CN112698085A - Weak current detection device based on single-molecule electrochemical analysis and analysis method thereof - Google Patents
Weak current detection device based on single-molecule electrochemical analysis and analysis method thereof Download PDFInfo
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- 238000001514 detection method Methods 0.000 title claims abstract description 35
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- 238000005070 sampling Methods 0.000 claims abstract description 21
- 238000006243 chemical reaction Methods 0.000 claims abstract description 20
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- 238000000835 electrochemical detection Methods 0.000 claims abstract description 13
- 230000003321 amplification Effects 0.000 claims abstract description 11
- 238000003199 nucleic acid amplification method Methods 0.000 claims abstract description 11
- 238000000034 method Methods 0.000 claims abstract description 9
- 239000002090 nanochannel Substances 0.000 claims description 12
- 238000001914 filtration Methods 0.000 claims description 8
- 238000010183 spectrum analysis Methods 0.000 claims description 4
- 238000000970 chrono-amperometry Methods 0.000 claims description 3
- 230000007613 environmental effect Effects 0.000 claims description 3
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- 108020004414 DNA Proteins 0.000 description 21
- 230000005540 biological transmission Effects 0.000 description 8
- 102000053602 DNA Human genes 0.000 description 7
- 239000010453 quartz Substances 0.000 description 5
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N silicon dioxide Inorganic materials O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 5
- 230000008859 change Effects 0.000 description 4
- 230000006399 behavior Effects 0.000 description 3
- 230000001186 cumulative effect Effects 0.000 description 3
- 150000002500 ions Chemical class 0.000 description 3
- 229910021607 Silver chloride Inorganic materials 0.000 description 2
- 230000000903 blocking effect Effects 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 238000002848 electrochemical method Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- HKZLPVFGJNLROG-UHFFFAOYSA-M silver monochloride Chemical compound [Cl-].[Ag+] HKZLPVFGJNLROG-UHFFFAOYSA-M 0.000 description 2
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Abstract
The invention provides a high-speed sampling detection device and an analysis method which can be directly used for single-molecule electrochemical detection, in particular to a device and a method which can amplify, filter, store, display and process and analyze weak current signals of single-molecule electrochemical detection. The device comprises a data interface and conversion device, a current amplifier, a control and data analysis system and a shielding box, wherein the data interface and conversion device, the current amplifier, the control and data analysis system are arranged in the shielding box; the single-molecule electrochemical detection is carried out in the shielding box, the shielding device is provided with an output interface, and the converted current signals can be displayed on a computer interface through the output interface for output. The device has the outstanding advantages of weak current amplification, high-speed sampling, signal analysis and processing, and can perform relevant electrochemical operations on a detection sample, such as linear scanning and the like.
Description
Technical Field
The invention belongs to the technical field of electrochemical measurement, and particularly relates to a single-molecule electrochemical ultralow current detection device and a data processing and analyzing method.
Background
In the aspects of physical, chemical and biological research, the extraction and detection of weak current signals always restrict the rapid development of a single-molecule behavior measurement method. Because of weak currentSignals are easily disturbed by the external environment, and with the addition of their low values, the weak current signals of the relevant single-molecule studies are usually buried in the background signal if the relevant signal is not processed. In addition, the time scale of behavior of a single molecule is very small (on the order of microseconds) from a physical and chemical standpoint. Therefore, for weak current signals (typically less than 10)-11A) High-frequency detection (sampling frequency is 10-500KHz) and analysis are the key points for realizing the observation of individual behaviors of single molecules by an electrochemical method. However, current electrochemical analysis uses electrochemical stations capable of detecting only currents greater than 10-9A, and high-speed sampling cannot be achieved. In addition, some existing weak signal detection systems, such as patch clamp, the device can only be applied to monitoring of ion channel current on the surface of cell membrane, and in the aspect of biological nanopore detection, the device cannot be used in detection of single molecule electrochemical reaction due to the device structure, and the data analysis of the device is complicated and needs to be carried out by using separate software, and the data generated by experiment cannot be obtained by what you see; in summary, there is no high-speed sampling detection device and analysis method for single-molecule electrochemical analysis.
Disclosure of Invention
In view of the above problems, the present invention is directed to a high-speed sampling detection device and analysis method that can be directly used for single-molecule electrochemical detection, and more particularly, to a device and method that can amplify, filter, store, display, process and analyze weak current signals of single-molecule electrochemical detection. The device has the outstanding advantages of weak current amplification, high-speed sampling, signal analysis and processing, and can perform relevant electrochemical operations on a detection sample, such as linear scanning and the like.
In order to achieve the purpose, the invention comprises a current detection port, a current amplifier, an analog-to-digital conversion and data processing module, a control and data analysis system, a shielding box for single-molecule electrochemical detection and the like (figure 1).
Specifically, the device comprises a data interface and conversion device, a current amplifier, a control and data analysis system and a shielding box, wherein the data interface and conversion device, the current amplifier and the control and data analysis system are arranged in the shielding box;
the data interface and conversion device is used for inputting a first current signal into the current amplifier, wherein the first current signal is a received weak current signal;
the current amplifier is used for amplifying the first current signal, converting the first current signal into a voltage signal and transmitting the voltage signal to the control and data analysis system;
the control and data analysis system comprises an analog-to-digital conversion and data processing module, and the analog-to-digital conversion and data processing module is used for reducing the voltage signal into a second current signal and filtering to eliminate low-frequency related noise so as to obtain single-molecule electrochemical data.
As a preferable technical solution of the present invention, the shielding device is provided with an output interface, and the current signal after the analysis of the monomolecular electrochemical data is converted can be displayed through the output interface.
As a preferable technical solution of the present invention, the shielding box is an independent device for shielding environmental electromagnetic noise during electrochemical detection.
As a preferable technical solution of the present invention, the data interface and the conversion device are bayonet nut connector BNC interfaces, and are disposed on the shielding box.
As a preferred embodiment of the present invention, the control and data analysis system instantly displays the data of the second current signal, and simultaneously performs fourier transform on the second current signal to obtain noise spectrum analysis data of the current signal.
In a preferred embodiment of the present invention, the detection device measures the first current signal by linear scanning and/or chronoamperometry.
As a preferred technical solution of the present invention, the stored single molecule electrochemical data is subjected to corresponding data processing and analysis by writing a Python processing program independently.
As a preferred embodiment of the present invention, the control and data analysis system instantly displays the data of the second current signal, and simultaneously performs fourier transform on the second current signal to obtain noise spectrum analysis data of the current signal.
In a preferred embodiment of the present invention, the detection device measures the first current signal by linear scanning and/or chronoamperometry.
As a preferable technical scheme of the invention, the shielding box for the single-molecule electrochemical detection is an independent device and is mainly used for shielding environmental electromagnetic noise during the electrochemical detection. A certain electric potential is applied to a nano channel or a nano electrode through a current detection port in a shielding box, a high-frequency sampling signal of weak current detected by monomolecular electrochemistry is obtained through a control and data analysis system and then enters a current amplifier through a BNC Bayonet Nut Connector (BNC for short), the current amplifier amplifies the weak current signal and converts the weak current signal into a voltage signal, the voltage signal is transmitted to the control and data analysis system, and an analog-to-digital conversion and data processing module (ADC) reduces the voltage signal into a second current signal according to the current amplification proportion given by the system and carries out filtering processing to eliminate low-frequency related noise. The control and data analysis system displays the current signal data in real time, and simultaneously performs Fourier transform on the current signal to obtain noise spectrum analysis data of the current signal (see figure 2). And (4) carrying out corresponding data processing and analysis on the stored single-molecule electrochemical data by a corresponding data processing program (self-writing Python program).
As a preferred technical solution of the present invention, the present invention further provides an analysis method of a weak current detection device based on single-molecule electrochemical analysis, the method specifically includes:
applying a specified potential to a nano channel or a nano electrode through a current detection port in a shielding box, and receiving a first current signal through a control and data analysis system, wherein the first current signal is a high-frequency sampled weak current signal obtained by single-molecule electrochemical detection;
a current amplifier is intervened through a BNC interface arranged on the shielding box, the current amplifier amplifies and converts the first current signal into a voltage signal, and the voltage signal is transmitted to the control and data analysis system;
and reducing the voltage signal into a second current signal through an analog-to-digital conversion and data processing module according to a current amplification ratio given by the system, filtering to eliminate low-frequency related noise to obtain single-molecule electrochemical data, analyzing the single-molecule electrochemical data, and outputting the single-molecule electrochemical data through an output interface, wherein the analog-to-digital conversion and data processing module is arranged in the control and data analysis system. As a preferable technical scheme of the invention, the sampling frequency of the high-frequency sampling signal is in the regulation range of 1-500 KHz.
As a preferable technical scheme of the invention, the amplification ratio of the current is 103-108V/A。
Has the advantages that:
1. the invention can change the amplification ratio of weak current according to the requirement of detection bandwidth, and the amplification ratio is 103-108V/A; current detection limit 10X10-15A
2. The invention can change the sampling frequency according to the experiment requirement, the adjusting range of the sampling frequency is 1-500KHz, and the requirements of single molecule electrochemical detection are fully met;
3. the control and data processing system of the invention realizes the anti-interference digital filtering function through the filtering processing function set by software, reduces the high-frequency and power frequency interference, can analyze and process the obtained data, and realizes the WYSIWYG (what you see is what you get) of the data.
Drawings
FIG. 1 is a schematic diagram of a low current high speed sampling and detecting device and a control system
FIG. 2 noise spectral data analysis
FIG. 3.54nm Quartz capillary tube Ionic Current at different voltages (KCl concentration in capillary tube 0.1mM)
FIG. 4.54nm nano-capillary linear scan data analysis results
FIG. 5. the current signal changes during the transport of lambda-DNA in a 38nm diameter nanocapillary, each current signal drop representing a lambda-DNA molecule passing through the nanocapillary.
FIG. 6 statistical analysis of data transmitted by lambda DNA in a 38nm diameter quartz capillary, each point in the plot representing a lambda DNA molecule, the abscissa representing the residence time of the DNA molecule in the nanocapillary and the ordinate representing the magnitude of the change in conductance of the nanocapillary caused by the transmission of the DNA molecule in the nanocapillary.
FIG. 7.500KHz current sampling signal diagram
Detailed Description
The following merely illustrates the principles of the invention. Therefore, although not explicitly described or shown in the present specification, those skilled in the art can implement the principle of the invention and invent various devices included in the concept and scope of the invention. Further, it is to be understood that all terms and embodiments of the appended claims are principally intended expressly to be only for understanding the concept of the invention, and are not to be construed as limiting the embodiments and aspects specifically enumerated herein.
The first embodiment is as follows: nanocapillary ion current detection
In order to measure weak current signals of the small-sized quartz nano-capillary, the concentration of electrolyte filled in the small-sized quartz nano-capillary with the diameter of 54nm is 0.1 mM. The voltage signal amplified by a current amplifier is acquired by a PCIe6353 data acquisition card (DAQ card) of NI (national instruments), the electrode adopts a two-electrode system (Ag/AgCl electrode), the voltage range is set to be 0.5V to-0.5V through a control system, the voltage change amplitude is 10mV/s, the current-voltage curve of the nano capillary needs to be measured is considered, the measurement of single molecule information is not involved, therefore, the sampling frequency of the current signal is set to be 1KHz, and the current amplification ratio is set to be 107V/A, reducing the finally amplified voltage signal into a current signal through a control and analysis system, storing and displaying the current signal, and processing the current signal through a Python program to obtain the current signal under different voltages acquired as shown in figure 3, wherein the result shows that the detection device can detect weak electricity in the nano channelThe flow is detected. Referring to fig. 4, the result of signal analysis shows that the apparatus and the analysis method can detect and analyze pA-level weak current.
Example two: detection of DNA transport in nanocapillary
In order to detect the transmission process of single DNA molecules in a quartz nanocapillary with the diameter of 38nm and the voltage of 0.5V, lambda-DNA is selected as a detection molecule, because the time for the single DNA molecules to pass through the nanochannel is very short (mu s), the sampling frequency of the device must be increased in order to record that the single DNA molecules pass through the nanochannel, the sampling frequency of 100KHz is selected in the test, when the detection bandwidth (the sampling frequency of signals) of the device is increased, the output amplitude of a current amplifier is reduced, and in order to control the amplitude reduction, the amplification ratio of the current amplifier is adjusted to 104V/A, collecting the amplified signals by using a PCIe6353 data acquisition card (DAQ card) of NI (national instruments) company, and performing low-pass filtering on the signals by using a two-electrode system (Ag/AgCl electrode) for 10 KHz. The measured DNA via current signals are shown in FIG. 5, and the scatter plot between the detected DNA residence time and the current response signals is shown in FIG. 6, which indicates that the apparatus and the analysis method are fully applicable to single-molecule electrochemical analysis. FIG. 7 is a graph of a 500KHz current sampling signal.
EXAMPLE III Single molecule Transmission Signal analysis
Since the signal generated by the DNA molecule during nanopore transmission is a time series signal, when a single DNA molecule passes through the nanochannel, the DNA molecule blocks the transmission of ions in the nanochannel, which in turn causes a decrease in ion current in the nanochannel. In addition, the transmission speed of the DNA molecules in the nanochannel is quite high, so that a time series signal with a large data volume is generated finally, and how to analyze and acquire the information transmitted by the DNA molecules in the signal becomes a problem to be solved in single molecule analysis. The invention integrates a processing program of time sequence analysis signals in a device, which adopts a cumulative sum algorithm (CUSUM), and judges whether DNA molecules are transmitted or not, the magnitude of the blocking current in the transmission process and the retention time of DNA in a nano channel by analyzing the difference between the cumulative sum value of the signals transmitted by the DNA molecules and the cumulative sum value of the signals transmitted by the DNA molecules through a computer program. In the algorithm, when the standard deviation of the baseline current signal in the nano-channel is calculated and the standard deviation of the blocking current in the DNA transmission process is set to be more than 5 times of the standard deviation of the baseline current signal, the DNA molecule can be considered to pass through the nano-channel.
As described above, although the present invention has been described with reference to the preferred embodiments, those skilled in the art can implement the present invention with various modifications and variations without departing from the spirit and scope of the present invention as set forth in the appended claims.
Claims (10)
1. The weak current detection device based on single-molecule electrochemical analysis is characterized by comprising a data interface and conversion device, a current amplifier, a control and data analysis system and a shielding box, wherein the data interface and conversion device, the current amplifier and the control and data analysis system are arranged in the shielding box;
the data interface and conversion device is used for inputting a first current signal into the current amplifier, wherein the first current signal is a received weak current signal;
the current amplifier is used for amplifying the first current signal, converting the first current signal into a voltage signal and transmitting the voltage signal to the control and data analysis system;
the control and data analysis system comprises an analog-to-digital conversion and data processing module, and the analog-to-digital conversion and data processing module is used for reducing the voltage signal into a second current signal and filtering to eliminate low-frequency related noise so as to obtain single-molecule electrochemical data.
2. The weak current detection device based on single-molecule electrochemical analysis according to claim 1, wherein the shielding device is provided with an output interface, and after the single-molecule electrochemical data is analyzed, the current signal can be displayed through the output interface after being converted.
3. The weak current detection device based on single molecule electrochemical analysis according to claim 1, wherein the shielding box is a stand-alone device for shielding environmental electromagnetic noise during electrochemical detection.
4. The weak current detection device based on single-molecule electrochemical analysis according to claim 1, wherein the data interface and conversion device is a Bayonet Nut Connector (BNC) interface disposed on the shielding box.
5. The weak current detection device based on single-molecule electrochemical analysis according to claim 1, wherein the control and data analysis system displays the data of the second current signal in real time, and simultaneously performs fourier transform on the second current signal to obtain the noise spectrum analysis data of the current signal.
6. The weak current detection device based on single molecule electrochemical analysis according to claim 4, wherein the first current signal is measured by linear scanning and/or chronoamperometry.
7. The weak current detection device based on single-molecule electrochemical analysis according to claim 4, wherein the stored single-molecule electrochemical data is processed and analyzed correspondingly by correspondingly and autonomously writing Python processing program.
8. A method for analyzing a weak current detection device based on single-molecule electrochemical analysis, the method comprising:
applying a specified potential to a nano channel or a nano electrode through a current detection port in a shielding box, and receiving a first current signal through a control and data analysis system, wherein the first current signal is a high-frequency sampled weak current signal obtained by single-molecule electrochemical detection;
a current amplifier is intervened through a BNC interface arranged on the shielding box, the current amplifier amplifies and converts the first current signal into a voltage signal, and the voltage signal is transmitted to the control and data analysis system;
and reducing the voltage signal into a second current signal through an analog-to-digital conversion and data processing module according to a current amplification ratio given by the system, filtering to eliminate low-frequency related noise to obtain single-molecule electrochemical data, analyzing the single-molecule electrochemical data, and outputting the single-molecule electrochemical data through an output interface, wherein the analog-to-digital conversion and data processing module is arranged in the control and data analysis system.
9. The analysis method according to claim 8, wherein the sampling frequency of the high-frequency sampling signal is adjusted in a range of 1-500 KHz.
10. Analytical method according to claim 8, characterised in that the amplification ratio of the current is 103-108V/A。
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