Fully automatic single cell mass spectrometry sample injector

Abstract

The present invention discloses a fully automatic single-cell mass spectrometry detection injector, comprising a fixed frame, an extraction system arranged on the fixed frame, a hollow nanoprobe arranged on the extraction system, a workbench arranged at the lower end of the fixed frame, a single-cell sample below the hollow nanoprobe, a fluorescence spectrum collection and analysis system arranged on the workbench of the fixed frame below the single-cell sample, an inverted fluorescence microscope arranged on the hollow nanoprobe, and the hollow nanoprobe is also connected to the injector. The present invention extracts substances in a single cell, such as cytoplasm, cell nucleus, etc., and then automatically sends them to the injector of a mass spectrometer for component analysis. Automatic sampling and transmission of samples to the mass spectrometer injection port can greatly reduce the operation time, ensure the sample quality, and improve the detection effect.

Description

Technical Field

[0001] The invention relates to a cell mass spectrometry detection sample injector, belonging to the technical field of biological detection. Background Art

[0002] Currently, single-cell mass spectrometry can only be performed by manually taking samples on a microscope and then sending them to the mass spectrometer injection port. This has the disadvantages of complex operation, long operation time, and easy damage to the sampling needle. It is especially not suitable for the injection of volatile samples. Summary of the invention

[0003] The technical problem to be solved by the present invention is to provide a safe and reliable fully automatic single cell mass spectrometry detection injector.

[0004] The technical solution of the present invention is:

[0005] A fully automatic single-cell mass spectrometry detection injector comprises a fixed frame, an extraction system is arranged on the fixed frame, a hollow nanoprobe is arranged on the extraction system, a workbench is arranged at the lower end of the fixed frame, a single-cell sample is placed on the workbench below the hollow nanoprobe, an inverted fluorescence microscope is arranged below the single-cell sample on the workbench, and the hollow nanoprobe is also connected to the injector.

[0006] The beneficial effects of the present invention are:

[0007] Extracting substances from a single cell: The hollow nanoneedle (minimum diameter 30nm) is precisely positioned inside the cell, such as the cytoplasm, nucleus, etc., through a fixed frame (accuracy 200nm), and then the substance is extracted through the extraction system. Other products cannot achieve this function;

[0008] Automatic transmission of extracted trace samples: The extraction system is equipped with an automatic transmission arm that can extend horizontally and rotate 90 degrees to send the sample to the mass spectrometer injection port for component analysis. The injection time for a single sample is about 30 seconds.

[0009] Good compatibility: This automatic injection system adopts a separate movable fixing system, and the injection needle can be adjusted in a wide range of distance and height. It can be used with various mass spectrometers, has excellent compatibility, and reduces the customer's use cost;

[0010] The present invention extracts substances in a single cell, such as cytoplasm, cell nucleus, etc., and then automatically sends them to the injection port of the mass spectrometer for component analysis. Automatic sampling and transmission of samples to the mass spectrometer injector can greatly reduce operation time, ensure sample quality, and improve detection effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1 It is a schematic diagram of the structure of the present invention;

[0012] Figure 2 It is a partially enlarged mass spectrum in an embodiment of the present invention;

[0013] Figure 3 It is the overall detection mass spectrum in the embodiment of the present invention.

[0014] In the figure: 1—fixed frame, 2—hollow nanoprobe, 3—single cell sample, 4—injector, 5—extraction system, 6—inverted fluorescence microscope. DETAILED DESCRIPTION

[0015] The present invention will be further described below in conjunction with specific drawings and embodiments.

[0016] A fully automatic single-cell mass spectrometry detection injector comprises a fixed frame 1, an extraction system 5 is arranged on the fixed frame 1, a hollow nanoprobe 2 is arranged on the extraction system 5, a workbench is arranged at the lower end of the fixed frame 1, a single-cell sample 3 is placed on the workbench below the hollow nanoprobe 2, an inverted fluorescence microscope 6 is arranged below the single-cell sample 3 on the workbench, and the hollow nanoprobe 2 is also connected to the injector 4.

[0017] When working, the extraction system 5 drives the hollow nanoprobe 2 to move up and down on the column of the fixed frame 1, adjusts the upper and lower distances between the hollow nanoprobe 2 and the single cell sample 3, and can also adjust the front and back and left and right positions. After the hollow nanoprobe 2 sucks the single cell sample 3 into the pinhole, it is rotated and transferred to the injector 4 for component analysis. The material in a single cell can be cytoplasm or cell nucleus. Automatic sampling and transmission of samples to the mass spectrometer injector 4 can greatly reduce the operation time, ensure the sample quality, and improve the detection effect.

[0018] The fixing frame 1 is produced by Jiangsu Ruiming Biological Company, model: GD02; the hollow nanoneedle 2 is produced by Jiangsu Ruiming Biological Company, model: RMA-PH; the single cell sample 3 can be produced by Cobioer Company, item number CBP60232; the sampler 4 is produced by Jiangsu Ruiming Biological Company, model: YTQ300; the extraction system 5 is produced by Jiangsu Ruiming Biological Company, model: FIS300; the inverted fluorescence microscope 6 is Olympus, model: CKX31.

[0019] Example:

[0020] Experimental materials: The samples are HeLa cells (human cervical cancer cells, produced by Cobioer, product number CBP60232),

[0021] Fixed frame 1: minimum adjustment accuracy 200nm, left and right adjustment distance range: 1mm-500mm, up and down adjustment distance range: 20mm-15000mm, angle adjustment range: 15 degrees-120 degrees; hollow nanoneedle 2: minimum diameter 30nm; injector 4: injection port inner diameter 1mm; extraction system 5: minimum extraction volume 15fL; inverted fluorescence microscope 6: can realize online fluorescence imaging function, minimum optical resolution is 1um; mass spectrometer: TransMIT, Germany, model: AP-SMALDI 10

[0022] Experimental process:

[0023] The sample cell 3 is attached to the bottom of the culture dish (before the experiment, the culture dish is filled with phosphate buffered saline solution - PBS, and placed in a carbon dioxide incubator and cultured at 20 degrees for 10 hours), and the culture dish is located in the middle of the stage of the inverted fluorescence microscope; the hollow nanoneedle 2 is moved to the top position of the cell through the horizontal axis (X axis) and the longitudinal axis (Z axis) of the fixed frame 1; the Z axis of the fixed frame is adjusted to insert the hollow nanoneedle 2 into the cell to a depth of about 100nm (the depth can be controlled by micromanipulation); it reaches the area of ​​the substance to be extracted; the extraction system 5 is started, the extraction working voltage is adjusted to 3V, the extraction time is adjusted to 5 seconds, and the extraction begins; after the extraction is completed, the fixed frame moves the probe upward (Z axis) for 300mm, then rotates 90 degrees counterclockwise, and finally moves 450mm horizontally, at which time the tip of the hollow probe has entered the sample port of the injector by 200mm; the injector starts to work, sucks out the trace substances in the hollow needle through the vacuum interface and sends them to the sample chamber of the mass spectrometer; the mass spectrometer starts to analyze the sample.

[0024] The experimental results are as follows:

[0025] Through mass spectrometry analysis, the various material components in the cells can be clearly analyzed. Figure 3 is the overall detection spectrum (0-280), Figure 2 This is a local enlarged spectrum (75.7-154), the six material components are Figure 2 It is marked with I-VI.

Claims

1. A fully automatic single cell mass spectrometry sample injector, comprising a fixed frame (1), characterized in that: The fixing frame (1) is provided with an extraction system (5), the extraction system (5) is provided with a hollow nanoprobe (2), a workbench is provided at the lower end of the fixing frame (1), a single cell sample (3) is provided on the workbench below the hollow nanoprobe (2), an inverted fluorescence microscope (6) is provided below the single cell sample (3) on the workbench, and the hollow nanoprobe (2) is also connected to a sample injector (4); The fixing frame has an accuracy of 200nm, the minimum diameter of the hollow nanoneedle is 30nm, and the inner diameter of the injection port of the injector (4) is 1mm; the extraction system (5) has a minimum extraction volume of 15fL; the inverted fluorescence microscope (6) can realize online fluorescence imaging function, and the minimum optical resolution is 1um; The detection method is as follows: the sample cell (3) is attached to the bottom of the culture dish. Before the experiment, the culture dish is filled with phosphate buffered saline solution (PBS) and placed in a carbon dioxide incubator for 10 hours at 20 degrees. The culture dish is located in the middle of the stage of an inverted fluorescence microscope. The hollow nanoneedle (2) is moved to the top of the cell by the horizontal X-axis and the vertical Z-axis of the fixed frame (1). The Z-axis of the fixed frame is adjusted to insert the hollow nanoneedle (2) into the cell to a depth of about 100nm to reach the area of ​​the substance to be extracted. The extraction system (5) is started, the extraction working voltage is adjusted to 3V, the extraction time is adjusted to 5 seconds, and the extraction is started. After the extraction is completed, the fixed frame moves the probe upward by 300mm, then rotates 90 degrees counterclockwise, and finally moves 450mm horizontally. At this time, the tip of the hollow probe has entered the sample port of the injector by 200mm. The injector starts to work, sucks out the trace substances in the hollow needle through the vacuum interface and sends them to the sample chamber of the mass spectrometer. The mass spectrometer starts to analyze the sample.

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