CN111815700B - Device and method for measuring mechanical properties of erythrocytes - Google Patents

Device and method for measuring mechanical properties of erythrocytes Download PDF

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CN111815700B
CN111815700B CN202010034302.6A CN202010034302A CN111815700B CN 111815700 B CN111815700 B CN 111815700B CN 202010034302 A CN202010034302 A CN 202010034302A CN 111815700 B CN111815700 B CN 111815700B
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interdigital transducer
red blood
channel
straight channel
blood cells
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CN111815700A (en
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陈希
刘洋
陈赵江
蔡建程
丁宇
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Zhejiang Normal University CJNU
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    • G—PHYSICS
    • G06—COMPUTING; CALCULATING OR COUNTING
    • G06T—IMAGE DATA PROCESSING OR GENERATION, IN GENERAL
    • G06T7/00—Image analysis
    • G06T7/60—Analysis of geometric attributes
    • G06T7/62—Analysis of geometric attributes of area, perimeter, diameter or volume
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M33/00—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus
    • C12M33/04—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus by injection or suction, e.g. using pipettes, syringes, needles
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/12—Means for regulation, monitoring, measurement or control, e.g. flow regulation of temperature
    • G—PHYSICS
    • G06—COMPUTING; CALCULATING OR COUNTING
    • G06T—IMAGE DATA PROCESSING OR GENERATION, IN GENERAL
    • G06T2207/00—Indexing scheme for image analysis or image enhancement
    • G06T2207/10—Image acquisition modality
    • G06T2207/10056—Microscopic image
    • G06T2207/10061—Microscopic image from scanning electron microscope
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

The invention discloses a device and a method for measuring the mechanical properties of erythrocytes, comprising an injection pump, a power adjustable signal generator, a surface acoustic wave driving array module, a temperature control module, a microscope, a CCD camera and a computer; the surface acoustic wave driving array module comprises a piezoelectric substrate, a first interdigital transducer, a second interdigital transducer, a third interdigital transducer, a fourth interdigital transducer, a fifth interdigital transducer, a sixth interdigital transducer and a micro-channel bonded on the piezoelectric substrate, wherein the first interdigital transducer, the second interdigital transducer, the third interdigital transducer, the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are deposited on the upper surface of the piezoelectric substrate; the piezoelectric substrate is tightly attached to the upper surface of the temperature control module; the red blood cells flowing into the micro-channel deform under the drive of the surface acoustic wave generated by the interdigital transducer, and the mechanical characteristics are calculated by analyzing the deformation parameters of the red blood cells shot by the CCD camera and using a mathematical model. The device and the method can realize the measurement of the mechanical properties of single red blood cells smaller than 10 microns and simultaneously reflect the elastic mechanical properties and the tension properties.

Description

Device and method for measuring mechanical properties of erythrocytes
Technical Field
The invention relates to the field of signal detection and physical parameter measurement, in particular to a device and a method for measuring mechanical properties of erythrocytes.
Background
Erythrocytes are mainly composed of phospholipid bilayer, membrane contractile protein and skeletal network. Lipid bilayers are very resistant to changes in local surface area, but the elasticity of the cytoskeleton allows the erythrocytes to withstand large tensile deformations while maintaining the structural integrity of the membrane. Erythrocyte deformation can affect the ability of cells to release various molecules and regulate their concentration in the blood, particularly when erythrocytes are subjected to strong deformation by surrounding fluids, they release Adenosine Triphosphate (ATP) and Adenosine Diphosphate (ADP). ATP is an energy source for intracellular functions, and extracellular ATP plays an important role as a signaling molecule in a variety of physiological processes. Furthermore, ADP is a factor of primary aggregation of platelets. Flow-induced deformation may also lead to rupture of the red blood cells, i.e. hemolysis. There is a causal relationship between the stress level of the membrane and hemolysis.
Since the red cell membrane is very thin, in the membrane model, the stress is integrated along the wall thickness, usually expressed in tension. Meanwhile, cytoskeleton in red blood cells and the like have elastic force. Thus, erythrocytes have both surface tension and elastic forces, collectively referred to as mechanical properties. Measurement of erythrocyte mechanical properties is critical to understanding the physiology and pathology of microcirculation. Many literature studies indicate that physiological diseases, such as uremic patients, hypertension, diabetes mellitus, retinopathy and the like, can be reflected on the mechanical properties of erythrocytes. Therefore, the measurement of the mechanical properties of the erythrocytes is of practical clinical significance.
The method for measuring the surface tension of a liquid is classified into a hydrostatic method and a kinetic method. The statics method includes capillary ascending method, drop volume method, du Muy ring method, wilhelmy disk method, hanging drop method, rotary drop method, and maximum bubble pressure method; the dynamic method is an oscillation jet method and a capillary wave method. Wherein the capillary rise method and the maximum bubble pressure method cannot be used to measure the liquid-liquid interfacial tension. The Wilhelmy disk method, maximum bubble pressure method, oscillating jet method, capillary wave method can be used to determine dynamic surface tension. Because the dynamics method is complex, the testing precision is not high, and the prior data acquisition and processing means are not advanced enough, so that the successful application examples of the measurement method are few. Therefore, in actual production, a statics measurement method is often used. The above methods have difficulty in achieving surface tension measurements for drops below 10 microns, while red blood cell sizes are much smaller than those of conventional drops, typically around 7 microns. In addition, erythrocyte membranes are fragile, and rupture of the membrane is easily caused by the above-mentioned various methods, so that measurement of erythrocyte membrane tension is difficult to achieve, and elastomechanical properties cannot be simultaneously reacted. Peng Hange (Peng Hange, zhou Tiean, tan Chengfang, etc.) QCM monitors the viscoelastic response of primary cardiomyocytes in rats under the action of different concentrations of myodynamic drugs [ J ]. Laser biologies report, 2019, 28 (3): 239-244.) it is proposed to use quartz crystal microbalances (quartz crystal microbalance, QCM) to monitor mechanical indicators such as cell viscoelastic parameters, but not to detect single cells, while only measuring viscoelastic properties but not to react to cell membrane tension. Yuan Weimo (Yuan Weimo, xue Chundong, liu Bo. A microfluidic chip for measuring single cell elastic modulus with high throughput [ J ]. Beijing biomedical engineering, 2019, 38 (5): 450-456.) although measurement of single cell elastic modulus can be achieved with a microfluidic chip, it cannot reflect cell membrane tension characteristics. Yang Qian (Yang Qian, sun Runan, zhang Jing, etc.) the detection of biomechanical properties of infection of erythrocyte membranes with Salmonella paratyphi B in combination with AFM [ J ]. Chinese science: life sciences, 2014,44 (2): 197-207.) atomic force microscopy and diffraction microscopy are used to achieve mechanical property measurements of erythrocyte membranes, but atomic force microscopy is too expensive.
Disclosure of Invention
The invention aims to overcome the defects of the existing measurement method, realize the measurement of the mechanical properties of single red blood cells with micrometer scale, especially less than 10 micrometers, and the measured values can simultaneously reflect the elastic mechanical properties and the tension properties of the single red blood cells. The invention provides a device and a method for measuring mechanical properties of erythrocytes by combining the prior art. The technical scheme of the invention is as follows:
the measuring device comprises an injection pump, a power adjustable signal generator, a surface acoustic wave driving array module, a temperature control module, a microscope, a CCD camera and a computer.
The surface acoustic wave driving array module comprises a piezoelectric substrate, a first interdigital transducer, a second interdigital transducer, a third interdigital transducer, a fourth interdigital transducer, a fifth interdigital transducer, a sixth interdigital transducer and a micro-channel bonded on the piezoelectric substrate, wherein the first interdigital transducer, the second interdigital transducer, the third interdigital transducer, the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are deposited on the upper surface of the piezoelectric substrate.
The piezoelectric substrate is made of 128-degree Y-X lithium niobate material and is tightly attached to the upper surface of the temperature control module.
The microchannel includes a microchannel inlet, a wedge-shaped channel, a straight channel, and a microchannel outlet. The injection pump is connected with the wedge-shaped channel through the micro-channel inlet, the other end of the wedge-shaped channel is connected with the straight channel, and the other end of the straight channel is a micro-channel outlet. The diameter of the through passage is 8-11 micrometers.
The first interdigital transducer, the second interdigital transducer and the third interdigital transducer are arranged on one side of the straight channel, and the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are arranged on the other side of the straight channel. The first interdigital transducer, the second interdigital transducer, the third interdigital transducer, the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are all in a fan ring shape, the inner arc aperture is 15-50 microns and is close to the straight channel, and the outer arc aperture is 200-250 microns and is far away from the straight channel. All interdigital transducers have the same working frequency and the working frequency band is 20-400 MHz. The number of the interdigital transducers of the first interdigital transducer and the fourth interdigital transducer is 10-30 pairs, and the interdigital transducers are symmetrically distributed on two sides of the straight channel; the number of the fork indexes of the second interdigital transducer and the fifth interdigital transducer is the same and is 30-50 pairs, and the second interdigital transducer and the fifth interdigital transducer are symmetrically distributed on two sides of the straight channel; the number of the fork indexes of the third interdigital transducer and the sixth interdigital transducer is the same and is 50-70 pairs, and the third interdigital transducer and the sixth interdigital transducer are symmetrically distributed on two sides of the straight channel; the first interdigital transducer, the second interdigital transducer and the third interdigital transducer are horizontally arranged in a row, and the inner arcs are on the same horizontal line and have equal center-to-center distances; the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are horizontally arranged in a row, and the inner arcs are on the same horizontal line and have equal center-to-center distances.
The two poles of the first interdigital transducer, the second interdigital transducer, the third interdigital transducer, the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are respectively and simultaneously connected with the positive pole and the negative pole of the power adjustable signal generator.
The microscope is connected with the CCD camera, and the CCD camera is connected with the computer.
The method for measuring the mechanical properties of the erythrocyte membrane comprises the following steps:
step 1, loading blood into a centrifuge tube, centrifuging in the centrifuge at a rotating speed of 3000 rpm for 5 minutes, and sucking and throwing away supernatant after centrifuging; repeating the above operation twice, dividing the obtained erythrocyte solution into two parts, diluting one part of the erythrocyte solution after centrifugation with 0.9% NaCl solution to obtain erythrocyte solution with pressure product of 5%, and storing the other part of erythrocyte solution in the environment of 37 ℃ without dilution;
step 2, respectively testing the viscosity mu of undiluted erythrocyte solution by a digital viscometer under the environment of 37 DEG C r And a red blood cell solution viscosity mu of 5% by volume after dilution c Relative viscosity μ' =μ is also noted r /μ c ；
Step 3, setting the temperature of the temperature control module 5 to be 37 ℃, and pushing the erythrocyte solution with the pressure product of 5% to the micro-channel through the injection pump 1;
and 4, slowly flowing the red blood cells in the wedge-shaped channel into the straight channel and flowing out of the straight channel through the micro-channel outlet, and shooting the red blood cells by a CCD camera through a microscope and transmitting the red blood cells to a computer.
Step 5, analyzing and measuring related parameters through video image software;
when the red blood cells just enter the straight channel, if the red blood cells are round, the measured sphere radius is recorded as r 0 ；
When the red blood cells are at the vertical crossing position of the straight channel and the connecting lines of the first interdigital transducer and the fourth interdigital transducer, the position is marked as x 1 The time is recorded as t 1 At this time, the red blood cell shape is elliptical, and the measured long half axis is p 1 And short half shaft q 1 Erythrocyte deformation parameter D 1 Denoted as D 1 ＝(p 1 -q 1 )/(p 1 +q 1 )；
When the red blood cells are at the vertical crossing position of the straight channel and the connecting lines of the second interdigital transducer and the fifth interdigital transducer, the position is marked as x 2 The time is recorded as t 2 The method comprises the steps of carrying out a first treatment on the surface of the The elliptic erythrocyte long half axis is measured to be p 2 And short half shaft q 2 Erythrocyte deformation parameter D 2 Denoted as D 2 ＝(p 2 -q 2 )/(p 2 +q 2 )；
When the red blood cells are at the vertical crossing position of the straight channel and the connecting line of the third interdigital transducer and the sixth interdigital transducer, the position is marked as x 3 The time is recorded as t 3 The method comprises the steps of carrying out a first treatment on the surface of the The elliptic erythrocyte long half axis is measured to be p 3 And short half shaft q 3 ；
When the red blood cells flow through the vertical crossing position of the connecting lines of the straight channel, the third interdigital transducer and the sixth interdigital transducer and the distance from the vertical crossing position is equal to the center-to-center distance between the fifth interdigital transducer and the sixth interdigital transducer, the vertical crossing position is marked as x 4 The time is recorded as t 4 And the elliptic erythrocyte long half axis is measured as p 4 And short half shaft q 4 The method comprises the steps of carrying out a first treatment on the surface of the This isAfter that, when the red blood cells finally become round to reach equilibrium, the radius is marked as r e ；
Distance Δx=x 4 -x 3 ＝x 3 -x 2 ＝x 2 -x 1 The method comprises the steps of carrying out a first treatment on the surface of the Erythrocyte deformation amount Δd 21 ＝D 2 -D 1 The method comprises the steps of carrying out a first treatment on the surface of the Flow velocity v 21 ＝Δx/Δt 21 At Deltat for erythrocytes 21 ＝t 2 -t 1 A distance Δx that flows over time; flow velocity v 32 ＝Δx/Δt 32 At Deltat for erythrocytes 32 ＝t 3 -t 2 A distance Δx that flows over time; α=Δv/Δx= (v 21 -v 32 ) And/Δx is the change in flow velocity over the distance Δx.
Step 6, if the red blood cells enter the straight channel and are round, the mechanical properties gamma of the red blood cells, including elastic force and tension, are calculated by the following formula:
wherein, κ= (2μ ' +3) (19 μ ' +16)/40 (μ ' +1);
if the red blood cells enter the straight channel and are non-circular, the mechanical properties gamma of the red blood cells, including elastic force and tension, are calculated by the following formula:
the beneficial effects of the invention are as follows: measurement of mechanical properties of individual erythrocytes on a micrometer scale, in particular less than 10 micrometers, can be achieved; the measured value can simultaneously reflect the elastic mechanical property and the tension property of the single red blood cell; compared with atomic force microscope, the cost is greatly reduced.
Drawings
In order to more clearly illustrate the embodiments of the invention or the technical solutions in the prior art, the drawings that are required in the embodiments or the description of the prior art will be briefly described, it being obvious that the drawings in the following description are only some embodiments of the invention, and that other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a schematic diagram of a measuring device;
fig. 2 is a side view of the structure of the measuring device.
Detailed Description
The invention is described in detail below with reference to the accompanying drawings, and the device for measuring the mechanical properties of the erythrocytes comprises the following technical scheme:
as shown in fig. 1 and 2, the measuring device comprises an injection pump 1, a power adjustable signal generator 2, a surface acoustic wave driving array module 3, a temperature control module 5, a microscope 6, a CCD camera 7 and a computer 8.
The saw driven array module 3 includes a piezoelectric substrate 37, a first interdigital transducer 31, a second interdigital transducer 32, a third interdigital transducer 33, a fourth interdigital transducer 34, a fifth interdigital transducer 35, a sixth interdigital transducer 36, and a micro channel 38 bonded on the piezoelectric substrate 37, which are deposited on the upper surface of the piezoelectric substrate 37.
The piezoelectric substrate 37 is made of 128-degree Y-X lithium niobate material and is tightly attached to the upper surface of the temperature control module 5.
The microchannel 38 includes a microchannel inlet 383, a wedge-shaped channel 381, a straight channel 382, and a microchannel outlet 384. The injection pump 1 is connected with a wedge-shaped channel 381 through a micro-channel inlet 383, the other end of the wedge-shaped channel 381 is connected with a straight channel 382, and the other end of the straight channel 382 is a micro-channel outlet 384. The diameter of the straight channel 382 is 8-11 microns.
The first interdigital transducer 31, the second interdigital transducer 32 and the third interdigital transducer 33 are on one side of the straight channel 382, and the fourth interdigital transducer 34, the fifth interdigital transducer 35 and the sixth interdigital transducer 36 are on the other side of the straight channel 382. The first interdigital transducer 31, the second interdigital transducer 32, the third interdigital transducer 33, the fourth interdigital transducer 34, the fifth interdigital transducer 35 and the sixth interdigital transducer 36 are all in a sector ring shape, the inner arc aperture is 15-50 microns and is close to the straight channel 382, and the outer arc aperture is 200-250 microns and is far away from the straight channel 382. All interdigital transducers have the same working frequency and the working frequency band is 20-400 MHz. The first interdigital transducer 31 and the fourth interdigital transducer 34 have the same interdigital numbers, are 10-30 pairs, and are symmetrically distributed on two sides of the straight channel 382; the number of the interdigital transducers 32 and 35 is 30-50 pairs, and the interdigital transducers are symmetrically distributed on two sides of the straight channel 382; the number of the interdigital transducers 33 and 36 is 50-70 pairs, and the interdigital transducers are symmetrically distributed on two sides of the straight channel 382; the first interdigital transducer 31, the second interdigital transducer 32 and the third interdigital transducer 33 are horizontally arranged in a row, and the inner arcs are on the same horizontal line and have equal center-to-center distances; the fourth interdigital transducer 34, the fifth interdigital transducer 35 and the sixth interdigital transducer 36 are horizontally arranged in a row, and the inner arcs are on the same horizontal line and have equal center-to-center distances.
The two poles of the first interdigital transducer 31, the second interdigital transducer 32, the third interdigital transducer 33, the fourth interdigital transducer 34, the fifth interdigital transducer 35 and the sixth interdigital transducer 36 are respectively connected with the positive pole and the negative pole of the power adjustable signal generator 2 at the same time.
The microscope 6 is connected with the CCD camera 7, and the CCD camera 7 is connected with the computer 8.
The method for measuring the mechanical properties of the erythrocyte membrane comprises the following steps:
step 1, loading blood into a centrifuge tube, centrifuging in the centrifuge at a rotating speed of 3000 rpm for 5 minutes, and sucking and throwing away supernatant after centrifuging; repeating the above operation twice, dividing the obtained erythrocyte solution into two parts, diluting one part of the erythrocyte solution after centrifugation with 0.9% NaCl solution to obtain erythrocyte solution with pressure product of 5%, and storing the other part of erythrocyte solution in the environment of 37 ℃ without dilution;
step 2, respectively testing the viscosity mu of undiluted erythrocyte solution by a digital viscometer under the environment of 37 DEG C r And a red blood cell solution viscosity mu of 5% by volume after dilution c Relative viscosity μ' =μ is also noted r /μ c ；
Step 3, setting the temperature of the temperature control module 5 to be 37 ℃, and pushing the erythrocyte solution with the pressure product of 5% to the micro-channel 38 through the injection pump 1;
in step 4, the red blood cells 4 in the wedge-shaped channel 381 slowly flow into the straight channel 382 and flow out through the micro-channel outlet 384, and the process is photographed by the CCD camera 7 through the microscope 6 and transmitted to the computer 8.
Step 5, analyzing and measuring related parameters through video image software;
when the red blood cells 4 just enter the straight channel 382, the measured sphere radius is denoted as r if the red blood cells are round 0 ；
When the red blood cells 4 are at the vertical crossing position 41 of the straight channel 382 and the connecting lines of the first 31 and fourth 34 interdigital transducers, the position is denoted as x 1 The time is recorded as t 1 At this time, the red blood cell shape is elliptical, and the measured long half axis is p 1 And short half shaft q 1 Erythrocyte deformation parameter D 1 Denoted as D 1 ＝(p 1 -q 1 )/(p 1 +q 1 )；
When the red blood cells 4 are at the vertical crossing position 42 of the straight channel 382 and the connecting lines of the second interdigital transducer 32 and the fifth interdigital transducer 35, the position is denoted as x 2 The time is recorded as t 2 The method comprises the steps of carrying out a first treatment on the surface of the The elliptic erythrocyte long half axis is measured to be p 2 And short half shaft q 2 Erythrocyte deformation parameter D 2 Denoted as D 2 ＝(p 2 -q 2 )/(p 2 +q 2 )；
When the red blood cells 4 are at the vertical crossing position 43 of the straight channel 382 and the connecting lines of the third 33 and sixth 36 interdigital transducers, the position is denoted as x 3 The time is recorded as t 3 The method comprises the steps of carrying out a first treatment on the surface of the The elliptic erythrocyte long half axis is measured to be p 3 And short half shaft q 3 ；
When the red blood cells 4 flow through the straight channel 382 and the vertical crossing position 43 of the connection lines of the third 33 and sixth 36 interdigital transducers, and the distance from this position 43 is equal to the center-to-center spacing of the fifth 35 and sixth 36 interdigital transducers, this position 44 is denoted as x 4 The time is recorded as t 4 And the elliptic erythrocyte long half axis is measured as p 4 And short half shaft q 4 The method comprises the steps of carrying out a first treatment on the surface of the Thereafter, the red blood cells 4 eventually become roundedAt equilibrium, its radius is denoted as r e ；
Distance Δx=x 4 -x 3 ＝x 3 -x 2 ＝x 2 -x 1 The method comprises the steps of carrying out a first treatment on the surface of the Erythrocyte deformation amount Δd 21 ＝D 2 -D 1 The method comprises the steps of carrying out a first treatment on the surface of the Flow velocity v 21 ＝Δx/Δt 21 At Deltat for erythrocytes 21 ＝t 2 -t 1 A distance Δx that flows over time; flow velocity v 32 ＝Δx/Δt 32 At Deltat for erythrocytes 32 ＝t 3 -t 2 A distance Δx that flows over time; α=Δv/Δx= (v 21 -v 32 ) Δx, which is the change in flow velocity over a distance Δx;
step 6, if the red blood cells 4 enter the straight channel 382 and are circular, the mechanical properties γ of the red blood cells, including elastic force and tension, are calculated by the following formula:
wherein, κ= (2μ ' +3) (19 μ ' +16)/40 (μ ' +1);
if the red blood cells 4 enter the straight channel 382 in a non-circular shape, the mechanical properties of the red blood cells, gamma, including elastic force and tension, are calculated by the following formula:
the above is merely a specific embodiment of the present invention, but the scope of the present invention is not limited thereto, and any changes or substitutions that do not undergo the inventive work should be covered in the scope of the present invention. Therefore, the protection scope of the present invention should be subject to the protection scope defined by the claims.

Claims (2)

1. A device for measuring mechanical properties of erythrocytes, characterized by: the system comprises an injection pump, a power adjustable signal generator, a surface acoustic wave driving array module, a temperature control module, a microscope, a CCD camera and a computer;
the surface acoustic wave driving array module comprises a piezoelectric substrate, a first interdigital transducer, a second interdigital transducer, a third interdigital transducer, a fourth interdigital transducer, a fifth interdigital transducer, a sixth interdigital transducer and a micro-channel bonded on the piezoelectric substrate, wherein the first interdigital transducer, the second interdigital transducer, the third interdigital transducer, the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are deposited on the upper surface of the piezoelectric substrate; the piezoelectric substrate is made of 128-degree Y-X lithium niobate material and is tightly attached to the upper surface of the temperature control module;
the micro-channel comprises a micro-channel inlet, a wedge-shaped channel, a straight channel and a micro-channel outlet; the injection pump is connected with the wedge-shaped channel through the micro-channel inlet, the other end of the wedge-shaped channel is connected with the straight channel, and the other end of the straight channel is a micro-channel outlet; the diameter of the straight channel is 8-11 micrometers;
the first interdigital transducer, the second interdigital transducer and the third interdigital transducer are arranged on one side of a straight channel, and the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are arranged on the other side of the straight channel; the first interdigital transducer, the second interdigital transducer, the third interdigital transducer, the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are all in a sector ring shape, the inner arc aperture is 15-50 microns and is close to the straight channel, and the outer arc aperture is 200-250 microns and is far away from the straight channel; all interdigital transducers have the same working frequency and the working frequency band is 20-400 MHz; the number of the interdigital transducers of the first interdigital transducer and the fourth interdigital transducer is 10-30 pairs, and the interdigital transducers are symmetrically distributed on two sides of the straight channel; the number of the fork indexes of the second interdigital transducer and the fifth interdigital transducer is the same and is 30-50 pairs, and the second interdigital transducer and the fifth interdigital transducer are symmetrically distributed on two sides of the straight channel; the number of the fork indexes of the third interdigital transducer and the sixth interdigital transducer is the same and is 50-70 pairs, and the third interdigital transducer and the sixth interdigital transducer are symmetrically distributed on two sides of the straight channel; the first interdigital transducer, the second interdigital transducer and the third interdigital transducer are horizontally arranged in a row, and the inner arcs are on the same horizontal line and have equal center-to-center distances; the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are horizontally arranged in a row, and the inner arcs are on the same horizontal line and have equal center-to-center distances;
the two poles of the first interdigital transducer, the second interdigital transducer, the third interdigital transducer, the fourth interdigital transducer, the fifth interdigital transducer and the sixth interdigital transducer are respectively connected with the positive pole and the negative pole of the power adjustable signal generator at the same time;
the microscope is connected with the CCD camera, and the CCD camera is connected with the computer.
2. A method for measuring mechanical properties of erythrocytes, applied to the device of claim 1, comprising the steps of:
step 1, loading blood into a centrifuge tube, centrifuging in the centrifuge at a rotating speed of 3000 rpm for 5 minutes, and sucking and throwing away supernatant after centrifuging; repeating the above operation twice, dividing the obtained erythrocyte solution into two parts, diluting one part of the erythrocyte solution after centrifugation with 0.9% NaCl solution to obtain erythrocyte solution with pressure product of 5%, and storing the other part of erythrocyte solution in the environment of 37 ℃ without dilution;
step 2, respectively testing the viscosity mu of undiluted erythrocyte solution by a digital viscometer under the environment of 37 DEG C r And a red blood cell solution viscosity mu of 5% by volume after dilution c Relative viscosity μ' =μ is also noted r /μ c ；
Step 3, setting the temperature of the temperature control module to be 37 ℃, and pushing the erythrocyte solution with the pressure product of 5% to the micro-channel through the injection pump 1;
step 4, the red blood cells in the wedge-shaped channel slowly flow into the straight channel and flow out through the micro-channel outlet, and the process is photographed by a CCD camera through a microscope and transmitted to a computer;
step 5, analyzing and measuring related parameters through video image software;
when the red blood cells just enter the straight channel, if the red blood cells are round, the measured sphere radius is recorded as r 0 ；
When the red blood cells are at the vertical crossing position of the straight channel and the connecting lines of the first interdigital transducer and the fourth interdigital transducer, the position is marked as x 1 The time is recorded as t 1 At this time, the red blood cell shape is elliptical, and the measured long half axis is p 1 And short half shaft q 1 Erythrocyte deformation parameter D 1 Denoted as D 1 ＝(p 1 -q 1 )/(p 1 +q 1 )；
When the red blood cells are at the vertical crossing position of the straight channel and the connecting lines of the second interdigital transducer and the fifth interdigital transducer, the position is marked as x 2 The time is recorded as t 2 The method comprises the steps of carrying out a first treatment on the surface of the The elliptic erythrocyte long half axis is measured to be p 2 And short half shaft q 2 Erythrocyte deformation parameter D 2 Denoted as D 2 ＝(p 2 -q 2 )/(p 2 +q 2 )；
When the red blood cells are at the vertical crossing position of the straight channel and the connecting line of the third interdigital transducer and the sixth interdigital transducer, the position is marked as x 3 The time is recorded as t 3 The method comprises the steps of carrying out a first treatment on the surface of the The elliptic erythrocyte long half axis is measured to be p 3 And short half shaft q 3 ；
When the red blood cells flow through the vertical crossing position of the connecting lines of the straight channel, the third interdigital transducer and the sixth interdigital transducer and the distance from the vertical crossing position is equal to the center-to-center distance between the fifth interdigital transducer and the sixth interdigital transducer, the vertical crossing position is marked as x 4 The time is recorded as t 4 And the elliptic erythrocyte long half axis is measured as p 4 And short half shaft q 4 The method comprises the steps of carrying out a first treatment on the surface of the Thereafter, when the red blood cells eventually become rounded to reach equilibrium, their radius is noted as r e ；
Distance Δx=x 4 -x 3 ＝x 3 -x 2 ＝x 2 -x 1 The method comprises the steps of carrying out a first treatment on the surface of the Erythrocyte deformation amount Δd 21 ＝D 2 -D 1 The method comprises the steps of carrying out a first treatment on the surface of the Flow velocity v 21 ＝Δx/Δt 21 At Deltat for erythrocytes 21 ＝t 2 -t 1 A distance Δx that flows over time; flow velocity v 32 ＝Δx/Δt 32 At Deltat for erythrocytes 32 ＝t 3 -t 2 A distance Δx that flows over time; α=Δv/Δx= (v 21 -v 32 ) Δx, which is the change in flow velocity over a distance Δx;
step 6, if the red blood cells enter the straight channel and are round, the mechanical properties gamma of the red blood cells, including elastic force and tension, are calculated by the following formula:
wherein, κ= (2μ ' +3) (19 μ ' +16)/40 (μ ' +1);
if the red blood cells enter the straight channel and are non-circular, the mechanical properties gamma of the red blood cells, including elastic force and tension, are calculated by the following formula:
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