CN103837462B - A kind of small-sized flow cytometer liquid-way system - Google Patents

Abstract

The invention discloses a liquid circuit system of a small flow cytometer, which comprises a flow chamber, a sample pipeline, a sheath fluid pipeline, a waste liquid pipeline and a gas pipeline. The sample to be tested is transported by a small-flow peristaltic pump and enters the flow chamber after passing through the pulsation buffer; the sheath liquid is transported by a large-flow peristaltic pump or diaphragm pump, and enters the flow chamber unilaterally or symmetrically after passing through the capsule filter element and the pulsation buffer; the micro air pump It is connected to the flow chamber via a one-way valve to realize the function of eliminating air bubbles in the pipeline. A pressure sensor is installed at the outlet of the flow chamber to monitor pressure changes and abnormalities. The liquid level sensor is placed in the sheath liquid pool and the waste liquid pool to set the liquid level warning. The filter screen is installed at the connection between the sheath liquid pool and the micro air pump to prevent impurities from entering. The invention has a simple structure, abandons the air compression pump used in the traditional flow cytometer, is easy to clean, and has the function of removing air bubbles, and can meet the application requirements of the miniaturization of the flow cytometer.

Description

A small flow cytometer liquid circuit system

technical field

The invention relates to the technical field of cell detection and analysis, in particular to a small flow cytometer liquid path system.

Background technique

Flow cytometer is an instrument capable of multi-parameter, quantitative analysis or sorting of cells or biological particles in a fast-flowing state, and has been widely used in clinical medicine, cytology, biology, microbiology And other fields, known as the CT of the cell biology laboratory. As one of the important components of the flow cytometer, the fluid circuit system is used to focus on the position of the cells or particles to be detected by using the principle of fluid dynamics, so that they pass through the laser detection area one by one. The cells to be tested are prepared into a single cell suspension, labeled with a fluorescent dye, placed in a sample tube, and enter the flow chamber under pumping pressure to form a sample flow; the sheath fluid is the matrix fluid that assists the sample flow to be normally detected, and is wrapped in Around the sample flow, keep it in the center of the flow chamber to ensure detection accuracy, and at the same time prevent the cells in the sample flow from getting close to the nozzle wall and blocking the nozzle hole. The sample and sheath fluid are combined in the flow cell and exit the flow cell to waste.

In the previous liquid circuit system of flow cytometer, the vacuum pump was often used as the power source of the entire liquid circuit system, supplemented by solenoid valves, check valves and pressure balance devices, to complete the delivery of samples, sheath fluid and waste disposal. Liquid discharge (US5395588, US4844610, EP0478392). Due to the large damping of the liquid circuit system, certain requirements are placed on the output power of the vacuum pump, resulting in a large and heavy vacuum pump, which increases the volume and weight of the entire instrument. And some flow cytometer patents propose to use a syringe pump supplemented by a multi-channel solenoid valve for sample delivery (US8394326, CN101451989). The flow rate of the sample driven by this method is stable and reduces the burden on the vacuum pump. However, the sample to be tested will enter the pump chamber of the syringe pump and the gap of the solenoid valve, which is not easy to clean, resulting in serious cross-contamination between different samples and affecting the measurement accuracy. Recently, a patent on flow cytometry proposes to use two peristaltic pumps to transport the sheath fluid and waste fluid, and use the pressure difference generated by the two to drive the sample flow (US7981661, US8187888). This method has a good effect in reducing the volume and weight of the instrument, but on the one hand, the sample pipeline is not easy to clean; Existence is often an important factor affecting the detection accuracy, and it is also very likely to directly lead to wrong results.

Contents of the invention

The purpose of the present invention is to overcome the above problems existing in the prior art, and to provide a liquid circuit system of a small flow cytometer, which can reduce the volume and weight of the existing flow cytometer, and is especially suitable for small flow cytometers. The sample is wrapped by the sheath fluid and passes through the laser detection area at a certain speed and distribution width. At the same time, it can realize the function of degassing and the cleaning function between different samples, so as to improve the detection accuracy.

In order to achieve the above-mentioned technical purpose and achieve the above-mentioned technical effect, the present invention is realized through the following technical solutions:

A liquid circuit system of a small flow cytometer, including a sample pipeline, a sheath fluid pipeline, a waste fluid pipeline and a gas pipeline;

A sampling needle, a sample pump and a sampling needle are sequentially installed on the sample pipeline;

One end of the sheath fluid pipeline is connected to the flow chamber and passes through the flow chamber, the other end of the sheath fluid pipeline is connected to the sheath fluid pool, and a capsule filter element is installed on the sheath fluid pipeline, and the sheath fluid pool A first liquid level sensor is installed at the bottom of the sheath liquid pool, and a first filter screen is installed at the outlet of the sheath liquid pool communicated with the atmosphere.

The waste liquid pipeline is connected to the outlet of the flow chamber and a waste liquid pool, and a second liquid level sensor is installed on the top of the waste liquid pool.

Two ends of the gas pipeline are connected with a one-way valve and a micro air pump, and the gas pipeline is connected with the sheath fluid pipeline through the one-way valve.

The sample pipeline is connected in series with the sample pump and the sample pulsation buffer, the sheath fluid pipeline is connected in series with the capsule filter element and the sheath fluid pulsation buffer, a pressure sensor is installed at the outlet of the flow chamber, and the A second filter screen is installed at the outlet where the micro air pump communicates with the atmosphere.

The sample pump is a small-flow peristaltic pump with a flow range of 1-200 μL/min; the sheath fluid pump is a large-flow peristaltic pump or a diaphragm pump with a flow range of 1-50 mL/min; the sheath fluid pulsation buffer adopts One of the air type, air bag type, and diaphragm type pulse dampers. The sample pulsation buffer adopts a straight-through pipeline with a thinner inner diameter, and the inner diameter is less than 250 μm, or several elastic pipeline holders clamp the sample pipeline. To achieve, the elastic pipe clamp includes a clamping seat and an elastic arm.

A sheath fluid pump is installed in the waste fluid pipeline or in the sheath fluid pipeline.

The gas pipeline is perpendicular to the directly opposite side of the flow chamber or perpendicular to the right side of the flow chamber.

When the gas pipeline is perpendicular to the opposite side of the flow chamber, the sheath fluid pipeline is divided into two branches connected to the flow chamber, which are at 90 degrees to the gas pipeline.

The beneficial effects of the present invention are:

1. Since the peristaltic pump is used as the sample pump and the diaphragm pump is used as the sheath liquid pump, the volume and weight of the flow cytometer are greatly reduced;

2. The micro air pump is used to realize the function of removing air bubbles, which solves the problem of detection errors or errors that may be caused by air bubbles;

3. The sample pump is reversed to realize the cleaning function of the sample pipeline, which makes the cleaning of the tube wall of the sample flow path more thorough and effective, reduces the possibility of the sample blocking the pipeline, and improves the detection accuracy at the same time;

4. The proposed (micro)flow pulsation damper, ie the elastic pipe holder, can also be used in other fluid systems.

Description of drawings

Fig. 1 is the overall structural diagram of embodiment 1;

Fig. 2 is the overall structural diagram of embodiment 2;

Fig. 3 is the overall structural diagram of embodiment 3;

Fig. 4 is the overall structural diagram of embodiment 4;

Fig. 5 is a control diagram of the present invention;

Fig. 6 is a structural diagram of the elastic pipe holder.

Explanation of symbols in the figure: 1. Flow chamber, 2. Injection needle, 3. Laser detection area, 4. Sample pipeline, 5. Sample pulsation buffer, 6. Sample pump, 7. Sample suction needle, 8. Sample loading Test tube, 9, sheath liquid pool, 1001, first liquid level sensor, 1002, second liquid level sensor, 1101, first filter screen, 1102, second filter screen, 12, sheath liquid pump, 13, capsule filter element, 14. Sheath fluid pulsation buffer, 15. Sheath fluid pipeline, 16. Check valve, 17. Gas pipeline, 18. Micro air pump, 19. Pressure sensor, 20. Waste fluid pipeline, 21. Waste fluid pool, 30, deck, 31, elastic arm.

detailed description

The present invention will be described in detail below with reference to the accompanying drawings and in combination with embodiments.

Example 1

Referring to Fig. 1, a small flow cytometer liquid circuit system enables the sample to pass through the laser detection area 3 at a certain speed and distribution width under the sheath fluid package, including the sample pipeline 4, the sheath fluid pipeline 15, Waste liquid pipeline 20 and gas pipeline 17;

A sample injection needle 2, a sample pump 6, and a sample suction needle 7 are sequentially installed on the sample pipeline 4;

One end of the sheath fluid pipeline 15 is connected to the flow chamber 1 and passes through the flow chamber 1, the other end of the sheath fluid pipeline 15 is connected to the sheath fluid pool 9, and a bladder type A filter element 13 and a sheath liquid pump 12, a first liquid level sensor 1001 is installed at the bottom of the sheath liquid pool 9, and a first filter screen 1101 is installed at the outlet of the sheath liquid pool 9 communicating with the atmosphere;

The waste liquid pipeline 20 is connected to the outlet of the flow chamber 1 and a waste liquid pool 21, and a second liquid level sensor 1002 is installed on the top of the waste liquid pool 21;

Both ends of the gas pipeline 17 are connected to a one-way valve 16 and a micro air pump 18 , and the gas pipeline 17 is connected to the sheath fluid pipeline 15 through the one-way valve 16 .

The sample pump 6 is a small-flow peristaltic pump with a flow range of 1-200 μL/min; the sheath fluid pump 12 is a large-flow peristaltic pump or a diaphragm pump with a flow range of 1-50 mL/min;

The sheath fluid is driven by the sheath fluid pump 12 and enters the flow chamber through the sheath fluid pipeline 15 to form a sheath fluid flow. The capsule filter element 13 is used to filter impurities in the sheath fluid, and at the same time has a certain buffering effect on the pulsation of the sheath fluid.

The effect of one-way valve 16 is to stop liquid from entering in the miniature air pump 18. Due to improper operation or other accidental factors, small air bubbles remain in the flow chamber 1, which will greatly affect the detection accuracy. Therefore, when an abnormality is found, it is necessary to implement the function of removing air bubbles. At this time, the micro air pump 18 inflates the sheath liquid pipeline 15 and the flow chamber 1 through the gas pipeline 17, and after the liquid in the flow chamber is completely emptied, the sheath liquid of the sample is loaded, so that the remaining fluid in the flow chamber Small air bubbles are expelled.

The liquid level sensors 1001 and 1002 monitor the liquid level, and generate signals when the sheath liquid is about to run out and the waste liquid is about to be full, and the main controller realizes an alarm, and shuts down all pumps at the same time, reminding the user to add sheath liquid or remove waste liquid. The filter screen 1101 is used to isolate external impurities to prevent them from falling into the sheath liquid pool.

As shown in Fig. 5, the control method is as follows: when the operation command is issued, the main controller turns on the sample pump 6 and the sheath liquid pump 12; when the stop command is issued, the main controller turns off the sample pump 6 and the sheath liquid pump 12. When the flow rate regulation command is issued, the main controller changes the speed regulation control voltage of the sample pump 6 .

Between loading different samples, the cleaning function needs to be performed. First, the previous sample tube to be tested needs to be removed, an empty waste liquid sample tube is placed, and the cleaning command is executed. The main controller turns on the reverse mode of the sample pump 6 , turn on the sheath liquid pump 12 to clean the entire fluid pipeline, and the waste liquid in the sample pipeline flows into the waste liquid sample tube. Execution stops after a few minutes. Load the sample with clean water, execute the run, stop after a while, and then load the next sample to be tested.

When executing the command to remove air bubbles, use clean water to load the sample, and the main controller sends a command to turn on the micro air pump 18, and then turn off the air pump 18 after a few seconds, then turn on the sample pump 6 and the sheath liquid pump 12, and turn off the sample pump 6 and the sheath pump after a few seconds Liquid pump 12.

When the sheath liquid level sensor or the waste liquid level sensor sends out a signal, the main controller controls the buzzer to generate a liquid level alarm, and shuts down all pumps at the same time.

Example 2

As shown in Figure 2, the sample pipeline 4 is connected in series with the sample pump 6 and the sample pulsation buffer 5, and the sheath fluid pipeline 15 is connected in series with the capsule filter element 13, the sheath fluid pump 12 and the sheath fluid pulsation buffer. Buffer 14, a pressure sensor 19 is installed at the outlet of the flow chamber 1, and a second filter screen 1102 is installed at the outlet of the micro air pump 18 communicated with the atmosphere.

The sheath fluid pulsation buffer 14 adopts one of air type, air bag type, and diaphragm type pulsation damper, and the sample pulsation buffer 5 adopts a straight-through pipeline with a smaller inner diameter, the inner diameter is less than 250 μm, or several elastic pipelines The sample pipeline is clamped by a holder, and the elastic pipeline holder includes a clamp 30 and an elastic arm 31 , as shown in FIG. 6 .

Since the sample flow rate is 10 μL/min-60 μL/min, the existing pulsation buffer has no effect on such a small flow rate, and the liquid volume of the sample itself is small, so too much sample cannot be retained in the buffer. Elastic tubing holders smooth out pulsation and reduce tubing volume to avoid sample waste. Its essence is to flatten part of the sample pipeline 4, as shown in Fig. 5, to make its inner diameter smaller, increase flow resistance, and reduce fluid pulsation. The inner diameter of the existing flow cytometer sample pipeline is 180 μm-250 μm, and it is difficult to manufacture fluid pipelines with smaller inner diameters. The use of pipeline holders can effectively reduce costs. The sheath fluid pulsation buffer 14 is used to smooth the flow pulsation when the sheath fluid is transported.

The pressure sensor 19 is mainly used to monitor pipeline blockage or abnormal pressure caused by large air bubbles. When the pressure sensor 19 outputs an abnormal signal, the main controller generates a pressure alarm, shuts down all the pumps simultaneously, and then executes the air bubble command. If the pressure is still abnormal after performing several times of degassing, the pipe should be blocked and the cleaning function can be performed.

The filter screen 1102 is used to isolate impurities in the atmosphere, preventing them from entering the flow chamber 1 and disturbing the flow path.

Example 3

As shown in FIG. 3 , the sheath liquid pump 12 is placed in the waste liquid pipeline 20 and acts as a waste liquid pump to drive the waste liquid mixed with the sample and the sheath liquid into the waste liquid pool. The flow of the sheath fluid is driven by the pressure difference between the sheath fluid pump 12 and the sample pump 6 .

Example 4

As shown in Figure 4, three inlets are opened on the flow chamber 1, the positions are 90° to each other, and the two inlets are connected together and connected to the sheath fluid pipeline 15, and the remaining outlets are connected through the sheath fluid pipeline 15 One-way valve 16, the one-way valve 16 is connected to the micro air pump 18 through the gas pipeline. The symmetrical injection method of the sheath fluid can reduce the impact on the injection needle 2 caused by the single side injection of the sheath fluid, and reduce the measurement error caused by the displacement of the sample outlet of the injection needle.

Claims (5)

1. a small-sized flow cytometer liquid-way system, it is characterised in that: include sample inlet (4), sheath liquid pipeline (15), waste liquid Pipeline (20) and gas piping (17)；

It is sequentially installed with sample introduction needle (2), sample pump (6) and suction needle (7) in described sample inlet (4);

One end of described sheath liquid pipeline (15) connects flow chamber (1) and passes through described flow chamber (1), described sheath liquid pipeline (15) Other end connecting utricle liquid pool (9), described sheath liquid pipeline (15) is provided with bellows filter core (13), the bottom of described sheath liquid pool (9) Being provided with the first liquid level sensor (1001), described sheath liquid pool (9) is provided with the first filter screen with the exit of atmosphere (1101);

Described waste-solution line (20) connects outlet and the waste liquid pool (21) of described flow chamber (1), the top of described waste liquid pool (21) Second liquid level sensor (1002) is installed;

The two ends of described gas piping (17) connect check valve (16) and micro air pump (18), and described gas piping (17) passes through institute State check valve (16) and connect described sheath liquid pipeline (15)；

Described sample pump (6) is low discharge peristaltic pump, and range of flow is at 1-200 μ L/min；Sheath liquid pump (12) is that big flow is wriggled Pump or membrane pump, range of flow is at 1-50mL/min；Sheath fluid pulsation damper (14) uses air type, gasbag-type, diaphragm type arteries and veins Rushing the one of damper, sample pulsation damper (5) uses the thinner straight-through pipeline of internal diameter, and internal diameter is less than 250 μm, or some The mode of individual elastic pipeline gripper sample inlet realizes, described elastic pipeline clamper includes deck (30) and elasticity Arm (31).

2. small-sized flow cytometer liquid-way system according to claim 1, it is characterised in that: described sample inlet (4) is gone here and there Connecing described sample pump (6) and sample pulsation damper (5), described sheath liquid pipeline (15) concatenates described bellows filter core (13) and sheath fluid Pulsation damper (14), pressure sensor (19), described micro air pump (18) and air are installed in the exit of described flow chamber (1) The exit of connection is provided with the second filter screen (1102).

3. small-sized flow cytometer liquid-way system according to claim 1, it is characterised in that: in described waste-solution line (20) Or described sheath liquid pipeline (15) is provided with sheath liquid pump (12).

4. small-sized flow cytometer liquid-way system according to claim 1, it is characterised in that: described gas piping (17) hangs down Directly in right opposite or the right flank being perpendicular to described flow chamber (1) of described flow chamber (1).

5. small-sized flow cytometer liquid-way system according to claim 4, it is characterised in that: described gas piping (17) hangs down Straight when the right opposite of described flow chamber (1), sheath liquid pipeline (15) is divided into Liang Ge branch to be connected to flow chamber (1), and with described Gas piping (17) becomes 90 degree.