CN111693600A - Method and device for promoting biomolecule sensing and biomolecule sensing system - Google Patents
Method and device for promoting biomolecule sensing and biomolecule sensing system Download PDFInfo
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Abstract
A method of facilitating biomolecule sensing, comprising: driving the resonator to produce an acoustic fluid in the liquid in the gigahertz (GHz) range; and driving the biological molecules in the flow field to be transported to a sensing interface of the biosensor in the action area of the acoustic fluid through the acoustic fluid in the GHz level. There is also provided an apparatus for facilitating sensing of biomolecules, comprising: an acoustic fluid system section including a resonator for generating acoustic fluid at GHz level; a cylindrical cavity located within the acoustic fluid action region of the resonator for limiting the range of the acoustic fluid field and for accommodating the sensing interface of the biomolecular sensor. A biomolecule sensing system is also provided. With the application, the biomolecule sensing function of the sensor can be promoted.
Description
Technical Field
The present application relates to the field of biomolecule detection technologies, and in particular, to a method and an apparatus for facilitating biomolecule sensing and a biomolecule sensing system.
Background
Biomolecule detection, such as tumor markers, detection of molecules such as specific DNA sequences, etc., plays an important role in the fields of cell function research, early detection and clinical application of diseases such as cancer, etc., and research and development of molecular drugs, etc.
Although great development and progress have been made in the research field, the biosensor has been slowly developed in terms of practical application. Among them, the practical requirements of enhancing molecular adsorption and reducing non-specific adsorption make the applicability of the sensor face a great challenge. For example, in practical detection, the performance of the sensor is limited by the diffusion of biomolecules at a solid-liquid interface, so that the sensor faces practical problems of low molecular adsorption efficiency, slow detection speed, difficulty in realizing detection of trace biomolecules in a complex sample and the like; non-specific adsorption can produce strong background noise, thereby affecting the detection limit, sensitivity, selectivity and device repeatability of target protein detection or producing false positive signals.
The existing methods for enhancing sensing, such as methods for driving fluid by physical fields of light, electricity and the like, have the characteristics of poor biocompatibility, complex equipment, strict requirements on the chargeability of biomolecules or the transparency of solution and the like, so that the methods lack controllability, universality, high efficiency and the like. Therefore, a system with biocompatibility, simple operation, miniaturization and modularity for enhancing molecular adsorption is needed to be used as a multifunctional platform for improving molecular adsorption characteristics and further improving the biomolecule detection technology based on affinity detection.
Disclosure of Invention
It is therefore one of the primary objectives of the present application to provide a method and apparatus for facilitating sensing of biomolecules and a biomolecule sensing system, so as to facilitate the biomolecule sensing function of the sensing interface through acoustic fluid.
The present application provides a method of facilitating biomolecule sensing, comprising:
driving the resonator to produce an acoustic fluid in the liquid in the gigahertz (GHz) range;
and driving the biological molecules in the flow field to be transported to a sensing interface of the biosensor in the action area of the acoustic fluid through the acoustic fluid in the GHz level.
Therefore, the application provides additional fluid driving force for the biomolecule adsorption process through GHz acoustic fluid, so that sufficient mixing of target molecules in a solution is realized, biomolecules in a depletion layer are effectively supplemented in the mixing process, the barrier effect of the depletion layer is broken, and therefore molecules near the active surface of the sensor keep sufficient reaction concentration. Particularly, when the driving mode is the impact mode, the fluid mode of directionally driving the biomolecules breaks through the solid-liquid diffusion limitation in the sensing process, shortens the transportation time of the biomolecules to the sensing surface, finally can enhance the molecular adsorption quantity, improve the detection sensitivity and the detection speed, and reduce the detection limit. Moreover, the multifunctional platform has strong biocompatibility, simplifies the equipment structure, is beneficial to miniaturization and modularization, and can be used as a multifunctional platform.
Optionally, the method further includes: and driving the biological molecules to perform fixed-point, quantitative and/or graphical modification on the sensing interface of the sensor by controlling the relative position and/or distance between the sensing interface and the resonator.
Due to the three-dimensional shape of the GHz acoustic fluid and the characteristic of high efficiency and controllability of the fluid mode, fixed-point modification and graphical modification of biomolecules on a sensing interface of the sensor are respectively realized by adjusting the relative position and distance between the GHz bulk acoustic wave resonator and the sensor, so that a more accurate sensing detection technology can be realized.
Optionally, the driving is performed in a manner that the resonator is directly opposite to the sensing interface.
From the above, in the direct alignment mode, under the action of the GHz acoustic fluid impact mode, the target molecules near the sensing interface of the sensor are subjected to a positive pressure, that is, the target molecules are driven directionally, so that the path time of the target molecules to the sensing interface of the sensor is shortened, and the absorption of the target molecules by the active surface of the sensor is further promoted. Therefore, under the synergistic effect of the enhanced mixing and directional driving of GHz acoustic fluid, the binding process of protein molecules is remarkably improved.
Optionally, the method further includes: the ability of the drive is controlled by controlling the duty cycle, signal strength and/or pulse period parameters of the pulsed excitation signal driving the resonator.
Therefore, the driving capability can be flexibly controlled through parameter adjustment, so that the driving force is controllable, technical support is further provided for fixed-point and quantitative modification, and a more accurate sensing detection technology can be realized.
Optionally, the method further includes: and removing the non-specific adsorbate on the sensor interface by controlling the position between the sensor interface and the resonator to be in a non-opposite mode.
Due to the characteristics of three-dimensional shape of GHz acoustic fluid and high efficiency and controllability of a fluid mode, the double functions of enhancing molecular adsorption and removing nonspecific adsorption can be realized by adjusting the relative position and distance between a GHz bulk acoustic wave resonator and a sensor, and the adsorption of nonspecific adsorbates on a sensor interface is reduced by removing nonspecific adsorption, so that the sensor interface can have more spaces to adsorb target biomolecules to be detected, and the accuracy and the sensitivity of sensing detection are improved.
The present application also provides an apparatus for facilitating sensing of biomolecules, comprising:
an acoustic fluid system section including a resonator for generating acoustic fluid at GHz level;
a cylindrical cavity located within the acoustic fluid action region of the resonator for limiting the range of the acoustic fluid field and for accommodating the sensing interface of the biomolecular sensor.
Therefore, the application provides additional directional fluid driving force for the biomolecule adsorption process through GHz acoustic fluid, so that sufficient mixing of target molecules in a solution is realized, biomolecules in a depletion layer are effectively supplemented in the mixing process, the barrier effect of the depletion layer is broken, and therefore molecules near the active surface of the sensor keep sufficient reaction concentration. Particularly, when the driving mode is the impact mode, the solid-liquid diffusion limitation in the sensing process is broken through, the supplement of molecules under a body phase is realized, the molecular adsorption quantity can be enhanced finally, the detection sensitivity and the detection speed are improved, and the detection limit is reduced. Moreover, the multifunctional platform has strong biocompatibility, simplifies the equipment structure, is beneficial to miniaturization and modularization, and can be used as a multifunctional platform.
Optionally, the method further includes: a three-dimensional displacement device carrying the resonator or fitted with the biomolecular sensor for controlling the relative position and/or distance between the resonator and the sensing interface.
By the above method, fixed-point modification and graphical modification of biomolecules on the sensing interface of the sensor are respectively realized by adjusting the relative position and distance between the GHz bulk acoustic wave resonator and the sensor, so that a more accurate sensing detection technology can be realized.
Optionally, the side wall of the cavity is transparent.
By the above, the transparent arrangement facilitates observation by an image acquisition device, such as a camera, or a microscope, etc. The observations include observations of relative position, distance, observation of acoustic fluid, etc., between the resonator 504 and the sensing interface of the sensor 508.
Optionally, the acoustic fluid system part comprises: the device comprises a central processing unit, a signal generator, a power amplifier and the resonator which are electrically connected in sequence.
The application also provides a biomolecule sensing system, which comprises a biomolecule sensor and the device in any one of the above technical schemes, wherein the sensing interface of the biomolecule sensor detachably extends into the cavity of the device for promoting biomolecule sensing.
By above, through the sensing interface of the biomolecule sensor and the cavity of the device for promoting the biomolecule sensing can be detachably arranged, so that the device for promoting the biomolecule sensing and the biomolecule sensor can be more flexibly arranged, namely, can be directly integrated, and can also be detachably arranged.
In summary, the present application can solve the following problems:
the GHz acoustic fluid is used together with other sensing systems, so that the solid-liquid diffusion limitation in the sensing process is broken, the molecular adsorption quantity is enhanced, the detection sensitivity and the detection speed of the sensing system are improved, and the detection limit is reduced. Carrying out fixed-point modification on a sensing interface of the sensor by adopting an acoustic fluid system; and realizing the graphical modification of the biological molecules by displacement control. The same acoustic fluid realizes the double functions of enhancing molecular adsorption and removing non-specific adsorption.
Drawings
FIG. 1 is a schematic diagram of a two-chamber model describing the process of transferring a biochemical molecule to be detected from a bulk solution to an adsorption interface and binding with a probe molecule of the adsorption interface;
FIG. 2 is a schematic diagram of GHz acoustic fluid-promoted protein molecule adsorption;
FIG. 3-a is a cut-away view of a side-view microscopic image of an acoustic fluid in an impact mode; 3-b are simulated graphs of pressure distribution in impact mode;
FIG. 4-a is a cut-away view of a side-view microscopic image of an acoustic fluid in shear mode; FIG. 4-b is a graph showing a simulation of pressure distribution in shear mode;
FIG. 5 is a schematic diagram of an embodiment of a biomolecule sensing device;
FIG. 6 is a flow diagram of an embodiment of a method of facilitating biomolecule sensing;
FIG. 7 is a schematic diagram of an embodiment of an apparatus for facilitating biomolecule sensing;
FIG. 8 is a schematic diagram of the real-time adsorption curve of SAv molecules under the action of GHz acoustic fluid;
fig. 9 (a) is a schematic diagram of duty cycle optimization, and (b) is a schematic diagram of SAv adsorption under GHz acoustic fluid treatment with different duty cycles;
fig. 10 is a flow field microscopic image of GHz acoustic fluid at different relative distances of the fiber optic probe and resonator in the vertical direction.
Detailed Description
In the following description, reference is made to "some embodiments" which describe a subset of all possible embodiments, but it is understood that "some embodiments" may be the same subset or different subsets of all possible embodiments, and may be combined with each other without conflict.
In the following description, references to the terms "first \ second \ third, etc. or module a, module B, module C, etc. are used solely to distinguish between similar objects and do not denote a particular order or importance to the objects, but rather the specific order or sequence may be interchanged as appropriate to enable embodiments of the application described herein to be practiced in an order other than that shown or described herein.
In the following description, reference to reference numerals indicating steps, such as S110, S120 … …, etc., does not necessarily indicate that the steps are performed in this order, and the order of the preceding and following steps may be interchanged or performed simultaneously, where permissible.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. The terminology used herein is for the purpose of describing embodiments of the present application only and is not intended to be limiting of the application.
Before further detailed description of the embodiments of the present application, terms and expressions mentioned in the embodiments of the present application, and their corresponding uses, functions, and so on in the present application will be described, and the terms and expressions mentioned in the embodiments of the present application are used for the following explanation.
1. Resonators in the GHz range, or Film Bulk Acoustic Resonators (FBAR) or solid-state bulk acoustic resonators (SMR) operating at GHz, were originally manufactured using silicon backplanes, MEMS (micro-electro-mechanical systems) technology and thin-Film technology for high-frequency wireless communication applications. The FBAR filter has the characteristics of high frequency, low loss, low temperature drift, steep filter skirt edge, extremely high Q value, working frequency, sensitivity, resolution, bearable power capacity, small volume and compatibility of a preparation process and a CMOS (complementary metal oxide semiconductor), and occupies the application field of most wireless communication fields. The SMR resonator has the advantages of the FBAR described above, and also has the advantage of more stable and reliable performance when compared to the FBAR when the experiment is performed in liquid. In this application, GHz resonators are innovatively used in the field of driving fluids to facilitate biomolecular sensor sensing.
2. The impact mode, in this application, the mode in which the positive pressure generated by the GHz bulk acoustic wave generated by the resonator in the GHz range acts on the sensing interface of the sensor (see fig. 3-a, 3-b) is referred to as the impact mode, or jet mode.
3. The shear mode, in this application, the mode in which the negative pressure generated by the GHz bulk acoustic wave generated by the resonator of the GHz order acts on the sensing interface of the sensor (see fig. 4-a, 4-b) is referred to as the shear mode, or shear flow mode.
The technical solution in the present application will be described below with reference to the accompanying drawings.
First, the effect of molecular mass transfer (or molecular diffusion) on sensing performance is described. Fig. 1 shows a sensor detection process described by a two-chamber model, in which the flow rate of a fluid between an adsorption interface and a bulk solution is zero, and a biochemical molecule to be detected in the bulk solution moves to the adsorption interface in a free diffusion manner and then is combined with a probe molecule of the adsorption interface, as shown in fig. 1. It can be seen that the binding process of the biochemical molecule to be detected from the bulk solution to the adsorption interface is generally divided into the following three steps:
1) the concentration distribution of the biochemical molecules to be detected in the body solution tends to be uniform due to the mass transfer process caused by the uneven concentration distribution of the biochemical molecules to be detected in the body solution.
2) The biochemical molecules to be detected penetrate through the diffusion layer of the body solution and the adsorption interface and freely diffuse to the sensing interface.
3) The biochemical molecules to be detected are combined with the probe molecules at the adsorption interface.
In theory, formulas may be used
The two-chamber model is described. Wherein [ A ]]0Is the concentration of the biochemical molecule to be measured in the bulk solution, [ A ]]sIs the concentration, k, of the biochemical molecule to be measured at the adsorption interfacemIs the convective diffusion coefficient, konAnd koffRespectively binding constant and dissociation constant of the binding of the biochemical molecule to be detected to the probe molecule, [ B ]]Is the density of probe molecules on the adsorption interface, [ AB ]]Is the concentration of the biochemical molecule to be detected bound to the probe molecule. Because the flow velocity of the fluid in the diffusion layer is zero, the biochemical molecules to be detected mainly move to the adsorption interface in a diffusion mode, the process usually needs a long time, and meanwhile, the concentration of the biochemical molecules to be detected on the adsorption interface is far less than that of the biochemical molecules to be detected in the body solution. Transferring this to massThe limitations present in the process are called mass transfer limitations.
Currently, methods for enhancing molecular mass transfer mainly employ active methods, such as electrically driven convective motion or thermally conductive convective means, to enhance mass transport of analytes. These methods typically employ energy dissipation or energy differentials to create fluid perturbations, such as convective motion or fluid circulation motion, within the liquid to provide thorough mixing of the bulk solution, to eliminate depletion layers, and to accelerate molecular motion of the analyte. The following is introduced:
the first prior art is as follows: the detection performance of localized surface plasmon resonance is enhanced by an electrically driven fluidic approach. Under the action of electroosmosis fluid, bulk phase solution containing molecules to be detected is fully mixed, so that the transportation process of the biomolecules is promoted, the biomolecules are effectively adsorbed on a sensing interface of the sensor, the detection performance of the sensor is finally improved, and the detection limit is reduced.
The defects of the prior art are as follows: the enhanced sensing method based on electrically driven fluid has limitations in practical use. Since the strength of the electroosmotic fluid is inversely proportional to the salt concentration of the solution, the electroosmotic fluid phenomenon is weakened in high salinity solutions such as biological solutions, and thus the efficiency of enhancing the molecular adsorption is reduced, so that the method is not versatile.
The second prior art is: fluids based on thermal effects, such as the use of focused laser light to illuminate a micro-reaction chamber, thereby introducing a temperature gradient inside the reaction chamber, facilitating fluid circulation within the reaction chamber. The method can enhance the mass transport of biomolecules and promote the kinetics of molecular binding.
The second existing defect of the prior art: the method based on the thermal effect needs to consider the activity of the biological molecules in practical application, the biological molecules can be inactivated due to the increased temperature, and the efficiency is reduced due to the insufficient flow field speed caused by the low temperature. Therefore, a compromise is required in the choice of parameters and the benefit of enhanced adsorption.
Based on the defects in the prior art, the application provides a method and a device for promoting biomolecule sensing, and the basic principle is that a high-frequency bulk acoustic wave resonance device with the resonance frequency of GHz or above is manufactured through micro-size to generate high-efficiency controllable acoustic fluid in liquid. Due to the three-dimensional form of the GHz acoustic fluid and the characteristic of high efficiency and controllability of the fluid mode, the GHz acoustic fluid provides additional directional fluid driving force for the biomolecule adsorption process, particularly, when the driving is in an impact mode, the solid-liquid diffusion limitation in the sensing process is broken through, the replenishment of molecules under the body phase is realized, the molecular adsorption quantity can be enhanced finally, the detection sensitivity and the detection speed are improved, and the detection limit is reduced. In addition, the fluid dynamics method has strong compatibility with other sensing systems, so that the enhancement effect of the fluid dynamics method can be quantified and regulated. In addition, the application also realizes fixed-point modification and graphical modification of biomolecules on a sensing interface of the sensor and realizes the double functions of enhancing molecular adsorption and removing nonspecific adsorption by adjusting the relative positions of the GHz bulk acoustic wave resonator and the sensor. Compared with the prior art I and the prior art II, the method has the advantages of strong biocompatibility, simplified equipment structure, contribution to miniaturization, modularization and capability of being used as a multifunctional platform. The present application is described in detail below.
[ analysis of the principles of the present application ]
First, the principle of facilitating biomolecule sensing according to the present application will be explained. Referring to the schematic shown in fig. 2, a mathematical model of enhanced adsorption of proteins was analyzed as follows: in the protein molecule interaction process, diffusion-limited binding kinetics is a typical problem, as shown in the upper part of the left side of fig. 2, the adsorption process of biomolecules to the sensing interface of the sensor under the action of a fluid field depends on the free diffusion movement of the molecules, and the slow and random movement mode limits the detection performance of the sensor and is characterized in that a depletion layer gradually appears on the active surface of the sensor, so that the whole detection process is slow and the detected signal intensity is low. When the application introduces a GHz acoustic fluid, as shown in the lower left part of FIG. 2, sufficient mixing of target molecules in a solution is realized under the action of the GHz acoustic fluid, the mixing process effectively replenishes biomolecules in a depletion layer, the barrier effect of the depletion layer is broken, and therefore molecules near the active surface of the sensor maintain sufficient reaction concentration. On the other hand, as shown in the lower right part of fig. 2, under the action of GHz acoustic fluid impact mode, the target molecules near the sensing interface of the sensor are subjected to positive pressure, that is, the target molecules are driven directionally, so that the path time of the target molecules to the sensing interface of the sensor is shortened, and the absorption of the target molecules by the active surface of the sensor is further promoted. Therefore, under the synergistic effect of the enhanced mixing and directional driving of GHz acoustic fluid, the binding process of protein molecules is remarkably improved. The GHz acoustic fluid impact mode can also be used to facilitate sensing as shown in fig. 3-a and 3-b, where it can be seen that the acoustic fluid in the impact mode generates a positive pressure greater than 0 to the sensing interface of the sensor, and the pressure at the critical sensing interface can reach above 6 Pa.
Next, the principle of the present application relating to the removal of non-specific adsorbates (NSB) in shear mode based on acoustic fluids was analyzed: namely, the removal effect of the GHz acoustic fluid on nonspecific adsorbate in the shear mode is analyzed. According to the embodiments shown in fig. 4-a and 4-b, the shear mode of the GHz acoustic fluid is realized by adjusting the horizontal relative position between the sensing interfaces of the resonator and the sensor, when the sensing interfaces of the resonator and the sensor deviate from the vertical relative position, as shown in fig. 4-a and 4-b, a negative pressure will be generated at the sensing interfaces of the sensor, that is, the shear mode described in this application, which is favorable for removing the non-specific adsorbate, and at this time, the GHz acoustic fluid in the shear mode can generate a sufficient removal force at the contact surface of the sensing interface and the liquid, so that the non-specific adsorbate proteins in the sensing interface can be peeled off, and the target biomolecules adsorbed by the sensing interface are retained.
Examples of the present application that facilitate biomolecule sensing devices
The device shown in fig. 5 is an example of a device facilitating biomolecule sensing, and the device shown in fig. 7 is an embodiment of a device facilitating biomolecule sensing. With reference to these two figures, the following description will be given by way of example of a biomolecular sensing application in facilitating an optical sensor based on optical interferometry, the device comprising:
the acoustic fluid system part comprises a central processing unit 501, a signal generator 502, a power amplifier 503 and a resonator 504 which are electrically connected in sequence, wherein the central processing unit 501 controls the signal generator 502 to generate a GHz-level high-frequency electric pulse signal, the GHz-level high-frequency electric pulse signal is amplified by the power amplifier 503 and then applied to an excitation end of the resonator 504, the resonator 504 is driven to resonate, and GHz high-frequency sound waves are generated, so that GHz acoustic fluid is triggered. The central processing unit 501 may be an external computer or a control module built in the signal generator 502, and the resonator 504 may be a film bulk acoustic resonator, a solid assembled bulk acoustic resonator, or some other radio frequency device capable of generating GHz bulk acoustic waves. And the fluid speed and the fluid mode of the GHz acoustic fluid are regulated and controlled through a radio frequency signal generator so as to accurately regulate and control the promoting effect.
A cylindrical cavity 505 is provided at the top of the resonator to limit the extent of the acoustic fluid field. The cylinder may be a cylinder, a square cylinder or a cylinder with any shape, wherein the sidewall of the cavity 505 may be transparent for observation by an image acquisition device, such as a camera or a microscope. The observations include observations of relative position, distance, observation of acoustic fluid, etc., between the resonator 504 and the sensing interface of the sensor 508.
The sensing interface of the sensor 508 is directed towards the resonator and extends into the cavity 505. In this example, the sensor 508 is an optical sensor based on optical interferometry, and the sensing interface thereof is an optical probe of the optical sensor, and the surface of the optical probe is chemically/biologically modified to serve as an active sensing interface for acquiring quantitative data of adsorption of biomolecules in real time. The optical probe is partially immersed inside a cavity 505 above the resonator 504. The acoustic fluid system portion then acts as a removable bio-driver to enhance the sensing interface adsorption of the optical sensor.
A sample container 506 for storing a liquid sample to be analyzed, which may also be provided with a sample injection port and a discharge port. The resonator 504, the cavity 505 and the sensing interface of the sensing system are placed in the liquid to be analyzed in the sample container 506, so that the solution to be analyzed is contained in the cavity 505. The manner of providing the sample container 506 is effective when applied to dynamic measurement of a liquid sample to be analyzed (dynamic measurement indicates that dynamic information of biomolecule absorption accompanying time variation can be obtained), dynamic information of the liquid sample can be continuously measured, i.e., real-time monitoring is realized, and interference of liquid change is reduced.
Alternatively, as shown in fig. 7, the sample container 506 may not be provided, and the liquid sample to be analyzed may be directly injected into the cavity 505, for example, by a pipette. This method is effective for static measurement (static measurement indicates that the change in the amount of adsorbed molecules of the sensor before and after adsorption is detected, and dynamic information is ignored).
A displacement system 507 can also be arranged and comprises a precise three-dimensional displacement device and a horizontal objective table driven by the three-dimensional displacement device; in accordance with the embodiment of fig. 5, the sample container 506 is placed on the horizontal stage such that movement of the sample container 506 may be controlled by the displacement system 507 to indirectly control the relative position and distance of the resonator 504 to the sensing interface of the sensor 508. Alternatively, and in accordance with the embodiment of fig. 7, the cavity 505 is placed on the horizontal stage to directly control the relative position and distance of the resonator 504 to the sensing interface of the sensor 508 via the displacement system 507. Alternatively, the sensing interface is fixed on the horizontal stage, so that the movement of the sensing interface can be controlled by the displacement system, and the relative position and distance between the sensing interface of the sensing system and the resonator can be controlled. The above-mentioned control of the relative position and distance may realize the following functions:
by matching with the displacement system 507, fixed-point modification can be performed on the sensing interface: by adjusting the vertical distance between the two, the enhancement range of the GHz acoustic fluid can be accurately controlled, and fixed-point and quantitative modification on a sensor sensing interface is realized; and the displacement system is matched, and graphical biomolecular modification can be carried out on a sensor sensing interface.
The displacement system is matched, and the dual functions of enhancing molecular adsorption and removing nonspecific adsorption can be realized: when the resonator 504 and the sensing interface of the sensor 508 are opposite, the generated acoustic fluid acts on the sensing interface in an impact mode, which is beneficial to enhancing molecular adsorption; when the resonator 504 and the sensor 508 are off-perpendicular relative to each other, the acoustic fluid generated at this time acts on the sensing interface in a shear mode that facilitates removal of non-specific adsorbates. Therefore, by changing the horizontal relative position of the two, the double functions of GHz acoustic fluid enhanced molecular adsorption and non-specific adsorption removal can be realized.
In view of the above, with the above system, the efficient mixing action of the generated GHz acoustic fluid and the impact mode perpendicular to the sensing interface provide a directional driving force for the liquid to be analyzed above the resonator, facilitate its transport to the sensor sensing interface, enhance the sensor response, and shorten the response time.
In addition, in the embodiment shown in fig. 5, the bulk acoustic wave generated by the resonator is directed vertically upward, which is easily understood to match with the orientation of the sensing interface according to actual conditions. For example, the generated bulk acoustic waves may be directed downward, sideways, etc. in a matched manner, all for the purpose of being disposed toward the sensing interface of the sensing system.
Examples of methods for facilitating biomolecule sensing
Referring to the flow chart shown in fig. 6, to facilitate an example of a method for sensing biomolecules, an example of the method of the present application will be described with reference to the example of fig. 5 and the detailed description of fig. 7, and the method comprises the following steps:
s610: when the resonator is immersed in liquid, a GHz radio frequency signal amplified by the power amplifier is applied to the resonator, so that high-frequency sound waves are generated, the liquid is driven to upwards propagate in the liquid and is quickly attenuated, and strong volume force is generated to push the liquid to upwards move to form acoustic fluid which efficiently and stably exists above the resonator.
S620: when a sensing interface of a sensor is immersed in a flow field in a cavity above a resonator, an acoustic fluid directly contacts the sensing interface (such as a glass optical fiber) of the sensor, so that biomolecules in a flow field area near the sensing interface are subjected to upward pressure to promote the biomolecules to be transported to the sensing interface, and subsequent bioadsorption and identification processes are performed.
By changing the relative position between the sensing interface and the resonator, the topography of the flow field can be changed to create an impact mode (shown in fig. 3-a, 3-b) and a shear mode (shown in fig. 4-a, 4-b). The impact mode has an influence on the actual biomolecule detection, see the pressure distribution of the acoustic streaming field in this mode shown in fig. 3-a, 3-b, wherein a pressure below 0 corresponds to a pressure in the fluid field in the downward direction, and a pressure above 0 indicates an upward pressure direction. For the shock mode in the figure, the local pressure at the active surface of the sensing interface is a positive value, and the magnitude of the pressure decreases as the downward vertical distance of the sensing interface increases. It can be seen that the biomolecules in the flow field region near the sensing interface are subjected to an upward pressure, and in actual detection, such a stress mode will promote the transportation of the biomolecules to the sensitive surface of the sensing interface for the subsequent bio-adsorption and identification processes.
S630: when the biomolecule needs to be modified on the sensing interface of the sensor in a fixed-point and quantitative manner, the enhancement range of the GHz acoustic fluid is accurately controlled by adjusting the vertical distance between the resonator and the sensor, so that the fixed-point and quantitative modification on the sensing interface of the sensor is realized, and the detection of the sensor has the characteristics of fixed-point and quantitative determination.
On the other hand, when the graphical biomolecular modification needs to be performed on the sensing interface of the sensor, the biomolecular modification (i.e. graphical biomolecular modification) on the specific position of the sensing interface of the sensor is realized by adjusting the facing position and/or the distance between the resonator and the sensor by matching with a displacement system.
S640: the non-specific adsorbate on the sensor interface is further removed by using a shear mode, which is as follows:
the effect of the shear mode on the actual biomolecule detection can be seen from the pressure distribution of the acoustic flow field in the mode shown in fig. 4-a and fig. 4-b, at this time, the biomolecule on the sensing interface is subjected to downward pressure, which indicates that the biomolecule is more easily pulled and peeled off from the sensing interface by the liquid, and since the adsorption force of the nonspecific adsorbate on the sensing interface is smaller than the specific adsorption force of the target biomolecule to be detected, the downward pressure applied at the sensing interface can be controlled by controlling the GHz acoustic fluid power or controlling the distance between the sensing interface and the resonator, so as to be used for removing the nonspecific adsorbate (such as the nonspecific protein molecule). Therefore, through analyzing the liquid pressure caused by the acoustic fluid, the relative position of the GHz acoustic fluid and the sensing interface can be modulated, so that the interaction is adjusted, the operation and the removal of the non-specific adsorbate are realized, the adsorption of the non-specific adsorbate on the sensor interface is reduced, the sensing detection precision can be improved, and the sensing interface can have more spaces to adsorb the detected target biomolecule.
From the above, the present application achieves enhancement of molecular adsorption in acoustic fluid-based impact modes. Specifically, the GHz acoustic fluid provides additional directional fluid driving force for the biomolecule adsorption process by controlling the three-dimensional form and the fluid mode of the GHz acoustic fluid, so that the solid-liquid diffusion limitation in the sensing process is broken, the supplement of molecules under the body phase is realized, the molecular adsorption quantity can be enhanced finally, the detection sensitivity and the detection speed are improved, and the detection limit is reduced. And by adjusting the relative position of the GHz bulk acoustic wave resonator and the sensor, fixed-point modification and graphical modification of biomolecules on a sensor interface of the sensor are respectively realized, and the functions of enhancing molecular adsorption and removing non-specific adsorption are realized.
In addition, other types of resonators (e.g., different resonant frequencies, different sizes and different shapes) can also be used as drivers for the acoustic fluid; the liquid medium in which the acoustic fluid is present may be various buffers containing target molecules for biological detection, serum solutions, and other viscous solutions associated with enhanced adsorption or enhanced removal; the sensor used in combination with the GHz acoustic fluid can be other sensors based on affinity detection, such as an electrochemical sensor, an optical sensor, a mechanical biosensor (such as a micro-cantilever and a quartz crystal microbalance) and the like; the target substance to be detected can be protein molecules, DNA molecules, micro/nano particles, plasmids or the substances after chemical modification.
[ Experimental data and Effect of the present application ]
Preparing a GHz acoustic fluid promoted protein molecule adsorption experiment: referring to fig. 7, a schematic diagram of an embodiment is shown, including a signal generator, a power supply, a power amplifier; a fluorescence microscope; the optical sensor employs a biomolecular interaction analysis system (BLItz) developed by ForteBio, USA, which employs a label-free detection technique based on optical interferometry; and a three-dimensional displacement stage.
GHz acoustic fluid promotes optical detection mode: a transparent cavity was placed on top of the resonator to limit the extent of the acoustic fluid field, and in experiments, an amplified radio frequency signal was applied to the excitation end of the resonator to trigger the acoustic fluid, which was the part acting as the acoustic fluid system. In connection with other systems, such as the optical sensor based on optical interferometry used in this example, the resonator is placed directly below the optical sensor with the optical probe partially immersed inside the cavity above the resonator. The resonator is used as a detachable biological driver to enhance the sensing interface adsorption of the optical sensor.
And then verifying the adsorption effect of the GHz acoustic fluid enhanced protein molecules. The fiber probe (BLItz detection part) with the surface adsorbing the Biotin is immersed in SAv (Biotin can be combined with SAv) solution, standing treatment is adopted for a control group, standing treatment is initially adopted for an experimental group, GHz acoustic fluid treatment is adopted for the experimental group, and the combination condition of two groups of molecules is detected in real time. In the absence of a fluid field, as shown in fig. 8, the adsorption process of biomolecules to the sensing interface of the sensor depends on the free diffusion movement of the molecules, and the slow and random movement mode limits the detection performance of the sensor. For the experimental group, the binding kinetics were improved immediately after application of the GHz acoustic fluid, and the amount of SAv absorption was also increased substantially. This increase in binding rate and signal response strength is a result of the combined effect of the intensive mixing and directional steering induced by the GHz acoustic fluid.
Parameter optimization study of duty cycle: in this section, optimized sensing performance is achieved by adjusting various parameters of the GHz acoustic fluid. In the following parameter research, parameters of the pulse excitation signal, such as duty ratio, signal intensity and pulse period, are optimized, so that the GHz acoustic fluid can both enhance the molecular mass transfer effect and provide sufficient reaction time. For example, the duty cycle of the pulsed excitation signal is optimized as shown in fig. 9. The fiber optic probe (BLItz sensing part) with the surface modified with Biotin was immersed in SAv solution, the control group was treated with standing (i.e., 0%), the 5 experimental groups were treated with GHz acoustic fluid, the applied power (indicated by the letter "P") was kept constant, the pulse period (indicated by the letter "T") of the RF signal was kept constant, and the duty cycle (indicated by the letter "T1") of the signal was set to 10%, 20%, and 50% in this order. SAv adsorption data as shown in the following figure, the enhancement effect is best when the signal excitation with 50% duty ratio is used, so the 50% duty ratio is set as the first optimization parameter. Similar to this experiment, the power amplitude and duty cycle are optimized separately, so that an optimized result can be obtained.
Distance parameter study: in addition to the previously studied signal parameters, the effect of the relative distance between the sensing interface of the sensor and the resonator on molecular adsorption was also studied. As shown in fig. 10, by adjusting the vertical distance between the resonator and the sensor, the enhanced range of GHz acoustic fluids can be accurately manipulated. The vertical position is regulated and controlled, so that fixed-point and quantitative modification on a sensing interface of the sensor is realized; and the sensor interface of the sensor can be subjected to graphical biomolecular modification by matching with a displacement system.
According to the experimental data, the GHz acoustic fluid is used with other sensing systems according to the principle of enhancing the molecular adsorption and the adjusting mechanism of the GHz acoustic fluid, so that the solid-liquid diffusion limitation in the sensing process is broken, the molecular adsorption quantity is enhanced, the detection sensitivity and the detection speed of the sensing system are improved, and the detection limit is reduced. Carrying out fixed-point modification on a sensing interface of the sensor by adopting an acoustic fluid system; and realizing the graphical modification of the biological molecules by displacement control. The same acoustic fluid realizes the double functions of enhancing molecular adsorption and removing non-specific adsorption.
In the several embodiments provided in the present application, it should be understood that the disclosed system, apparatus and method may be implemented in other ways. For example, the above-described apparatus embodiments are merely illustrative, and for example, the division of the units is only one logical division, and other divisions may be realized in practice, for example, a plurality of units or components may be combined or integrated into another system, or some features may be omitted, or not executed. In addition, the shown or discussed mutual coupling or direct coupling or communication connection may be an indirect coupling or communication connection through some interfaces, devices or units, and may be in an electrical, mechanical or other form.
It is to be noted that the foregoing is only illustrative of the preferred embodiments of the present application and the technical principles employed. It will be understood by those skilled in the art that the present application is not limited to the particular embodiments described herein, but is capable of various obvious changes, rearrangements and substitutions as will now become apparent to those skilled in the art without departing from the scope of the application. Therefore, although the present application has been described in more detail with reference to the above embodiments, the present application is not limited to the above embodiments, and may include other equivalent embodiments without departing from the spirit of the present application.
Claims (10)
1. A method of facilitating biomolecule sensing, comprising:
driving the resonator to generate an acoustic fluid in the liquid in the order of gigahertz;
and (3) driving the biological molecules in the flow field to be transported to the sensing interface of the biosensor in the action area of the acoustic fluid through the acoustic fluid in the gigahertz level.
2. The method of claim 1, further comprising:
and driving the biological molecules to perform fixed-point, quantitative and/or graphical modification on the sensing interface of the sensor by controlling the relative position and/or distance between the sensing interface and the resonator.
3. A method according to claim 1 or 2, characterized in that the driving is performed in such a way that the resonator is located directly opposite the sensing interface.
4. The method of claim 3, further comprising: the ability of the drive is controlled by controlling the duty cycle, signal strength and/or pulse period parameters of the pulsed excitation signal driving the resonator.
5. The method of claim 1 or 2, further comprising:
and removing the non-specific adsorbate on the sensor interface by controlling the position between the sensor interface and the resonator to be in a non-opposite mode.
6. An apparatus for facilitating biomolecule sensing, comprising:
an acoustic fluid system section including a resonator for generating acoustic fluid in the gigahertz range;
a cylindrical cavity located within the acoustic fluid action region of the resonator for limiting the range of the acoustic fluid field and for accommodating the sensing interface of the biomolecular sensor.
7. The apparatus of claim 6, further comprising:
a three-dimensional displacement device carrying the resonator or fitted with the biomolecular sensor for controlling the relative position and/or distance between the resonator and the sensing interface.
8. The device of claim 6, wherein the chamber sidewall is transparent.
9. The apparatus of claim 6, wherein the acoustic fluid system portion comprises: the device comprises a central processing unit, a signal generator, a power amplifier and the resonator which are electrically connected in sequence.
10. A biomolecular sensing system comprising a biomolecular sensor and a device according to any of claims 6 to 9, the sensing interface of the biomolecular sensor being releasably extendable into the cavity of the device for facilitating sensing of the biomolecule.
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