Sensing device for analyzing fluids using sound waves

Abstract

A method for detecting an analyte in a fluid, the method comprising the step of pre-arranging a sensing device (100) comprising at least one SAW sensor (110), said or each SAW sensor (110) comprising a substrate (115) having an outer surface (115') comprising at least one piezoelectric portion, at least one transmitting interdigital transducer (111) disposed on said piezoelectric portion of said outer surface (115'), said transmitting interdigital transducer (111) being arranged to emit surface acoustic waves in response to an input electrical signal, at least one reflector electrode (112) disposed on said outer surface (115'), said reflector electrode (112) being arranged to reflect said acoustic waves back towards said transmitting interdigital transducer (111). Then the following steps: adsorbing a plurality of probe molecules on said outer surface (115') of said substrate (115) and / or on said or each transmitting interdigital transducer (111) and / or on said or each reflector electrode (112), sending n input electrical signals having respective frequencies f i to said transmitting interdigital transducer (111), where i = 1, and subsequently transmitting at least one surface acoustic wave by said or each transmitting interdigital transducer (111), reflecting said or each transmitted surface acoustic wave by said or each reflector electrode (112), identifying said n frequencies f i of said input electrical signals, corresponding to at least one resonance frequency f T of a surface acoustic wave generating a power exceeding a predetermined threshold P r , conveying said fluid on said outer surface (115') and / or on said or each transmitting interdigital transducer (111) and / or on said or each reflector electrode (112), removing said fluid through said outer surface (115') and / or through said or each transmitting interdigital transducer (111) and / or through said or each reflector electrode (112), verifying a possible change in the value of at least one previously identified resonance frequency f r .

Description

Technical Field

[0001] The present invention relates to the analysis of fluids and the detection of specific analytes of interest.

[0002] In particular, the present invention relates to the detection of these analytes by surface acoustic waves generated by nanostructured sensors.

[0003] Description of the prior art

[0004] The field of micro- and nanotechnology, so-called "microfluidics", i.e., technologies that allow the manipulation of biological and non-biological fluids on a micrometer scale, has become of great interest due to the rapid technological advances achieved in the last decades.

[0005] In fact, analysis in the microfluidic regime through chips equipped with suitable nanostructured sensors offers considerable advantages over standard equipment for laboratory analysis, since it allows the creation of fully automated systems of limited size and requiring minimal operator intervention, in addition to increasing the efficiency of the method due to the use of smaller amounts of reagents while maintaining high sensitivity. All this translates into significantly reduced costs, reduced complexity of the method itself and increased portability of the analytical tool.

[0006] An example of such a device is disclosed in "A Rayleigh Surface Acoustic Wave (R-SAW) Resonator Biosensor based on Positive and Negative Reflectors with Sub-Nanomolar Detection limit" by M. Agostini, G. Greco and M. Snipers, January 2018. The biosensor object of the article includes a piezoelectric substrate on which a metal structure capable of generating surface acoustic waves (SAW), in particular Rayleigh type (R-SAW), by applying radio frequency is produced.

[0007] His technology may be of particular interest in the field of timely diagnostics, that is, methods of analyzing patients with the goal of determining the presence of a pathology at an early stage. In fact, molecules whose concentrations are observed to change in the biological fluids of patients may be an indicator of damage or dysfunction of internal organs ("biomarkers") even before more serious symptoms are observed. For example, the possibility of identifying the onset of cancers such as breast cancer, pancreatic cancer or prostate cancer is known thanks to the monitoring of specific biomarkers. Changes in the concentrations of these biomarkers can usually only be detected by complex laboratory analyses, especially in the early stages of the disease. For this reason, in recent years, academic and industrial research has shown great interest in the development of easy-to-use, portable, timely diagnostic systems with high sensitivity, low cost and rapid and reliable measurements. Their use is not limited to the biomedical field. In fact, after appropriate identification of the analyte of interest, these devices can also be used for environmental monitoring and the detection of contaminants in food.

[0008] However, in the field of fluid analysis, in order to identify specific analytes, the instruments used mostly employ optical sensors, which are difficult to integrate on small chips, while the application of the above-mentioned acoustic biosensor surfaces is still a pioneering field of research and there are no examples of effective methods in this case.

[0009] US2006049714 describes a chemical acoustic wave sensor for monitoring the concentration of an analyte in a substance such as blood. The chemical sensor may include one or more interdigital transducers and a selective coating formed on a piezoelectric substrate. The coating and the interdigital transducer are used to convert an electrical signal into a surface wave. The device includes an antenna adapted to receive one or more input signals that excite the acoustic device and generate an output signal related to the concentration of the analyte of interest.

[0010] However, this system does not provide the possibility of introducing turbulence into the fluid to mix the fluid itself and increase the possibility of generating molecular bonds between the analyte and the molecular probe, thereby improving the efficiency of detecting the analyte itself. In fact, only SH-SAW type sound waves are used in US2006049714, which are not sufficient to generate the turbulence required for the above purposes. Summary of the invention

[0011] It is therefore a feature of the present invention to provide a method for detecting an analyte in a fluid that allows analysis to be performed with an accuracy and / or precision equal to or greater than known methods, utilizing a nanostructured device with a significantly reduced overall size and a distinct advantage in portability.

[0012] Another feature of the present invention is to provide a method that can reduce costs compared to traditional methods.

[0013] It is yet another feature of the present invention to provide such a method which allows for improved reproducibility of the analysis.

[0014] These and other objects are achieved by a method for detecting an analyte in a fluid, the method comprising the steps of:

[0015] - a pre-arranged sensing device comprising at least one SAW sensor, the or each SAW sensor comprising:

[0016] - a substrate having an outer surface, said outer surface comprising at least one piezoelectric portion;

[0017] - at least one transmitting interdigital transducer arranged on a piezoelectric portion of said outer surface, said transmitting interdigital transducer being arranged to transmit surface acoustic waves in response to an input electrical signal;

[0018] - at least one reflector electrode arranged on said outer surface, said reflector electrode being arranged to reflect acoustic waves towards said transmitting interdigital transducer;

[0019] - adsorbing a plurality of probe molecules on said outer surface of the substrate and / or on said or each emitting interdigital transducer and / or on said or each electrode reflector;

[0020] - will have respective frequencies f i n input electrical signals are sent to a transmitting interdigital transducer, where i=1, ..., n, and at least one surface acoustic wave is subsequently transmitted by the or each transmitting interdigital transducer;

[0021] - reflecting the or each emitted surface acoustic wave by the or each reflector electrode;

[0022] - Identify the n frequencies f of the input electrical signal i , corresponding to the power generated exceeding the predetermined threshold P T At least one resonant frequency f of the surface acoustic wave r ;

[0023] - conveying a fluid on the outer surface and / or on the or each transmitting interdigital transducer and / or on the or each electrode reflector;

[0024] - removing fluid through the outer surface and / or through the or each transmitting interdigital transducer and / or through the or each electrode reflector;

[0025] - verifying a possible change in the value of at least one resonance frequency fr previously identified.

[0026] In particular, the identified resonant frequency or one of the identified resonant frequencies may be selected and it may be assessed whether, after the fluid has passed, an electrical signal at this frequency still generates an acoustic wave of power equal to the previously detected power.

[0027] Alternatively, new multiple electrical signals may be emitted and a new resonant frequency (or multiple new frequencies) detected and compared to the frequencies identified prior to the passage of fluid.

[0028] In both cases, if the parameters that influence the position of the resonance frequency in an undesired way, such as temperature and pressure, do not change and if the reference resonance frequency does not change, it means that it has not changed the mass density of the outer surface of the substrate or the electrodes of the transducer and reflector, and therefore this means that the analyte being analyzed, i.e. the molecule capable of forming a bond with the molecular probe adsorbed on the sensor, is not present in the fluid being analyzed.

[0029] Rather, the change in resonant frequency is a function of the change in mass density at the external surface and therefore a function of the concentration of the analyte present in the fluid being analyzed.

[0030] The change in mass density may be positive in the case where the analyte binds to the molecular probe, or negative in the case where the analyte cleaves the molecular probe.

[0031] Compared to devices utilizing optical sensors coupled to transducers, this solution is completely electrical and integrated on a chip, allowing for a significantly smaller footprint and increased practicality, making the device an easily transportable fluid analysis laboratory.

[0032] Advantageously, the step of adsorbing a plurality of probe molecules is performed on the or each emitting interdigital transducer and / or on the or each electrode reflector.

[0033] In particular, the sensing device further comprises an auxiliary interdigital transducer and, simultaneously with or upstream of the step of delivering the fluid on the outer surface, a step of transmitting a signal having a frequency f through the auxiliary interdigital transducer is provided. t The step of surface acoustic waves is used to introduce turbulence in a fluid.

[0034] In this way, the probability of creating a molecular bond between the analyte and the molecular probe increases, thereby improving the detection efficiency of the analyte itself.

[0035] Advantageously, the frequency f of the n surface acoustic waves emitted by the transmitting interdigital transducer is i Greater than 800MHz, while the frequency f emitted by the auxiliary interdigital transducer t Less than 800MHz.

[0036] Advantageously, the sensing device comprises at least two SAW sensors and a control unit and further provides for comparing the surface acoustic waves reflected by the reflector electrodes of the at least two SAW sensors to reduce noise and determine the or each resonant frequency f with greater accuracy. r steps.

[0037] Compared to a setup using a single SAW sensor, this solution can improve the accuracy of the analysis since the differential measurement acts as a filter for the response signal.

[0038] In particular, the sensing device includes at least one microfluidic channel arranged to transport a fluid along a predetermined path on an outer surface so as to contain the fluid in a minimum volume for analysis.

[0039] This solution allows the use of reduced fluid volumes, reduced consumption, and increased surface / volume ratio compared to prior art devices, thereby improving diagnostic performance, thus making it possible to significantly increase the number of analyses performed on the same device.

[0040] Advantageously, there are at least two resonance frequencies f defined r steps.

[0041] This solution allows more than one signal to be received simultaneously, enabling a comparison to be made reducing the noise of the signals themselves.

[0042] According to another aspect of the present invention, a sensing device for detecting an analyte in a fluid is claimed, the sensing device comprising at least one SAW sensor, the or each SAW sensor comprising:

[0043] - a substrate having an outer surface, said outer surface comprising at least one piezoelectric portion;

[0044] - a transmitting interdigital transducer arranged on the outer surface, the transmitting interdigital transducer being arranged to transmit surface acoustic waves in response to an input electrical signal;

[0045] - at least one reflector electrode arranged on said outer surface, said reflector electrode being arranged to reflect acoustic waves towards said transmitting interdigital transducer.

[0046] The present invention therefore proposes a device that can be fully integrated within a chip, capable of performing high sensitivity analysis relative to traditional instruments, thus allowing the implementation of the above-mentioned method.

[0047] Advantageously, there is also provided an auxiliary interdigital transducer arranged to transmit a signal having a frequency f t The surface acoustic waves are used to introduce turbulence in the fluid, and the frequency f of the n surface acoustic waves emitted by the transmitting interdigital transducer is iGreater than 800MHz, while the frequency f emitted by the auxiliary interdigital transducer t Less than 800MHz.

[0048] In particular, at least two SAW sensors and a control unit are also provided, which are arranged to compare surface acoustic waves reflected by reflector electrodes of the at least two SAW sensors to reduce noise.

[0049] In particular, at least one microfluidic channel is provided, which is arranged to transport a fluid along a predetermined path on the outer surface so as to contain the fluid in a minimum volume for performing the analysis.

[0050] In particular, the or each SAW sensor is an R-SAW sensor adapted to generate surface acoustic waves of the Rayleigh type [Rayleigh Surface Acoustic Waves].

[0051] Advantageously, the piezoelectric portion of the substrate is made of lithium niobate [LN].

[0052] This material allows the transmission of R-SAW surface acoustic waves in at least one direction.

[0053] In particular, the substrate has been cut "Y-cut 128° X-rotated".

[0054] In this way, the substrate allows the transmission of R-SAW surface acoustic waves in at least two directions. BRIEF DESCRIPTION OF THE DRAWINGS

[0055] Other features and / or advantages of the present invention will become more apparent from the following description of exemplary embodiments thereof, which are illustrative and non-limiting, with reference to the accompanying drawings, in which:

[0056] - Figure 1 shows a flow chart of a method for detecting an analyte in a fluid according to the present invention;

[0057] - Figure 2A A possible exemplary embodiment of a sensing device for detecting an analyte in a fluid according to the present invention is shown;

[0058] - Figure 2B Another possible exemplary embodiment of a sensing device is shown;

[0059] - Figure 3A , 3B , 3C, 3D, 3E and 3F show some possible arrangements of interdigital transducers and reflector electrodes on the outer surface of the substrate. DETAILED DESCRIPTION

[0060] refer to Figure 1Flow chart 300, a method for detecting an analyte in a fluid according to the present invention comprises pre-arranging a possible exemplary embodiment thereof as shown in Figure 2A The first step of the sensing device 100

[301] .

[0061] In particular, Figure 2A The sensor device 100 includes four SAW sensors 110, each of which includes a substrate 115 having an outer surface 115', the outer surface having at least one piezoelectric portion. Figure 3A , 3B , 3C, 3D, 3E and 3F, each SAW sensor 110 includes at least one transmitting interdigital transducer 111 and at least one reflector electrode 112 arranged on the piezoelectric portion of the outer surface 115'. In particular, the transmitting interdigital transducer 111 is suitable for transmitting surface acoustic waves in response to an input electrical signal, and the reflector electrode 112 is suitable for reflecting the acoustic waves toward the transmitting interdigital transducer 111.

[0062] Furthermore, the sensor device 100 may comprise at least one auxiliary interdigital transducer 120, which is arranged to emit a signal having a frequency f t Surface acoustic waves are used to induce turbulence in fluids.

[0063] Still reference Figure 1 300 , the method then provides a step of adsorbing a plurality of probe molecules on the outer surface 115 ′ of the substrate 115 and / or on the electrodes of the transmitting interdigital transducer 111 and the electrodes of the reflector electrode 112 [ 302 ].

[0064] Then the respective frequencies f i The step of sending n input electrical signals, where i=1, ..., n, to the transmitting interdigital transducer 111, and then transmitting at least one surface acoustic wave

[303] .

[0065] This is followed by a step of reflecting the or each emitted surface acoustic wave by the or each reflector electrode 112

[304] .

[0066] Based on the reflected signal and / or the transmitted signal, the n frequencies f of the input electrical signal can be identified i , corresponding to the power generated exceeding the predetermined threshold P T At least one resonant frequency f of the surface acoustic wave r

[305] This identification can be done graphically by identifying peaks in the spectrum of the reflected signal that are above a certain value, i.e., peaks that actually produce sound waves and therefore peaks where the energy of the reflected signal is low or zero.

[0067] There then exists a step of delivering a fluid at the probe molecules, ie on the outer surface 115 ′ of the substrate 115 and / or on the electrodes of the transmitting interdigital transducer 111 and the reflector electrode 112 [ 306 ].

[0068] refer to Figure 2B In a preferred exemplary embodiment, the device 100 includes at least one microfluidic channel 130 arranged to provide a transport step by directing the fluid along a predetermined path on the outer surface 115' so as to contain the fluid in a minimum volume for analysis. The microfluidic channel 130 may also include an analysis chamber 130' at each SAW sensor 110 for allowing the fluid to reside and / or move on the electrodes of such SAW sensor 110.

[0069] In a preferred exemplary embodiment, the device 100 comprises at least one auxiliary interdigital transducer 120 arranged to emit a signal having a frequency f t Surface acoustic waves are used to induce turbulence in fluids.

[0070] This is followed by the step of removing fluid from the area to which it was transported

[307] .

[0071] Then it is necessary to verify at least one resonant frequency f previously identified. r Steps for possible changes in the value of

[308] .

[0072] The foregoing description of some exemplary embodiments will fully disclose the present invention from a conceptual point of view so that others, by applying current knowledge, will be able to modify and / or adapt specific exemplary embodiments in various applications without further study and without departing from the present invention, and therefore, it means that such adaptations and modifications must be considered equivalent to the specific embodiments. The components and materials that implement the different functions described herein can have different properties without departing from the field of the invention for this reason. It should be understood that the wording or terminology adopted herein is for the purpose of description rather than limitation.

Claims

1. A method for detecting an analyte in a fluid, the method comprising the following steps: - a pre-arranged sensor device (100) comprising at least one SAW sensor (110), the or each SAW sensor (110) comprising: - a substrate (115) having an outer surface (115') comprising at least one piezoelectric portion; - at least one transmitting interdigital transducer (111) arranged on said piezoelectric portion of said outer surface (115'), said transmitting interdigital transducer (111) being arranged to transmit surface acoustic waves in response to an input electrical signal; - at least one reflector electrode (112) arranged on the outer surface (115'), the reflector electrode (112) being arranged to reflect the surface acoustic waves towards the transmitting interdigital transducer (111); - an auxiliary interdigital transducer (120) arranged on said piezoelectric portion of said outer surface (115'); - adsorbing a plurality of probe molecules on the or each emitting interdigital transducer (111) and / or on the or each reflector electrode (112); - will have respective frequencies f i sending n input electrical signals to the transmitting interdigital transducer (111), where i=1, ..., n, and subsequently transmitting at least one surface acoustic wave by the or each transmitting interdigital transducer (111); - reflecting the or each emitted surface acoustic wave by the or each reflector electrode (112); - identifying the n frequencies f of the input electrical signal i , corresponding to the power generated exceeding the predetermined threshold P T At least one resonant frequency f of the surface acoustic wave r ; - conveying the fluid on the outer surface (115') and / or on the or each transmitting interdigital transducer (111) and / or on the or each reflector electrode (112); - the auxiliary interdigital transducer (120) transmits a signal having a frequency f t surface acoustic waves for introducing turbulence in the fluid; - removing said fluid through said outer surface (115') and / or from said or each transmitting interdigital transducer (111) and / or from said or each reflector electrode (112); - Verify at least one previously identified resonant frequency f r possible changes in the value of ; The method is characterized in that the or each SAW sensor (110) is an R-SAW sensor adapted to generate Rayleigh surface acoustic waves, And in that the sensing device (100) includes at least two SAW sensors (110) and a control unit and wherein a step of reducing noise by comparing the surface acoustic waves reflected by the reflector electrodes (112) of the at least two SAW sensors (110) is also provided, the surface acoustic waves reflected by the reflector electrode (112) or each reflector electrode (112) having the same wavelength.

2. The method for detecting an analyte in a fluid according to claim 1, wherein the frequency f of the n surface acoustic waves emitted by the transmitting interdigital transducer (111) is i greater than 800 MHz, and the frequency f emitted by the auxiliary interdigital transducer (120) t Less than 800MHz.

3. A method for detecting an analyte in a fluid according to claim 1, wherein the sensing device (100) comprises at least one microfluidic channel (130), which is arranged to transport the fluid along a predetermined path on the outer surface (115') so as to contain the fluid in a minimum volume for performing the analysis.

4. The method for detecting an analyte in a fluid according to claim 1, wherein the piezoelectric portion of the substrate (115) is made of lithium niobate [LN].

5. The method for detecting an analyte in a fluid according to claim 1, wherein there are at least two resonant frequencies f r steps.

6. A sensing device (100) for detecting an analyte in a fluid, the sensing device (100) comprising at least one SAW sensor (110), the or each SAW sensor (110) comprising: - a substrate (115) having an outer surface (115') comprising at least one piezoelectric portion; - a transmitting interdigital transducer (111) arranged on the outer surface (115'), the transmitting interdigital transducer (111) being arranged to transmit surface acoustic waves in response to an input electrical signal; - at least one reflector electrode (112) arranged on the outer surface (115'), the reflector electrode (112) being arranged to reflect the surface acoustic waves towards the transmitting interdigital transducer (111); - an auxiliary interdigital transducer (120) arranged to transmit a signal having a frequency f t surface acoustic waves for introducing turbulence in the fluid; The sensor device (100) is characterized in that The or each SAW sensor (110) is an R-SAW sensor adapted to generate Rayleigh surface acoustic waves; And at least two SAW sensors (110) and a control unit are provided, the control unit being arranged to reduce noise by comparing the surface acoustic waves reflected by the reflector electrodes (112) of the at least two SAW sensors (110), the surface acoustic waves reflected by the reflector electrode (112) or each reflector electrode (112) having the same wavelength.

7. The sensing device (100) according to claim 6, wherein the frequency f of the n surface acoustic waves emitted by the transmitting interdigital transducer (111) is i greater than 800 MHz, and the frequency f emitted by the auxiliary interdigital transducer (120) t Less than 800MHz.

8. The sensing device (100) according to claim 6, wherein at least one microfluidic channel (130) is provided, which is arranged to transport the fluid along a predetermined path on the outer surface (115') so as to contain the fluid in a minimum volume for performing the analysis.

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