CN118549303B - 3D prints biological particle deformation detection device based on flow impedance technique - Google Patents

Abstract

A 3D printed biological particle deformation detection device based on flow impedance technology includes two parts: a flow channel layer and an electrode layer. The flow channel layer is provided with a focusing flow channel, two sheath flow channels, an extrusion flow channel and a collision flow channel; the electrode layer is provided with a shape detection electrode, an extrusion detection electrode and a collision detection electrode. The shape detection electrode is located at the bottom of the focusing flow channel, the extrusion detection electrode is located at the bottom of the extrusion flow channel, and the collision detection electrode is located at the bottom of the collision flow channel. The focusing flow channel focuses the biological particles at a specific position to increase the accuracy of morphological detection; the extrusion flow channel causes the biological particles to be squeezed and deformed by the fluids on both sides, and the extrusion detection electrode obtains the impedance signal when the biological particles are squeezed; the collision flow channel causes the biological particles to directly collide with the collision flow channel and deform, and the collision detection electrode records the deformation and the recovery state after deformation. Moreover, the device is designed based on flow impedance technology, has a high throughput advantage compared to image detection, and is suitable for biochemical fields such as single cell research.

Description

A 3D printed biological particle deformation detection device based on flow impedance technology

Technical Field

The present invention relates to the field of power system automation, and in particular to a 3D printed biological particle deformation detection device based on flow impedance technology.

Background Art

The measurement of deformation characteristics of biological particles such as cells is of great significance in the fields of life sciences, clinical medicine, chemistry and chemical engineering. At present, the mainstream method for measuring the deformation of biological particles still mainly adopts the method of image detection, that is, a high-speed camera is used to continuously shoot the deformed biological particles, and the deformation characteristics of the biological particles are obtained by image analysis. However, the detection flux of such methods is extremely low, usually one per few seconds, which is difficult to apply to the high-throughput detection of large background cells. In addition, such methods often rely on expensive high-speed cameras and require professional personnel to operate, and their detection costs are often high. In recent years, with the advancement of microfluidics technology, flow impedance detection technology has made great progress in the detection of biological particles. Taking cells as an example, flow impedance detection technology realizes non-label detection of cells by analyzing the disturbance of the electric field when the cells pass through the electric field area. For example, this method can realize the type identification of white blood cells and tumor cells. However, existing flow impedance detection devices are usually difficult to achieve high-throughput detection of biological particle deformation. In view of the advantages of high throughput and non-labeling in the detection of biological particles, it is of great significance to develop a device based on flow impedance cytometry that can detect the deformation characteristics of biological particles. It has great application value in the fields of disease diagnosis and single cell physical properties research.

Summary of the invention

In order to solve the above technical problems, the present invention proposes a 3D printing biological particle deformation detection device based on flow impedance technology, which is mainly designed based on flow impedance detection technology and inertial microfluidics technology, and its production method involves 3D printing technology of heterogeneous materials.

To achieve the above object, the technical solution adopted by the present invention is:

A 3D printed biological particle deformation detection device based on flow impedance technology, composed of a flow channel layer and an electrode layer, the flow channel layer and the electrode layer are sealed up and down; divided into 4 detection areas and respectively used to measure the shape of biological particles, the detection area used to measure the deformation of biological particles caused by fluid extrusion, the detection area used to measure the deformation of biological particles caused by collision with the flow channel wall, and the detection area used to measure the deformation of biological particles caused by collision with the flow channel wall. The detection area used to measure the shape of biological particles has one detection area, which is detection area one, the detection area used to measure the deformation of biological particles caused by fluid extrusion has one detection area, the detection area used to measure the deformation of biological particles caused by fluid extrusion has two detection areas, which are symmetrically arranged, namely detection area three and detection area five, and the detection area used to measure the deformation of biological particles caused by collision with the flow channel wall has two symmetrically arranged, namely detection area four and detection area six;

The detection area one is formed by the focusing flow channel and the shape detection electrode one, the shape detection electrode two, and the shape detection electrode three; the detection area two is formed by the extrusion flow channel and the extrusion detection electrode one, the extrusion detection electrode two, and the extrusion detection electrode three; the detection area three is formed by the collision flow channel two and the collision detection electrode five and the collision detection electrode six; the detection area four is formed by the collision flow channel two and the collision detection electrode seven and the collision detection electrode eight; the detection area five is formed by the collision flow channel one and the collision detection electrode three and the collision detection electrode four; the detection area six is formed by the collision flow channel two and the collision detection electrode one and the collision detection electrode two.

Preferably, the flow channel layer is provided with a sample inlet, sheath liquid inlet one and sheath liquid inlet two, sample outlet one and sample outlet two, as well as electrode hole one, electrode hole two, electrode hole three, electrode hole four, electrode hole five, electrode hole six, electrode hole seven, electrode hole eight and electrode hole nine, and the bottom surface of the flow channel layer is provided with a focusing flow channel, sheath liquid flow channel one and sheath liquid flow channel two, an extrusion flow channel, and a collision flow channel one and a collision flow channel two, and the sheath liquid flow channel one and the sheath liquid flow channel two are located on both sides of the extrusion flow channel; the collision flow channel one and the collision flow channel two are directly connected and vertically connected to the extrusion flow channel, and when biological particles enter the collision flow channel one or the collision flow channel two, they collide with the flow channel wall to produce deformation, and the biological particles flow out through the collision flow channel one or the collision flow channel two.

Preferably, the electrode layer is provided with shape detection electrode 1, shape detection electrode 2, shape detection electrode 3, extrusion detection electrode 1, extrusion detection electrode 2, extrusion detection electrode 3, and collision detection electrode 1, collision detection electrode 2, collision detection electrode 3, collision detection electrode 4, collision detection electrode 5, collision detection electrode 6, collision detection electrode 7, and collision detection electrode 8, wherein shape detection electrode 1, shape detection electrode 3, extrusion detection electrode 1, extrusion detection electrode 3, collision detection electrode 1, collision detection electrode 4, collision detection electrode 5, and collision detection electrode 8 are sensing electrodes, and shape detection electrode 2, extrusion detection electrode 2, collision detection electrode 2, collision detection electrode 3, collision detection electrode 6, and collision detection electrode 7 are excitation electrodes; the sizes of all electrodes match the sizes of biological particles.

Preferably, the focusing channel, the extrusion channel, the collision channel 1 and the collision channel 2 are straight flow channels with rectangular cross-sections.

Preferably, the electrode layer is made of heterogeneous materials by three-dimensional printing, the electrode part is a conductive material, and the non-electrode part is an insulating material, and the flow channel layer is made of non-conductive materials by three-dimensional printing.

Preferably, when the flow channel layer and the electrode layer are assembled from top to bottom, they are sealed by a hot pressing process or by a sealant.

Principle description:

In this device, the sample liquid containing biological particles is injected into the flow channel of the device through the device inlet. The front end of the flow channel is a straight flow channel with a high aspect ratio. Under the inertial flow of a limited flow rate, the biological particles can be focused at a specific position of the flow channel, so that they pass through the shape detection area (detection area one) at a specific position; the principle is that the biological particles in the straight flow channel with a high aspect ratio are focused at a specific position of the flow channel due to the inertial lift force. Subsequently, the fluid in the sheath liquid flow channel flows into the extrusion flow channel, and the biological particles are thus squeezed and deformed by the fluid (detection area two). The biological particles then flow through the extrusion flow channel to the collision flow channel, where they are deformed after colliding with the wall (detection area three), and restore their original shape at the end of the collision flow channel (detection area four).

In the detection area of the present invention, the basic principle is that when biological particles pass through the electric field area, the electric field disturbance will be generated. By analyzing the disturbance effect of biological particles on the electric field, the morphology and deformation characteristics of biological particles can be measured. The present invention has four detection areas in total. Detection area one is used to measure the morphological information of biological particles, detection area two is used to measure the characteristics of biological particles being deformed by fluid extrusion, detection area three is used to measure the characteristics of biological particles colliding with the wall of the flow channel to generate deformation, and detection area four is used to measure the characteristics of the recovery state of biological particles after deformation. During the detection process, the signal generator of the phase-locked amplifier emits an excitation electric field, which is a multi-frequency AC electric field, which is respectively connected to the excitation electrodes of the detection areas one to four, and the sensing electrodes of the detection areas one to four are short-circuited as shown in the figure below and then connected to the transimpedance amplifier. The two currents in the transimpedance amplifier are again connected to the phase-locked amplifier to demodulate the weak signal, and finally the morphology and deformation signals of the biological particles are obtained.

Since the excitation electric field is a multi-frequency AC electric field, the impedance sensing signal includes low-frequency and high-frequency parts. When the biological particles are in the detection area 1, the shape and internal characteristics of the biological particles can be obtained by analyzing the amplitude phase of the impedance signal in this area. When the biological particles are in the detection area 2, the biological particles are squeezed by the fluid and are elongated in the X-axis direction. At this time, the disturbance caused by the biological particles in the electric field is greater, so the impedance signal is shown as having a larger amplitude and peak width. By analyzing the amplitude and peak width between the detection area 1 and the detection area 2, the deformation of the biological particles caused by the fluid can be obtained. Similarly, the biological particles are deformed due to collision in the detection area 3. By analyzing the changes in the impedance signal of the biological particles at various frequencies, the collision deformation information of the biological particles can be obtained. It should be noted that the previous experimental studies have shown that there is a certain difference between the extrusion deformation and the collision deformation of the biological particles. Since the distance between the detection area 4 and the detection area 3 is specially designed, the biological particles are recovering to their original shape when they are in the detection area 4'. By analyzing the impedance signal, information on the deformation and recovery speed of the biological particles can also be obtained. In addition, since the detection areas five and six are symmetrically distributed with the detection areas three and four, the effects when the biological particles flow through these areas are the same, so they will not be described in detail.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG1 is an exploded view of the assembly of a biological particle deformation detection device manufactured by 3D printing according to the present invention;

FIG2 is a schematic diagram of the structure of the flow channel layer;

FIG3 is a schematic diagram of the detailed structure of the front side of the flow channel layer;

FIG4 is a schematic diagram of the detailed structure of the back side of the flow channel layer;

FIG5 is a partial schematic diagram of the back side of the flow channel layer;

FIG6 is a schematic diagram of the structure of an electrode layer;

FIG7 is a schematic diagram of the detailed structure of the front side of the electrode layer;

FIG8 is a schematic diagram of the detailed structure of the front side of the electrode layer;

FIG9 is a second schematic diagram of the detailed structure of the front side of the electrode layer;

FIG10 is a third schematic diagram of the detailed structure of the front side of the electrode layer;

FIG11 is a schematic diagram of the detection area after the flow channel layer and the electrode layer are assembled;

FIG12 is a local schematic diagram of the detection area after the flow channel layer and the electrode layer are assembled;

FIG13 is a second schematic diagram of the local detection area after the flow channel layer and the electrode layer are assembled;

FIG14 is a third local schematic diagram of the detection area after the flow channel layer and the electrode layer are assembled;

FIG15 is a schematic diagram of the detection principle;

FIG16 is an original signal of biological particle deformation measurement by the device of the present invention;

The component names are as follows:

1. Flow channel layer; 2. Electrode layer; 11. Sample inlet; 12. Sheath liquid inlet 1; 13. Sample outlet 1; 14. Electrode hole 1; 15. Electrode hole 2; 16. Electrode hole 3; 17. Sample outlet 2; 18. Electrode hole 4; 19. Electrode hole 5; 110. Electrode hole 6; 111. Sheath liquid inlet 2; 112. Electrode hole 7; 113. Electrode hole 8; 114. Electrode hole 9; 115. Focusing channel; 116. Sheath liquid channel 1; 117. Extrusion channel; 118. Collision channel 1; 119. Collision channel two; 120, sheath liquid channel two; 21, shape detection electrode one; 22, shape detection electrode two; 23, shape detection electrode three; 24, extrusion detection electrode one; 25, extrusion detection electrode two; 26, extrusion detection electrode three; 27, collision detection electrode one; 28, collision detection electrode two; 29, collision detection electrode three; 210, collision detection electrode four; 211, collision detection electrode five; 212, collision detection electrode six; 213, collision detection electrode seven; 214, collision detection electrode eight.

DETAILED DESCRIPTION

The present invention is further described in detail below in conjunction with the accompanying drawings and specific embodiments:

As shown in FIG1 , the 3D printed biological particle deformation detection device based on flow impedance technology of the present invention is composed of a flow channel layer 1 and an electrode layer 2, and the flow channel layer 1 and the electrode layer 2 are sealed up and down to prevent leakage;

As shown in Fig. 2-5, the flow channel layer 1 is provided with a sample inlet 11, a sheath liquid inlet 1 12 and a sheath liquid inlet 2 111, a sample outlet 1 13 and a sample outlet 2 17, as well as an electrode hole 14, an electrode hole 2 15, an electrode hole 3 16, an electrode hole 4 18, an electrode hole 5 19, an electrode hole 6 110, an electrode hole 7 112, an electrode hole 8 113, and an electrode hole 9 114. The bottom surface of the flow channel layer 1 is provided with a focusing flow channel 115, a sheath liquid flow channel 1 116 and a sheath liquid flow channel 2 120, an extrusion flow channel 117, and a collision flow channel 1 118 and a collision flow channel 2 119; the focusing flow channel 115 is a straight flow channel with a rectangular cross section. The sheath liquid flow channel 1 116 and the sheath liquid flow channel 2 120 are located on both sides of the extrusion flow channel 117. The biological particles are focused at a specific equilibrium position through the focusing flow channel 115 and then enter the extrusion flow channel 117. Under the action of the fluid in the sheath liquid flow channel 1 116 and the sheath liquid flow channel 2 120, the biological particles are squeezed and deformed; the collision flow channel 1 118 and the collision flow channel 2 119 are directly connected and vertically connected to the extrusion flow channel 117. When the biological particles enter the collision flow channel 1 118 or the collision flow channel 2 119, they collide with the flow channel wall and deform. It should be noted that the biological particles flow out through the collision flow channel 1 118 or the collision flow channel 2 119;

As shown in Fig. 6-10, the electrode layer is provided with shape detection electrode 1 21, shape detection electrode 2 22, shape detection electrode 3 23, extrusion detection electrode 1 24, extrusion detection electrode 2 25, extrusion detection electrode 3 26, collision detection electrode 1 27, collision detection electrode 2 28, collision detection electrode 3 29, collision detection electrode 4 210, collision detection electrode 5 211, collision detection electrode 6 212, collision detection electrode 7 213, and collision detection electrode 8 214, wherein shape detection electrode 1 21, shape detection electrode 3 23, extrusion detection electrode 1 24, extrusion detection electrode 3 26, collision detection electrode 1 27, collision detection electrode 4 210, collision detection electrode 5 211, and collision detection electrode 8 214 are sensing electrodes, and shape detection electrode 2 22, extrusion detection electrode 2 25, collision detection electrode 2 28, collision detection electrode 3 29, collision detection electrode 6 212, and collision detection electrode 7 213 are excitation electrodes; the sizes of all electrodes match the sizes of biological particles (such as cells).

As shown in Figures 11-14, the focusing channel 115 and the shape detection electrode 1 21, the shape detection electrode 2 22, and the shape detection electrode 3 23 together form a detection area 1, the extrusion channel 117 and the extrusion detection electrode 1 24, the extrusion detection electrode 2 25, and the extrusion detection electrode 3 26 together form a detection area 2, the collision channel 2 119 and the collision detection electrode 5 211 and the collision detection electrode 6 212 together form a detection area 3, and the collision channel 2 119 and the collision detection electrode 7 213 and the collision detection electrode 8 214 together form a detection area 4; in addition, since the collision channel 1 118 and the collision channel 2 119 are symmetrical, biological particles can only be discharged from one of the channels, so the area formed by the collision channel 1 118 and the collision detection electrode 3 29 and the collision detection electrode 4 210 is recorded as the detection area 5, and the area formed by the collision channel 2 119 and the collision detection electrode 1 27 and the collision detection electrode 2 28 is recorded as the detection area 6.

As shown in FIG15 , due to the effect of the focusing channel 115, the biological particles are focused at a specific position in the channel and pass through the detection area 1; in the detection area 2, the fluids in the sheath liquid channel 1 16 and the sheath liquid channel 2 120 merge into the extrusion channel 117, and the biological particles are thus squeezed and deformed by the fluid; in the detection area 3, since the collision channel 2 119 is vertically connected to the extrusion channel 117, the biological particles are deformed after colliding with the wall, and restore their original shape at the end of the collision channel 2 119 (i.e., the detection area 4). It should be noted that in order to measure the recovery speed of the biological particles after collision deformation, , the detection area three and the detection area four need to be separated by a long distance (50-100 microns); the shape detection electrode one 21, the shape detection electrode two 22, and the shape detection electrode three 23 are used to detect the shape of biological particles, the extrusion detection electrode one 24, the extrusion detection electrode two 25, and the extrusion detection electrode three 26 are used to detect the deformation of biological particles caused by fluid extrusion, the collision detection electrode five 211 and the collision detection electrode six 212 are used to detect the deformation of biological particles caused by collision, and the collision detection electrode seven 213 and the collision detection electrode eight 214 are used to detect the recovery speed of biological particles after deformation. The circuit connection method is shown in Figure 5. The signal generator of the phase-locked amplifier emits an excitation electric field, which is a multi-frequency AC electric field, and is respectively connected to the excitation electrodes of the detection areas one to four, namely, the shape detection electrode two 22, the squeeze detection electrode two 25, the collision detection electrode three 29, and the collision detection electrode six 212. The shape detection electrode one 21 of the detection area one, the squeeze detection electrode one 24 of the detection area two, the collision detection electrode five 211 of the detection area three, and the collision detection electrode four 210 of the detection area five are short-circuited together and then connected to the transimpedance amplifier. The shape detection electrode three 23 of the detection area one, the squeeze detection electrode three 26 of the detection area two, the collision detection electrode eight 214 of the detection area three, and the collision detection electrode one 27 of the detection area five are short-circuited together and then connected to the transimpedance amplifier. The two currents in the transimpedance amplifier are again connected to the phase-locked amplifier to demodulate the weak signal, and finally the morphology and deformation signals of the biological particles are obtained.

As shown in FIG16 , since the excitation electric field is a multi-frequency AC electric field (including a low-frequency electric field f1 and a high-frequency electric field f2 in this embodiment), the impedance sensing signal includes two parts, low frequency and high frequency, namely (1) in FIG9 and (2) in FIG9 . When the biological particles are in the detection area 1, the shape and internal characteristics of the biological particles can be obtained by analyzing the amplitude phase of the impedance signal in the area. When the biological particles are in the detection area 2, the biological particles are elongated in the X-axis direction due to the fluid extrusion. At this time, the disturbance caused by the biological particles in the electric field is greater, so the impedance signal is shown as having a larger amplitude and peak width, etc. By analyzing the amplitude and peak width between the detection area 1 and the detection area 2, the deformation of the biological particles caused by the fluid extrusion can be obtained. Similarly, the biological particles are deformed due to collision in the detection area 3. By analyzing the changes in the impedance signal of the biological particles at various frequencies, the collision deformation information of the biological particles can be obtained. It should be noted that the previous experimental studies have shown that there is a certain difference between the extrusion deformation and the collision deformation of the biological particles. Since the distance between detection area 4 and detection area 3 is specially designed, the biological particles are recovering to their original shape when in detection area 4, and information on the deformation and recovery speed of the biological particles can also be obtained by analyzing the impedance signal. In addition, since detection areas 5 and 6 are symmetrically distributed with detection areas 3 and 4, the effects of biological particles flowing through these areas are the same, so they will not be repeated.

The above description is only a preferred embodiment of the present invention and does not constitute any other form of limitation to the present invention. Any modification or equivalent change made based on the technical essence of the present invention still falls within the scope of protection required by the present invention.

Claims (4)

1. A 3D printed biological particle deformation detection device based on flow impedance technology, comprising a flow channel layer (1) and an electrode layer (2), characterized in that: the flow channel layer (1) and the electrode layer (2) are sealed up and down; divided into four detection areas, which are respectively a detection area for measuring the shape of biological particles, a detection area for measuring the deformation of biological particles caused by fluid compression, a detection area for measuring the deformation of biological particles caused by collision with the flow channel wall, and a detection area for measuring the deformation of biological particles caused by collision with the flow channel wall, wherein one detection area for measuring the shape of biological particles is detection area one, one detection area for measuring the deformation of biological particles caused by fluid compression is detection area two, two detection areas for measuring the deformation of biological particles caused by collision with the flow channel wall are symmetrically arranged, namely detection area three and detection area five, and two detection areas for measuring the deformation of biological particles caused by collision with the flow channel wall are symmetrically arranged, namely detection area four and detection area six; The detection area one is formed by the focusing channel (115) and the shape detection electrode one (21), the shape detection electrode two (22), and the shape detection electrode three (23); the detection area two is formed by the extrusion channel (117) and the extrusion detection electrode one (24), the extrusion detection electrode two (25), and the extrusion detection electrode three (26); the detection area three is formed by the collision channel two (119) and the collision detection electrode five (211), and the collision detection electrode six (212); the detection area four is formed by the collision channel two (119) and the collision detection electrode seven (213), and the collision detection electrode eight (214); the detection area five is formed by the collision channel one (118) and the collision detection electrode three (29), and the collision detection electrode four (210); and the detection area six is formed by the collision channel two (119) and the collision detection electrode one (27), and the collision detection electrode two (28); The flow channel layer (1) is provided with a sample inlet (11), a sheath liquid inlet 1 (12) and a sheath liquid inlet 2 (111), a sample outlet 1 (13) and a sample outlet 2 (17), as well as an electrode hole 1 (14), an electrode hole 2 (15), an electrode hole 3 (16), an electrode hole 4 (18), an electrode hole 5 (19), an electrode hole 6 (110), an electrode hole 7 (112), an electrode hole 8 (113) and an electrode hole 9 (114); the bottom surface of the flow channel layer (1) is provided with a focusing flow channel (115), a sheath liquid flow channel 1 (116) and a sheath liquid flow channel 2 (117). 20), an extrusion flow channel (117), and a collision flow channel 1 (118) and a collision flow channel 2 (119), wherein the sheath liquid flow channel 1 (116) and the sheath liquid flow channel 2 (120) are located on both sides of the extrusion flow channel (117); the collision flow channel 1 (118) and the collision flow channel 2 (119) are directly connected and vertically connected to the extrusion flow channel (117), and when the biological particles enter the collision flow channel 1 (118) or the collision flow channel 2 (119), they collide with the flow channel wall surface to generate deformation, and the biological particles flow out through the collision flow channel 1 (118) or the collision flow channel 2 (119); The electrode layer (2) is provided with a shape detection electrode 1 (21), a shape detection electrode 2 (22) and a shape detection electrode 3 (23), a squeeze detection electrode 1 (24), a squeeze detection electrode 2 (25) and a squeeze detection electrode 3 (26), and a collision detection electrode 1 (27), a collision detection electrode 2 (28), a collision detection electrode 3 (29), a collision detection electrode 4 (210), a collision detection electrode 5 (211), a collision detection electrode 6 (212), a collision detection electrode 7 (213) and a collision detection electrode 8 (214), wherein the shape detection electrode Electrode one (21), shape detection electrode three (23), squeeze detection electrode one (24), squeeze detection electrode three (26), collision detection electrode one (27), collision detection electrode four (210), collision detection electrode five (211) and collision detection electrode eight (214) are sensing electrodes, and shape detection electrode two (22), squeeze detection electrode two (25), collision detection electrode two (28), collision detection electrode three (29), collision detection electrode six (212) and collision detection electrode seven (213) are excitation electrodes; the sizes of all electrodes match the size of biological particles. 2. According to claim 1, a 3D printed biological particle deformation detection device based on flow impedance technology is characterized in that: the focusing flow channel (115), the extrusion flow channel (117), the collision flow channel 1 (118) and the collision flow channel 2 (119) are straight flow channels with rectangular cross-sections. 3. According to claim 1, a 3D printed biological particle deformation detection device based on flow impedance technology is characterized in that: the electrode layer (2) is made of heterogeneous materials by three-dimensional printing, the electrode part is a conductive material, and the non-electrode part is an insulating material, and the flow channel layer (1) is made of non-conductive materials by three-dimensional printing. 4. A 3D printed biological particle deformation detection device based on flow impedance technology according to claim 1, characterized in that: when the flow channel layer (1) and the electrode layer (2) are assembled from top to bottom, they are sealed by a hot pressing process or by a sealant.