single cell in the first flow cell, the detector receives the scattered light and fluorescence, and the analysis module outputs the detection results of the cells; (A4) The cells discharged from the first flow cell are aerosolized; (A5) The control unit adjusts the operation of the mass spectrometry unit according to the detection result.

[0008] Compared with the prior art, the present invention has the following beneficial effects: 1. The cells are driven by the pretreatment unit, avoiding the use of sheath fluid, realizing the combination of spectroscopy and mass spectrometry, and bringing into play the respective advantages of the two methods; 2. A linkage between spectroscopy and mass spectrometry has been established, and the work of mass spectrometry is controlled by the detection results of spectroscopy. For example, if cell abnormalities are identified by spectroscopy, the ion analysis in mass spectrometry can be stopped. This significantly reduces the burden on the mass spectrometry analysis unit, such as transmission pressure and calculation pressure, and correspondingly improves work efficiency, which can further increase throughput. BRIEF DESCRIPTION OF THE DRAWINGS

[0009] The disclosure of the present invention will become easier to understand with reference to the accompanying drawings. It is easy for those skilled in the art to understand that these drawings are only used to illustrate the technical solution of the present invention and are not intended to limit the scope of protection of the present invention. In the drawings: Figure 1 It is a schematic diagram of the structure of the single cell analysis device based on mass spectrometry and spectroscopy of the present invention; Figure 2 It is a schematic diagram of the structure of the circulation pool of the present invention.

[0010] In the accompanying drawings, 11-sample pool, 12-delivery pump, 21-pretreatment unit, 211-second circulation pool, 212-first circulation pool, 213-connecting part, 214-vibration module, 31-nebulizer, 41-light source, 42-detector, 43-analysis module, 51-mass spectrometry unit, 61-control unit. DETAILED DESCRIPTION

[0011] Figure 1-Figure 2 The following description describes optional specific embodiments of the present invention to teach those skilled in the art how to implement and reproduce the present invention. In order to teach the technical solution of the present invention, some conventional aspects have been simplified or omitted. Those skilled in the art will understand that variations or replacements derived from these specific embodiments will be within the scope of the present invention. Those skilled in the art will understand that the following features can be combined in various ways to form multiple variations of the present invention. Thus, the present invention is not limited to the following optional specific embodiments, but is only limited by the claims and their equivalents.

[0012] Example 1.

[0013] A single cell analysis device based on mass spectrometry and spectroscopy according to an embodiment of the present invention is as follows: Figure 1 As shown, including: The first circulation pool 212 and the optical detection unit include a light source 41 , a detector 42 and an analysis module 43 .

[0014] Preprocessing unit 21, such as Figure 2 As shown, the pretreatment unit 21 includes a second circulation pool 211 and a vibration module 214. The vibration module 214 is arranged close to the outer wall of the second circulation pool 211, so that the cells in the second circulation pool 211 are arranged and flow in sequence along the central axis of the second circulation pool 211, and the outlet of the second circulation pool 211 is connected to the inlet of the first circulation pool 212.

[0015] A nebulizer 31 and a mass spectrometer analysis unit 51 , wherein the nebulizer 31 is used to nebulize the cells output from the first circulation pool 212 and send the cells to the mass spectrometer analysis unit 51 .

[0016] The control unit 61 is used to control the operation of the mass spectrometry analysis unit 51 according to the detection result of the analysis module 43.

[0017] In order to reduce the working pressure of the mass spectrometry analysis unit 51, further, the working mode of the control unit 61 is: If the detection result of the analysis module 43 is abnormal, the mass spectrometry analysis unit 51 stops ion analysis. The abnormality includes multiple cells entering at the same time or cell debris.

[0018] In order to better drive cells and complete spectral detection, further, the cross-section of the second circulation pool 211 perpendicular to the central axis direction thereof is a ring, and the cross-section of the first circulation pool 212 perpendicular to the central axis direction thereof is a rectangle.

[0019] In order to improve working stability, further, the outlet of the second circulation pool 211 and the inlet of the first circulation pool 212 are connected through a connecting portion 213, and the inner cross-section of the connecting portion 213 transitions from a circle to a rectangle.

[0020] The single cell analysis method of the embodiment of the present invention, that is, the working method of the single cell analysis device of the embodiment of the present invention, comprises the following steps: (A1) Liquid containing cells enters the second circulation cell 211 .

[0021] (A2) The vibration module 214 emits mechanical waves to drive the cells in the second flow cell 211 to move to the central axis of the second flow cell 211, and to be arranged and moved forward one by one.

[0022] (A3) The cells enter the first circulation pool 212 in sequence, the excitation light emitted by the light source 41 is incident on the single cells in the first circulation pool 212, the detector 42 receives the scattered light and fluorescence, and the analysis module 43 outputs the detection results of the cells.

[0023] (A4) The cells discharged from the first flow cell 212 are atomized.

[0024] (A5) The control unit 61 adjusts the operation of the mass spectrometry unit 51 according to the detection result.

[0025] In order to reduce the working pressure of the mass spectrometry analysis unit 51, further, the working mode of the control unit 61 is: If the detection result is abnormal, the mass spectrometry analysis unit 51 stops ion analysis; the abnormality includes multiple cells entering at the same time or cell fragments.

[0026] If the detection result is normal, the mass spectrometry analysis unit 51 performs ion analysis.

[0027] Example 2.

[0028] An application example of a single cell analysis device and method based on mass spectrometry in Example 1 of the present invention.

[0029] In this application example, Figure 1As shown, the delivery pump 12, the first circulation pool 212, the pretreatment unit 21 and the atomizer 31 are connected in sequence.

[0030] The light source 41 and the detector 42 are arranged on the side of the first circulation pool 212 , and the detection result output by the analysis module 43 is sent to the control unit 61 .

[0031] The mass spectrometry unit 51 adopts ICP-MS, in which the torch is arranged vertically, for ionization of single cells after atomization. The mass spectrometry part includes an ion deflection module, an ion transmission module and a detector, wherein the ion deflection module is used to deflect ions in the vertical direction by 90 degrees, and the ions enter the transmission module.

[0032] like Figure 2 As shown, the cross section of the second circulation pool 211 perpendicular to the central axis direction thereof is a circular ring, and the vibration module 214 is made of piezoelectric ceramics and is attached to the outer wall of the second circulation pool 211. The cross section of the first circulation pool 212 perpendicular to the central axis direction thereof is a rectangular shape. The connecting portion 213 connects the outlet of the second circulation pool 211 and the inlet of the first circulation pool 212, and the inner cross section of the connecting portion 213 transitions from a circular shape to a rectangular shape, so that the first circulation pool 212 and the second circulation pool 211 are connected together.

[0033] The single cell analysis method of the embodiment of the present invention, that is, the working method of the single cell analysis device of the embodiment of the present invention, comprises the following steps: (A1) Under the suction of the delivery pump 12 , the liquid containing cells in the sample pool 11 enters the second circulation pool 211 .

[0034] (A2) The vibration module 214 emits mechanical waves, which propagate and reflect in the second circulation pool 211, driving the cells in the liquid in the second circulation pool 211 to move to the central axis of the second circulation pool 211, and to be arranged and move forward one by one.

[0035] (A3) The cells pass through the connection part 213 and enter the first circulation pool 212 in sequence. The excitation light emitted by the light source 41 is incident on the single cells in the first circulation pool 212. The detector 42 receives the scattered light and fluorescence. The analysis module 43 outputs the detection results of the cells. The specific analysis process is the existing technology in this field.

[0036] (A4) Cells exiting the first flow cell 212 are nebulized and ionized by the vertical torch.

[0037] (A5) The control unit 61 adjusts the operation of the mass spectrometry unit 51 according to the detection result, specifically in the following manner: If the detection result is abnormal, such as multiple cells entering at the same time, or cell fragments, the mass spectrometry analysis unit 51 stops the operation of the ion deflection module, and no ions enter the downstream ion transmission module, that is, the ion analysis is stopped.

[0038] If the detection result is normal, the mass spectrometry unit 51 performs ion analysis. Specifically, the ions pass vertically through the sampling cone and then are horizontally transmitted through the ion deflection module. Neutral particles are separated due to inertia and deflected ions.

Claims

1. A single cell analysis device based on mass spectrometry, comprising a first flow cell and an optical detection unit, wherein the optical detection unit comprises a light source, a detector and an analysis module; characterized in that: The single cell analysis device further comprises: A pretreatment unit, the pretreatment unit comprising a second flow cell and a vibration module, the vibration module being arranged close to the outer wall of the second flow cell so that the cells in the second flow cell are sequentially arranged and flow along the central axis of the second flow cell, and the outlet of the second flow cell is connected to the inlet of the first flow cell; A nebulizer and a mass spectrometry unit, wherein the nebulizer is used to nebulize the cells output from the first flow cell and send them to the mass spectrometry unit; A control unit is used to control the operation of the mass spectrometry analysis unit according to the detection result of the analysis module.

2. The single cell analysis device based on mass spectrometry according to claim 1, characterized in that: The working mode of the control unit is: If the detection result of the analysis module is abnormal, the mass spectrometry analysis unit stops ion analysis.

3. The single cell analysis device based on mass spectrometry according to claim 2, characterized in that: Such abnormalities include simultaneous entry of multiple cells or cellular fragments.

4. The single cell analysis device based on mass spectrometry according to claim 1, characterized in that: The cross section of the second flow cell perpendicular to the central axis direction thereof is a circular ring, and the cross section of the first flow cell perpendicular to the central axis direction thereof is a rectangular shape.

5. The single cell analysis device based on mass spectrometry according to claim 4, characterized in that: The outlet of the second circulation cell and the inlet of the first circulation cell are connected via a connecting portion, and the inner cross-section of the connecting portion transitions from a circle to a rectangle.

6. The single cell analysis device based on mass spectrometry according to claim 1, characterized in that: The mass spectrometry unit adopts ICP-MS.

7. The single cell analysis method implemented by the single cell analysis device based on mass spectrometry according to claim 1, comprising the following steps: (A1) The liquid containing cells enters the second flow cell; (A2) the vibration module emits mechanical waves to drive the cells in the second flow cell to move to the central axis of the second flow cell, to be arranged one by one and to move forward; (A3) The cells enter the first flow cell in sequence, the excitation light emitted by the light source is incident on the single cell in the first flow cell, the detector receives the scattered light and fluorescence, and the analysis module outputs the detection results of the cells; (A4) The cells discharged from the first flow cell are aerosolized; (A5) The control unit adjusts the operation of the mass spectrometry unit according to the detection result.

8. The single cell analysis method according to claim 7, characterized in that The working mode of the control unit is: If the detection result is abnormal, the mass spectrometry unit stops ion analysis; If the detection result is normal, the mass spectrometry unit performs ion analysis.

9. The single cell analysis method according to claim 8, characterized in that Such abnormalities include simultaneous entry of multiple cells or cellular fragments.

10. The single cell analysis method according to claim 7, characterized in that: In step (A4), the nebulized cells are ionized.

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