A flow cytometer liquid path system based on positive pressure sample loading and a control method thereof

Abstract

The application discloses a flow cytometer liquid path system based on positive pressure sample loading and a control method thereof, and relates to the field of flow cytometry detection. The sample loading focusing is achieved by positive pressure driving of sheath liquid by a quantitative pump and positive pressure driving of a sample by a plunger pump. The flow cytometer liquid path system comprises a sheath liquid barrel, a cleaning liquid barrel, a shutdown liquid barrel, a waste liquid barrel, a quantitative pump, a plunger pump, a diaphragm pump, a capsule filter, a cleaning swab, a sample needle and a flow cell. The bottom of the cleaning swab is provided with an inlet, the top of the cleaning swab is provided with an outlet, and the cleaning swab is provided with a cleaning channel for the sample needle to pass through. The flow cell is provided with a sheath liquid inlet on one side and a sheath liquid outlet on the other side, and the bottom of the flow cell is provided with a sample inlet and the top of the flow cell is provided with a sample outlet. The bottom end of the capsule filter is provided with an inlet, the top end of the capsule filter is provided with an outlet, and the side of the capsule filter is further provided with a side outlet. The quantitative pump and the plunger pump are used for positive pressure sample loading, the quantitative pump can realize finer subdivision, the liquid control precision is higher, and the pulsation problem driven by the peristaltic pump is avoided.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of flow cytometry, in particular to a flow cytometer liquid path system based on positive pressure sample loading. BACKGROUND

[0002] Flow cytometry is a device for automatic analysis of cells or micrometer-sized particles. It can quickly measure, store and display a series of important biophysical and biochemical parameters of dispersed cells or particles suspended in a liquid. It is currently increasingly used in clinical applications and is known as cell CT. The existing flow cytometer liquid path generally uses a peristaltic pump, a plunger pump or positive pressure for sample injection, and a sheath liquid is also driven by a peristaltic pump.

[0003] However, the existing technology has the following problems: the peristaltic pump is pulse-driven, so whether it is a sample or a sheath liquid, it will cause pulsation of the liquid flow. This pulsation will cause fluctuations in the detection results when passing through the laser focusing area, thereby affecting the measurement results. The service life of the peristaltic pump tube is very short and needs to be replaced regularly, usually every six months, which is high in maintenance cost. Even if the peristaltic pump tube is within its service life, slight deformation of the peristaltic pump tube will still cause changes in the liquid flow, thereby causing changes in the liquid flow rate and ultimately resulting in fluctuations in the results. When the temperature changes, the density and viscosity of the sheath liquid will change slightly, and the flow rate of the peristaltic pump is related to the viscosity and density of the liquid, so temperature fluctuations will cause slight fluctuations in the flow rate.

[0004] In addition, air bubbles often cannot be discharged in the liquid path system, which requires disassembly of the pipeline to discharge the air bubbles, and the air bubbles cannot be monitored and discharged in real time. The flow rate of the plunger pump generally deviates from the actual flow rate, and when performing absolute counting by volume, the measurement volume will not be accurate. At the same time, ordinary flow cytometers only use a sheath liquid tank and a waste liquid tank and do not have a multi-liquid maintenance liquid path system, which requires longer maintenance time and higher labor maintenance cost. SUMMARY

[0005] The present application proposes a flow cytometer liquid path system based on positive pressure sample loading and a control method thereof to solve the above problems. The sheath liquid is driven by a quantitative pump and the sample is driven by a plunger pump under positive pressure for sample loading and focusing.

[0006] The technical scheme of the present application is as follows: the flow cytometer liquid path system comprises a sheath liquid tank 1, a cleaning liquid tank 2, a shutdown liquid tank 3, a waste liquid tank 4, a quantitative pump 5, a plunger pump 7, a diaphragm pump 14, a bladder filter 9, a cleaning swab 10, a sample needle 11 and a flow cell 13.

[0007] The bottom of the cleaning swab 10 is provided with an inlet, the top is provided with an outlet, and a cleaning channel is provided in the cleaning swab 10 for the sample needle 11 to enter and exit; one side of the flow cell 13 is provided with a sheath liquid inlet, the other side is provided with a sheath liquid outlet, and the bottom of the flow cell 13 is provided with a sample inlet, and the top is provided with a sample outlet; the bottom end of the capsule filter 9 is provided with an inlet, the top end is provided with an outlet, and the side is further provided with a side outlet;

[0008] The sheath liquid barrel 1, the cleaning liquid barrel 2, and the shutdown liquid barrel 3 are connected to the inlet of the quantitative pump 5, the outlet of the quantitative pump 5 is connected to the inlet of the plunger pump 7, the inlet of the cleaning swab 10, and the inlet of the capsule filter 9, the outlet of the capsule filter 9 is connected to the sheath liquid inlet of the flow cell 13, the plunger pump 7 is also connected to the sample needle 11 and the sample inlet of the flow cell 13, the sheath liquid outlet of the flow cell 13 and the outlet of the cleaning swab 10 are both connected to the inlet of the diaphragm pump 14, and the outlet of the diaphragm pump 14, the sample outlet of the flow cell 13, and the side outlet of the capsule filter 9 are all connected to the waste liquid barrel 4.

[0009] The flow cytometer liquid path system further comprises a first electromagnetic valve LV01, a second electromagnetic valve LV02, a third electromagnetic valve LV03, a fourth electromagnetic valve LV04, a fifth electromagnetic valve LV05, a sixth electromagnetic valve LV06, a seventh electromagnetic valve LV07, an eighth electromagnetic valve LV08, a ninth electromagnetic valve LV09, and a tenth electromagnetic valve LV10.

[0010] The first electromagnetic valve LV01, the second electromagnetic valve LV02, and the ninth electromagnetic valve LV09 are two-position three-way electromagnetic valves with two inlets and one outlet, and the third electromagnetic valve LV03 and the eighth electromagnetic valve LV08 are two-position three-way electromagnetic valves with one inlet and two outlets; when not powered on, they are in Figure 1 the position shown, and after being powered on, they are switched to the other position;

[0011] The fourth electromagnetic valve LV04, the fifth electromagnetic valve LV05, the sixth electromagnetic valve LV06, the seventh electromagnetic valve LV07, and the tenth electromagnetic valve LV10 are two-position two-way electromagnetic valves with one inlet and one outlet; when not powered on, they are in Figure 1 the off state shown, and after being powered on, they are switched to the other position, entering the on state;

[0012] The connections described in this paragraph are all achieved through pipelines: the sheath liquid barrel 1 and the cleaning liquid barrel 2 are respectively connected to the two inlets of the first electromagnetic valve LV01, the outlet of the first electromagnetic valve LV01 and the shutdown liquid barrel 3 are respectively connected to the two inlets of the second electromagnetic valve LV02, the outlet of the second electromagnetic valve LV02 is connected to the inlet of the quantitative pump 5, and the outlet of the quantitative pump 5 is connected to the inlet of the third electromagnetic valve LV03.

[0013] One of the outlets of the third electromagnetic valve LV03 is connected to the inlet of the cleaning swab 10, and the other outlet of the third electromagnetic valve LV03 is simultaneously connected to the inlets of the fourth electromagnetic valve LV04, the seventh electromagnetic valve LV07 and the capsule filter 9. The fourth electromagnetic valve LV04 is also connected to the plunger pump 7, and the seventh electromagnetic valve LV07 is also connected to the sheath liquid inlet of the flow cell 13. The outlet of the capsule filter 9 is connected to the sheath liquid inlet of the flow cell 13 through the sixth electromagnetic valve LV06, and the side outlet of the capsule filter 9 is connected to the waste liquid tank 4 through the fifth electromagnetic valve LV05.

[0014] The plunger pump 7 is connected to the inlets of the fourth electromagnetic valve LV04 and the eighth electromagnetic valve LV08. One of the outlets of the eighth electromagnetic valve LV08 is connected to the sample needle 11, and the other outlet is connected to the sample inlet of the flow cell 13.

[0015] The sample outlet of the flow cell 13 is connected to the waste liquid tank 4 through the tenth electromagnetic valve LV10. The sheath liquid outlet of the flow cell 13 and the outlet of the cleaning swab 10 are connected to the two inlets of the ninth electromagnetic valve LV09, respectively. The outlet of the ninth electromagnetic valve LV09 is connected to the waste liquid tank 4 through the diaphragm pump 14.

[0016] A pressure sensor 6 is connected between the quantitative pump 5 and the third electromagnetic valve LV03.

[0017] A bubble sensor 8 is connected between the third electromagnetic valve LV03 and the capsule filter 9.

[0018] A flow sensor 12 is connected between the eighth electromagnetic valve LV08 and the flow cell 13.

[0019] The control method includes a sample loading method, a general cleaning method and a reinforced cleaning method.

[0020] The sample loading method is as follows: first, the plunger pump 7 sucks a column of air through the idle sample needle 11, then the plunger pump 7 draws the sample through the sample needle 11 inserted into the sample pool, and then the sample is sent into the flow cell 13.

[0021] At the same time, the sheath liquid in the sheath liquid tank 1 is sent into the flow cell 13 through the capsule filter 9 by the quantitative pump 5. The sheath liquid and the sample enter the flow cell 13 at the same time, forming a stable sheath flow state, and then the sample flow information is collected.

[0022] The control method includes a general cleaning method, which includes a sample needle outer wall general cleaning method and a sample needle inner wall general cleaning method.

[0023] The general cleaning method for the outer wall of the sample needle is as follows: the sheath liquid is sent into the cleaning swab 10 by the quantitative pump 5, and is drawn away by the diaphragm pump 14, the sample needle 11 is inserted into the cleaning swab 10 and taken out, and the general cleaning of the outer wall of the sample needle 11 is completed.

[0024] The general cleaning method for the inner wall of the sample needle is as follows: the sheath liquid is sent into the sample needle 11 by the quantitative pump 5 through the plunger pump 7, and the sample needle 11 is kept above the cleaning swab 10, the waste liquid flowing out is drawn away by the diaphragm pump 14, and the general cleaning of the inner wall of the sample needle 11 is completed.

[0025] The control method includes a reinforced cleaning method, which is divided into two steps of preliminary cleaning of the cleaning liquid and re-cleaning of the sheath liquid.

[0026] In the first step, the sample needle and the flow cell are cleaned with the cleaning liquid respectively.

[0027] When cleaning the sample needle, the outer wall of the sample needle is preliminarily cleaned as follows: the cleaning liquid is sent into the cleaning swab 10 by the quantitative pump 5, and is drawn away by the diaphragm pump 14, in the process, the sample needle 11 is inserted into the cleaning swab 10 and taken out, and the preliminary cleaning of the outer wall of the sample needle 11 is completed.

[0028] Then the sample needle is refilled as follows: a test tube containing the cleaning liquid is taken and prepared; the plunger pump 7 is opened to draw a section of air through the idle sample needle 11, then the bottom of the sample needle 11 is placed in the test tube, the plunger pump 7 is opened again to draw a small amount of cleaning liquid through the sample needle, so that the cleaning liquid fills the inside of the sample needle 11.

[0029] The specific method for cleaning the flow cell with the cleaning liquid is as follows: the cleaning liquid is drawn from the cleaning liquid barrel 2 by the quantitative pump 5, and is sent into the flow cell 13 after bypassing the capsule filter, then is divided into two paths and enters the waste liquid barrel 4 from the sheath liquid outlet and the sample outlet of the flow cell 13 respectively, the quantitative pump 5 and the diaphragm pump 14 are closed to make the flow cell 13 full of cleaning liquid.

[0030] In the second step, the cleaning liquid is soaked in the sample needle and the flow cell for more than 5 minutes, and then the sample needle and the flow cell are washed with the sheath liquid respectively.

[0031] When the sample needle is cleaned with the sheath liquid, the outer wall of the sample needle is preliminarily cleaned as follows: the sheath liquid is sent into the cleaning swab 10 by the quantitative pump 5, and is drawn away by the diaphragm pump 14, in the process, the sample needle 11 is inserted into the cleaning swab 10 and taken out, and the reinforced cleaning process of the outer wall of the sample needle 11 is completed.

[0032] Then the inner wall of the sample needle is cleaned, specifically as follows: the cleaning liquid originally filled in the sample needle 11 is discharged through the plunger pump 7, then the sheath liquid is sent into the sample needle 11 through the quantitative pump 5, and the sample needle 11 is kept above the cleaning swab 10, the waste liquid is pumped away through the diaphragm pump 14, and the reinforced cleaning process of the inner wall of the sample needle 11 is completed.

[0033] The specific method for cleaning the flow cell with the sheath liquid is as follows: the cleaning liquid is pumped from the sheath liquid barrel 1 through the quantitative pump 5, and then sent into the flow cell 13 after bypassing the capsule filter, and then divided into two paths, respectively entering the waste liquid barrel 4 from the sheath liquid outlet and the sample outlet of the flow cell 13, and the reinforced cleaning process of the flow cell 13 is completed.

[0034] The control method comprises a shutdown liquid filling method, and the shutdown liquid filling method comprises a sample needle filling method and a flow cell filling method.

[0035] The sample needle filling method is as follows: the shutdown liquid in the shutdown liquid barrel 3 is sent into the sample needle 11 through the quantitative pump 5 and the plunger pump 7, and the sample needle 11 is kept above the cleaning swab 10, the waste liquid is pumped away through the diaphragm pump 14, and the sample needle 11 is filled with the shutdown liquid after the quantitative pump 5 and the diaphragm pump 14 stop.

[0036] The flow cell filling method is as follows: the shutdown liquid is pumped from the shutdown liquid barrel 3 through the quantitative pump 5, and then sent into the flow cell 13 after bypassing the capsule filter, and then divided into two paths, respectively entering the waste liquid barrel 4 from the sheath liquid outlet and the sample outlet of the flow cell 13, and the flow cell 13 is filled with the shutdown liquid by closing the quantitative pump 5 and the diaphragm pump 14.

[0037] The beneficial effects of the present application are as follows:

[0038] Firstly, the quantitative pump and the plunger pump are used for positive pressure sampling in the present application, the quantitative pump can realize more fine subdivision, the liquid control precision is higher, the pulsation problem driven by the peristaltic pump is avoided, and the maintenance problem caused by pipeline aging is avoided.

[0039] Secondly, the bubble sensor is used in the liquid path system to monitor the situation of bubbles appearing in the pipeline, so that the bubbles can be discharged in time and the pipeline leakage situation can be monitored.

[0040] Thirdly, the flow sensor is used in the liquid path system to monitor the actual sample flow, which is compared with the flow provided by the plunger pump, so that the actual flow compensation is completed, and the volume method absolute counting is realized.

[0041] Fourthly, the float sensor is used in the liquid path system to monitor the liquid level in the barrel, so that the liquid amount in the barrel can be maintained conveniently.

[0042] Five, the liquid system uses a pressure sensor to monitor the pipeline pressure overrun, facilitating timing, pipeline and device maintenance.

[0043] Six, the liquid system uses sheath liquid, cleaning liquid, shutdown liquid, realizes sample loading, cleaning, flushing and other maintenance. BRIEF DESCRIPTION OF DRAWINGS

[0044] Figure 1 is a structural schematic diagram of an embodiment of the liquid system of the present application.

[0045] In the drawings: 1-sheath liquid barrel, 1-1-float sensor, 2-cleaning liquid barrel, 3-shutdown liquid barrel, 4-waste liquid barrel, 5-dosing pump, 5-1-dosing pump cleaning channel, 6-pressure sensor, 7-plunger pump, 8-bubble sensor, 9-bag filter, 10-cleaning swab, 11-sample needle, 12-flow sensor, 13-flow cell, 14-diaphragm pump. EMBODIMENT

[0046] In order to clearly illustrate the technical features of the present patent, the present patent will be described in detail below through specific embodiments, and in conjunction with the accompanying drawings.

[0047] As shown in Figure 1 , it includes sheath liquid barrel 1, float sensor 1-1, cleaning liquid barrel 2, shutdown liquid barrel 3, waste liquid barrel 4, dosing pump 5, dosing pump cleaning channel 5-1, pressure sensor 6, plunger pump 7, bubble sensor 8, bag filter 9, cleaning swab 10, sample needle 11, flow sensor 12, flow cell 13, diaphragm pump 14, wherein the sheath liquid barrel 1 and the cleaning liquid barrel 2 are connected to the first electromagnetic valve, the first electromagnetic valve and the shutdown liquid barrel 3 are connected to the second electromagnetic valve, the second electromagnetic valve is communicated with the dosing pump 5, the dosing pump cleaning channel 5-1 is connected between the second electromagnetic valve and the cleaning liquid barrel 2, the dosing pump 5 and the third electromagnetic valve are communicated, the pressure sensor 6 is connected between the dosing pump 5 and the third electromagnetic valve, the third electromagnetic valve and the lower port of the cleaning swab 10 are communicated, the upper port of the cleaning swab 10 is communicated with the ninth electromagnetic valve, the ninth electromagnetic valve and the diaphragm pump 14 are communicated, and the diaphragm pump 14 is communicated with the waste liquid barrel 4.

[0048] Further, the third electromagnetic valve and the lower port of the plunger pump 7 are communicated, the fourth electromagnetic valve is arranged between the third electromagnetic valve and the lower port of the plunger pump 7, the upper port of the plunger pump 7 is communicated with the eighth electromagnetic valve, the eighth electromagnetic valve is communicated with the sample needle 11, and the sample needle 11 is arranged in the cleaning swab 10.

[0049] Further, the third electromagnetic valve and the left end liquid inlet of the flow cell 13 are communicated, the seventh electromagnetic valve is arranged between the third electromagnetic valve and the left end liquid inlet of the flow cell 13, the lower end liquid inlet of the flow cell 13 is communicated with the flow sensor 12, and the flow sensor 12 is communicated with the eighth electromagnetic valve.

[0050] Further, the third electromagnetic valve is communicated with the lower end liquid inlet of the capsule filter 9, the bubble sensor 8 is connected between the third electromagnetic valve and the lower end liquid inlet of the capsule filter 9, the upper end liquid outlet of the capsule filter 9 is communicated with the left end liquid inlet of the flow cell 13, the sixth electromagnetic valve is arranged between the upper end liquid outlet of the capsule filter 9 and the left end liquid inlet of the flow cell 13, and the right end liquid outlet of the flow cell 13 is communicated with the ninth electromagnetic valve.

[0051] Further, the side end liquid outlet of the capsule filter 9 is communicated with the waste liquid barrel 4, the fifth electromagnetic valve is arranged between the side end liquid outlet of the capsule filter 9 and the waste liquid barrel 4, the upper end liquid outlet of the flow cell 13 is communicated with the waste liquid barrel 4, and the tenth electromagnetic valve is arranged between the upper end liquid outlet of the flow cell 13 and the waste liquid barrel 4.

[0052] Working principle of the present application:

[0053] It should be noted that the opening in the following refers to switching the two-position two-way electromagnetic valve or the two-position three-way electromagnetic valve from the state shown in the figure to another position.

[0054] In work, the plunger pump 7 first sucks a section of air column through the idle sample needle 11, and then sucks a section of sample, and the sample is stored in the pipeline between the upper port of the plunger pump 7 and the eighth electromagnetic valve, then the quantitative pump 5 is opened, the third electromagnetic valve is opened, the sixth electromagnetic valve is opened, the eighth electromagnetic valve is opened, the tenth electromagnetic valve is opened, the plunger pump 7 is pushed, the sheath liquid passes through the first electromagnetic valve, the second electromagnetic valve, the quantitative pump, the third electromagnetic valve, enters the capsule filter 9, and then enters the flow cell through the sixth electromagnetic valve from the side port of the flow cell, and the sample enters the flow cell from the bottom port of the flow cell through the eighth electromagnetic valve and the flow sensor 12.

[0055] The sheath liquid and the sample form a certain flow ratio and enter the flow cell 13 at the same time, and form a stable sheath flow state, and then the sample flow information is collected through forward scattering light, side scattering light and fluorescence signal.

[0056] Generally, after the sample is loaded, the inner and outer walls of the sample needle 11 are usually cleaned with sheath liquid, and when intensive cleaning is required, i.e. during daily shutdown maintenance, the inner and outer walls of the sample needle 11 are cleaned with cleaning liquid first and then sheath liquid.

[0057] Specifically:

[0058] Generally, after the sample is loaded,

[0059] Firstly, clean the outer wall of the sample needle: after the sample is loaded, open the diaphragm pump 14 to form a negative pressure environment, open the quantitative pump 5 to start pumping the sheath liquid in the sheath liquid barrel 1, and then send it into the cleaning swab 10 through the third electromagnetic valve. The sample needle 11 goes down and up in the cleaning swab, and finally returns to the original position. The sheath liquid forms a liquid flow which circulates through the outlet at the upper end of the cleaning swab 10, is discharged through the ninth electromagnetic valve and the diaphragm pump 14, and then enters the waste liquid barrel 4, thereby completing the cleaning of the outer wall of the sample needle 11.

[0060] Then, clean the inner wall of the sample needle: open the diaphragm pump 14 to form a negative pressure environment, open the third electromagnetic valve, open the fourth electromagnetic valve, and open the quantitative pump 5 to start pumping the sheath liquid in the sheath liquid barrel 1. The sheath liquid enters the sample needle 11 through the plunger pump 7. The sample needle 11 stays above the cleaning swab 10. The waste liquid which is flushed in the opposite direction in the sample needle 11 is discharged through the upper end of the cleaning swab 10, and then enters the waste liquid barrel 4 through the diaphragm pump 14, thereby completing the cleaning of the inner wall of the sample needle 11.

[0061] Generally, after the sample is loaded, the flow cell only contains sheath liquid and no sample, so the flow cell can be temporarily cleaned.

[0062] When the cleaning needs to be strengthened, it is divided into two steps: preliminary cleaning with cleaning liquid and re-cleaning with sheath liquid. The preliminary cleaning with cleaning liquid needs to use cleaning liquid to clean the sample needle and the flow cell respectively.

[0063] When the sample needle is cleaned, the outer wall of the sample needle is first cleaned, and then the sample needle is filled. The preliminary cleaning of the outer wall of the sample needle: open the diaphragm pump 14 to form a negative pressure environment, open the quantitative pump 5 and the first electromagnetic valve to start pumping the cleaning liquid in the cleaning liquid barrel 2, and then send it into the cleaning swab 10 through the third electromagnetic valve. The sample needle 11 goes down and up in the cleaning swab, and finally returns to the original position. The cleaning liquid forms a liquid flow which circulates through the outlet at the upper end of the cleaning swab 10, is discharged through the ninth electromagnetic valve and the diaphragm pump 14, and then enters the waste liquid barrel 4, thereby completing the preliminary cleaning of the outer wall of the sample needle 11.

[0064] Fill the sample needle: first take a test tube containing cleaning liquid; open the plunger pump 7 to pump a section of air through the idle sample needle 11, and then place the bottom of the sample needle 11 in the test tube. Open the plunger pump 7 again to pump a small amount of cleaning liquid through the sample needle, so that the cleaning liquid fills the inside of the sample needle 11.

[0065] Preliminary cleaning of flow cell: because the cleaning liquid will have fluorescence signal, after passing through the capsule filter 9, there will be a small amount of residue in it, so that in the subsequent detection with sheath liquid into the flow cell, produce fluorescence interference, affect the final results, so the case also set the seventh solenoid valve; when cleaning the flow cell, bypass the capsule filter 9 and the sixth solenoid valve; the process of preliminary cleaning of flow cell includes: opening the diaphragm pump 14, the first solenoid valve, the quantitative pump 5, the third solenoid valve, the seventh solenoid valve, the ninth solenoid valve, the tenth solenoid valve, so that the quantitative pump 5 from the cleaning liquid barrel 2 into the flow cell 13 after the cleaning liquid is divided into two ways to ensure that the flow cell is full of cleaning liquid;

[0066] Wait for more than 5 minutes for the cleaning liquid to soak in the sample needle and the flow cell, and then flush the sample needle and the flow cell with sheath liquid respectively;

[0067] When cleaning the sample needle with sheath liquid, first clean the outer wall of the sample needle, then clean the inner wall of the sample needle. Clean the outer wall of the sample needle: open the diaphragm pump 14 to form a negative pressure environment, open the quantitative pump 5 to start extracting sheath liquid in the sheath liquid barrel 1, and send it into the cleaning swab 10 through the third solenoid valve. The sample needle 11 goes down and up in the cleaning swab and finally resets. The sheath liquid forms a liquid flow circulating through the outlet at the upper end of the cleaning swab 10, is discharged into the waste liquid barrel 4 through the ninth solenoid valve and the diaphragm pump 14, and the process of strengthening cleaning of the outer wall of the sample needle 11 is completed.

[0068] Clean the inner wall of the sample needle: first, discharge the cleaning liquid originally filled in the sample needle through the plunger pump 7, then open the diaphragm pump 14 to form a negative pressure environment, open the quantitative pump 5, the third solenoid valve and the fourth solenoid valve, so that the sheath liquid in the sheath liquid barrel 1 is extracted by the quantitative pump 5, then sent into the sample needle 11 through the plunger pump 7, and finally discharged. The cleaning liquid or sheath liquid discharged from the sample needle 11 will fall into the cleaning swab 10 and be discharged into the waste liquid barrel 4 through the outlet at the upper end of the cleaning swab 10 through the ninth solenoid valve and the diaphragm pump 14, and the process of strengthening cleaning of the inner wall of the sample needle 11 is completed.

[0069] Final cleaning of flow cell: open the diaphragm pump 14, the quantitative pump 5, the third solenoid valve, the seventh solenoid valve, the ninth solenoid valve and the tenth solenoid valve, so that the sheath liquid extracted from the sheath liquid barrel 1 by the quantitative pump 5 enters the flow cell 13 and is finally discharged, completing the process of strengthening cleaning of the flow cell 13.

[0070] During shutdown or product transportation every day, the sample needle 11 and the flow cell 13 need to be filled with shutdown liquid to prevent crystallization in the quantitative pump 5, the plunger pump 7, the diaphragm pump 14 and the flow cell 13 and the sample needle 11 due to residual sheath liquid after shutdown or product transportation. The shutdown liquid contains a bacteriostatic agent to prevent bacteria from growing in the system.

[0071] As to filling shutdown liquid, it is divided into sample needle filling and flow cell filling. The sample needle is filled: the diaphragm pump 14 is opened to form a negative pressure environment, the second electromagnetic valve, the quantitative pump 5, the third electromagnetic valve and the fourth electromagnetic valve are opened, so that the shutdown liquid in the shutdown liquid barrel 3 is extracted by the quantitative pump 5 and then sent into the sample needle 11 through the plunger pump 7, the leaked shutdown liquid is discharged through the outlet at the upper end of the cleaning swab 10 through the ninth electromagnetic valve and the diaphragm pump 14, so that the sample needle 11 is filled with the shutdown liquid after the quantitative pump 5 and the diaphragm pump 14 stop.

[0072] The flow cell is filled: the diaphragm pump 14, the second electromagnetic valve, the quantitative pump 5, the third electromagnetic valve, the seventh electromagnetic valve, the ninth electromagnetic valve and the tenth electromagnetic valve are opened, so that the shutdown liquid extracted from the shutdown liquid barrel 3 by the quantitative pump 5 is divided into two paths and finally discharged to form a passage, at this time, the quantitative pump 5 and the diaphragm pump 14 are closed, so that the flow cell 13 is filled with the shutdown liquid.

[0073] In addition, the outer wall of the sample needle can also be cleaned by the shutdown liquid: the diaphragm pump 14 is opened to form a negative pressure environment, the quantitative pump 5 and the second electromagnetic valve are opened to extract the shutdown liquid, so that the shutdown liquid enters the quantitative pump 5 through the quantitative pump cleaning channel 5-1 and is sent into the cleaning swab 10 through the third electromagnetic valve, the sample needle 11 is lowered and raised in the cleaning swab and is finally reset; the shutdown liquid forms a liquid flow which is discharged through the outlet at the upper end of the cleaning swab 10 through the ninth electromagnetic valve and the diaphragm pump 14 and enters the waste liquid barrel 4, thereby completing the shutdown liquid cleaning of the outer wall of the sample needle 11.

[0074] In addition:

[0075] The bubble sensor 8 monitors the situation of bubbles in the pipeline, a small amount of bubbles will gather at the upper end of the capsule filter 9, when the bubbles in the pipeline accumulate to a certain amount, the sheath liquid sampling will be affected, at this time, the bubble discharge operation is performed, the third electromagnetic valve is opened, the fifth electromagnetic valve is opened, the quantitative pump 5 is opened to enter the sheath liquid, so that the bubbles in the capsule filter 9 are discharged from the side of the capsule filter 9 through the fifth electromagnetic valve, and the waste liquid enters the waste liquid barrel 4.

[0076] The pressure sensor 6 behind the quantitative pump 5 monitors the situation that the pipeline pressure exceeds the limit, when the pipeline pressure exceeds the limit, the quantitative pump 5 is stopped and the pipeline and the device are checked, the timing, the pipeline and the device maintenance are performed.

[0077] The flow sensor 12 is used to monitor the actual sample flow, compared with the flow provided by the plunger pump 7, the actual plunger pump 7 flow compensation is completed, which can effectively realize the volume method absolute counting.

[0078] The float sensor 1-1 is arranged in the sheath liquid barrel 1, the cleaning liquid barrel 2, the shutdown liquid barrel 3 and the waste liquid barrel 4, which is used to monitor the liquid level in the barrels, so as to add the sheath liquid, the cleaning liquid and the shutdown liquid in time and pour out the waste liquid in time.

[0079] The present application has many specific embodiments and the foregoing merely preferred embodiments and, it will be apparent that those skilled in the art can make modifications of detail, while staying within the scope of the application, which is defined by the appended claims.

Claims

1. A flow cytometer fluidics system based on positive pressure loading, characterized in that, The flow cytometer liquid path system comprises a sheath liquid barrel (1), a cleaning liquid barrel (2), a shutdown liquid barrel (3), a waste liquid barrel (4), a quantitative pump (5), a plunger pump (7), a diaphragm pump (14), and a capsule filter (9), a cleaning swab (10), a sample needle (11), and a flow cell (13); The bottom of the cleaning swab (10) is provided with an inlet, and the top is provided with an outlet, and a cleaning channel for the sample needle (11) to pass through is formed in the cleaning swab (10); one side of the flow cell (13) is provided with a sheath liquid inlet, and the other side is provided with a sheath liquid outlet, and the bottom of the flow cell (13) is provided with a sample inlet, and the top is provided with a sample outlet; the bottom end of the capsule filter (9) is provided with an inlet, the top end is provided with an outlet, and the side part is further provided with a side outlet; The sheath liquid barrel (1), the cleaning liquid barrel (2), and the shutdown liquid barrel (3) are connected to the inlets of the quantitative pump (5), the outlet of the quantitative pump (5) is connected to the inlets of the plunger pump (7), the cleaning swab (10), and the capsule filter (9), the outlet of the capsule filter (9) is connected to the sheath liquid inlet of the flow cell (13), the plunger pump (7) is further connected to the sample needle (11) and the sample inlet of the flow cell (13), the sheath liquid outlet of the flow cell (13) and the outlet of the cleaning swab (10) are both connected to the inlet of the diaphragm pump (14), and the outlet of the diaphragm pump (14), the sample outlet of the flow cell (13), and the side outlet of the capsule filter (9) are all connected to the waste liquid barrel (4); The flow cytometer liquid path system further comprises a first electromagnetic valve (LV01), a second electromagnetic valve (LV02), a third electromagnetic valve (LV03), a fourth electromagnetic valve (LV04), a fifth electromagnetic valve (LV05), a sixth electromagnetic valve (LV06), a seventh electromagnetic valve (LV07), an eighth electromagnetic valve (LV08), a ninth electromagnetic valve (LV09), and a tenth electromagnetic valve (LV10); The first electromagnetic valve (LV01), the second electromagnetic valve (LV02), and the ninth electromagnetic valve (LV09) are two-position three-way electromagnetic valves with two inlets and one outlet, and the third electromagnetic valve (LV03) and the eighth electromagnetic valve (LV08) are two-position three-way electromagnetic valves with one inlet and two outlets; The fourth electromagnetic valve (LV04), the fifth electromagnetic valve (LV05), the sixth electromagnetic valve (LV06), the seventh electromagnetic valve (LV07), and the tenth electromagnetic valve (LV10) are two-position two-way electromagnetic valves with one inlet and one outlet; The sheath liquid barrel (1) and the cleaning liquid barrel (2) are respectively connected to the two inlets of the first electromagnetic valve (LV01), the outlet of the first electromagnetic valve (LV01) and the shutdown liquid barrel (3) are respectively connected to the two inlets of the second electromagnetic valve (LV02), the outlet of the second electromagnetic valve (LV02) is connected to the inlet of the quantitative pump (5), and the outlet of the quantitative pump (5) is connected to the inlet of the third electromagnetic valve (LV03). One of the outlets of the third solenoid valve (LV03) is connected to the inlet of the cleaning swab (10), and the other outlet of the third solenoid valve (LV03) is simultaneously connected to the inlet of the fourth solenoid valve (LV04), the seventh solenoid valve (LV07) and the capsule filter (9), the fourth solenoid valve (LV04) is further connected to the plunger pump (7), the seventh solenoid valve (LV07) is further connected to the sheath liquid inlet of the flow cell (13), the outlet of the capsule filter (9) is connected to the sheath liquid inlet of the flow cell (13) through the sixth solenoid valve (LV06), and the side outlet of the capsule filter (9) is connected to the waste liquid barrel (4) through the fifth solenoid valve (LV05); The plunger pump (7) is connected to the inlets of the fourth solenoid valve (LV04) and the eighth solenoid valve (LV08), one of the outlets of the eighth solenoid valve (LV08) is connected to the sample needle (11), and the other outlet is connected to the sample inlet of the flow cell (13); The sample outlet of the flow cell (13) is connected to the waste liquid barrel (4) through the tenth solenoid valve (LV10), and the sheath liquid outlet of the flow cell (13) and the outlet of the cleaning swab (10) are connected to the two inlets of the ninth solenoid valve (LV09) respectively, and the outlet of the ninth solenoid valve (LV09) is connected to the waste liquid barrel (4) through the diaphragm pump (14); The pressure sensor (6) is further connected between the quantitative pump (5) and the third solenoid valve (LV03); The bubble sensor (8) is further connected between the third solenoid valve (LV03) and the capsule filter (9).

2. A flow cytometer fluidics system based on positive sample loading according to claim 1, wherein, The flow sensor (12) is further connected between the eighth solenoid valve (LV08) and the flow cell (13).

3. A control method for a positive pressure on-deck flow cytometer fluidics system as defined in claim 1, wherein The control method comprises a sample loading method, a general cleaning method and a reinforced cleaning method; The sample loading method is as follows: first, the plunger pump (7) sucks a column of air through the idle sample needle (11), then the plunger pump (7) draws the sample through the sample needle (11) inserted into the sample pool, and then the sample is sent into the flow cell (13); At the same time, the sheath liquid in the sheath liquid barrel (1) is sent into the flow cell (13) through the capsule filter (9) by the quantitative pump (5), and the sheath liquid and the sample enter the flow cell (13) at the same time to form a stable sheath flow state, and then the sample flow information is collected.

4. A control method for a positive pressure sample-based flow cytometer fluidic system as defined in claim 1, characterized by, The control method comprises a general cleaning method, and the general cleaning method comprises a sample needle outer wall general cleaning method and a sample needle inner wall general cleaning method; The sample needle outer wall general cleaning method is as follows: the sheath liquid is sent into the cleaning swab (10) by the quantitative pump (5), and is drawn away by the diaphragm pump (14), the sample needle (11) is inserted into the cleaning swab (10) and then taken out, and the general cleaning of the outer wall of the sample needle (11) is completed; The sample needle inner wall general cleaning method is as follows: the sheath liquid is sent into the inside of the sample needle (11) through the plunger pump (7) by the quantitative pump (5), and the sample needle (11) is kept above the cleaning swab (10), the waste liquid flowing out is drawn away by the diaphragm pump (14), and the general cleaning of the inner wall of the sample needle (11) is completed.

5. A control method for a positive pressure on-deck flow cytometer fluidics system as defined in claim 1, wherein The control method comprises a reinforced cleaning method, which is divided into two steps of preliminary cleaning of the cleaning liquid and re-cleaning of the sheath liquid; In the first step, the sample needle and the flow cell are cleaned respectively by using the cleaning liquid; When cleaning the sample needle, the outer wall of the sample needle is preliminarily cleaned, and the specific method is as follows: the cleaning liquid is sent into the cleaning swab (10) by the quantitative pump (5), and is drawn away by the diaphragm pump (14), in the process, the sample needle (11) is inserted into the cleaning swab (10) and taken out, thereby completing the preliminary cleaning of the outer wall of the sample needle (11); Then, the sample needle is refilled, and the specific method is as follows: a test tube containing the cleaning liquid is taken, and is ready for use; the plunger pump (7) is opened to draw a section of air through the idle sample needle (11), then the bottom of the sample needle (11) is placed in the test tube, and the plunger pump (7) is opened again to draw a small amount of cleaning liquid through the sample needle, so that the cleaning liquid fills the inside of the sample needle (11); The specific method for cleaning the flow cell by using the cleaning liquid is as follows: the cleaning liquid is drawn from the cleaning liquid barrel (2) by the quantitative pump (5), and is sent into the flow cell (13) after bypassing the capsule filter, then is divided into two paths and enters the waste liquid barrel (4) from the sheath liquid outlet and the sample outlet of the flow cell (13) respectively, the quantitative pump (5) and the diaphragm pump (14) are closed to make the flow cell (13) full of the cleaning liquid; In the second step, the cleaning liquid is soaked in the sample needle and the flow cell for more than 5 minutes, and then the sample needle and the flow cell are washed by the sheath liquid respectively; When the sample needle is cleaned by the sheath liquid, the outer wall of the sample needle is preliminarily cleaned, and the specific method is as follows: the sheath liquid is sent into the cleaning swab (10) by the quantitative pump (5), and is drawn away by the diaphragm pump (14), in the process, the sample needle (11) is inserted into the cleaning swab (10) and taken out, thereby completing the reinforced cleaning process of the outer wall of the sample needle (11); Then, the inner wall of the sample needle is cleaned, and the specific method is as follows: the cleaning liquid originally filled in the sample needle (11) is discharged by the plunger pump (7), then the sheath liquid is sent into the inside of the sample needle (11) by the quantitative pump (5) through the plunger pump (7), and the sample needle (11) is kept above the cleaning swab (10), the waste liquid is drawn away by the diaphragm pump (14), thereby completing the reinforced cleaning process of the inner wall of the sample needle (11); The specific method for cleaning the flow cell by using the sheath liquid is as follows: the cleaning liquid is drawn from the sheath liquid barrel (1) by the quantitative pump (5), and is sent into the flow cell (13) after bypassing the capsule filter, then is divided into two paths and enters the waste liquid barrel (4) from the sheath liquid outlet and the sample outlet of the flow cell (13) respectively, thereby completing the reinforced cleaning process of the flow cell (13).

6. A control method for a positive pressure sample-based flow cytometer fluidic system as defined in claim 1, wherein The control method comprises a shutdown liquid filling method, and the shutdown liquid filling method comprises a sample needle filling method and a flow cell filling method; The sample needle filling method is as follows: the shutdown liquid in the shutdown liquid barrel (3) is sent into the inside of the sample needle (11) by the quantitative pump (5) through the plunger pump (7), and the sample needle (11) is kept above the cleaning swab (10), the waste liquid is drawn away by the diaphragm pump (14), after the quantitative pump (5) and the diaphragm pump (14) are stopped, the inside of the sample needle (11) is full of the shutdown liquid; The flow cell filling method is: the shutdown liquid is extracted from the shutdown liquid barrel (3) by the quantitative pump (5), and is sent into the flow cell (13) after bypassing the capsule filter, and then is divided into two ways, respectively entering the waste liquid barrel (4) from the sheath liquid outlet and the sample outlet of the flow cell (13), and the quantitative pump (5) and the diaphragm pump (14) are closed to fill the flow cell (13) with the shutdown liquid.

Images