CN104849322A - Impedance biosensor and bio-impedance detection analysis method - Google Patents

Abstract

The present invention relates to an impedance biosensor and a bioimpedance analysis method. The impedance biosensor of the present invention includes a microprocessor, an impedance measurement module and a microelectrode module; the microprocessor controls the impedance measurement module to complete the microelectrode module Acquisition and processing analysis of the impedance; the impedance measurement module realizes fast and accurate measurement by setting an impedance measurement chip, a high-pass filter, a voltage follower, an IV buffer and a range regulator, and the biosensor of the present invention has a cost Low, wide frequency band, high precision, quantitative analysis and on-site detection and other advantages.

Description

A kind of impedance biosensor and bioimpedance detection and analysis method

technical field

The invention relates to the technical field of biological impedance detection, in particular to an impedance biosensor and a biological impedance detection and analysis method.

Background technique

Food safety and animal diseases are one of the most prominent problems facing our country. At present, food safety accidents are in a period of frequent occurrence, and highly pathogenic animal diseases also occur from time to time. The screening of pathogenic microorganisms is the key to food safety and animal disease prevention and control. Since the existing routine detection methods are usually unable to achieve on-site rapid detection, there is an urgent need to develop rapid detection technologies for pathogenic microorganisms.

Impedance biosensor is a new type of biological detection technology, which has the advantages of fast detection speed, high sensitivity and simple operation, and has attracted the attention of researchers and enterprises in the fields of agriculture, food, environment and hygiene. Previous studies have shown that its detection accuracy is usually between large-scale laboratory analysis instruments (such as real-time fluorescent quantitative PCR) and conventional rapid detection methods (such as colloidal gold test strips), and it is expected to achieve simple, fast and low-cost detection of pathogenic microorganisms. detection. At present, impedance biosensors are usually composed of impedance biochips and impedance analyzers. Impedance analysis is generally completed by large-scale impedance analysis instruments. Agilent of the United States, Solartron of the United Kingdom, and Zahner of Germany have all developed impedance analysis instruments or electrochemical workstations; In addition, some studies have reported some small impedance detection devices, such as the impedance detection devices independently developed by Ying Yibin (authorized public number CN 203310795 U), Hu Yaohua (authorized public number CN 203241371 U) and others. Ying Yibin's handheld impedance detection device adds pre-processing between the analog signal and the detection electrode, which improves the accuracy and stability of impedance detection. Hu Yaohua's handheld impedance detection device can realize multi-target measurement and has an alarm function.

However, the existing small impedance detection devices have problems such as narrow frequency band, low detection speed and accuracy, and relatively simple functions.

Contents of the invention

The technical problem to be solved by the present invention is to provide an impedance biosensor to realize rapid and accurate measurement and analysis of bioimpedance within a wide frequency range.

In order to solve the above technical problems, the present invention discloses an impedance biosensor, which includes a microprocessor, an impedance measurement module, and a microelectrode module, and target microorganisms are adsorbed on the microelectrode module to change the microelectrode The impedance of the module; the microprocessor controls the impedance measurement module to complete the acquisition of the impedance of the microelectrode module, and processes and analyzes the impedance to obtain the target microbial concentration;

The impedance measurement module includes:

An impedance measurement chip, used to measure the impedance of the microelectrode module, the impedance measurement chip is connected to the microprocessor, and the microprocessor provides control signals for the impedance measurement chip;

A high-pass filter, whose input end is connected to the excitation signal output end of the impedance measurement chip, for filtering low-frequency interference signals;

A voltage follower, whose input end is connected to the output end of the high-pass filter, and whose output end is connected to the input end of the microelectrode module, is used to adjust the DC bias voltage of the excitation signal output by the impedance measurement chip, thereby eliminating a DC component difference between the output and input of the microelectrode module;

I-V buffer, its input terminal is connected to the output terminal of the microelectrode module, and its output terminal is connected to the input pins VIN and RFB of the impedance measurement chip through a feedback resistor, for converting the current signal output by the microelectrode module into a voltage signal and input it to the impedance measurement chip, and the impedance measurement chip processes the received voltage signal to obtain the impedance of the micro-electrode module, and then transmits the impedance of the micro-electrode module to the micro-electrode module processor;

And a range adjuster, the input end of which is connected to the output end of the microelectrode module, and the output end of which is connected to the output end of the I-V buffer, for providing multiple measurement ranges for the impedance measurement chip.

Preferably, the microprocessor adopts an ARM9 processor, and the connection mode between the ARM9 processor and the impedance measurement chip is:

The pin TOUT0 of the internal timer T0 of described ARM9 processor is connected with the external clock pin MCLK of described impedance measurement chip, provides clock signal for described impedance measurement chip; The I ² C interface clock lead of described ARM9 processor The pin I ² CSCL and the I ² C interface data pin I ² CSDA are respectively connected to the I ² C interface clock pin SCL and the I ² C interface data pin SDA of the impedance measurement chip.

Preferably, the microprocessor includes an analysis and calculation unit, which obtains the electron transfer resistance of the microelectrode module at this time according to the impedance of the microelectrode module measured by the impedance measurement chip, and calculates the electron transfer resistance of the microelectrode module at this time. The difference between the electron transfer resistance of the electrode module and the electron transfer resistance when the microelectrode module does not adsorb the target microorganism is used as the change value of the electron transfer resistance, and the relationship model between the change value of the electron transfer resistance and the concentration of the target microorganism is analyzed to obtain Corresponding target microbial concentration;

The analysis and calculation unit includes a model construction subunit, which converts the detection system formed by the microelectrode module into an equivalent circuit, and calculates the impedance of the equivalent circuit, and then obtains the actual value according to the impedance of the equivalent circuit. part, then combined with the real part of the measured impedance and the corresponding angular frequency, use the Newton down-hill algorithm to obtain the electron transfer resistance; obtain the electron transfer resistance at this time and the electron transfer when the microelectrode module does not adsorb the target microorganism The difference in resistance is used as the change value of the electron transfer resistance, and a relationship model between the change value of the electron transfer resistance and the concentration of the target microorganism is established.

In this step, the impedance measurement chip measures the real part and the imaginary part (hexadecimal) of the impedance, and transmits the measured value to the microprocessor, and the microprocessor first corresponds to the impedance according to the impedance measurement chip The actual real and imaginary parts of the impedance of the microprocessor module can be obtained from the conversion formula in the product specification of the product, and then the electron transfer resistance is obtained by using the following formula,

$\mathrm{<span\; class="notranslate">\; R</span>}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; +</span>\frac{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}{{<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; \&omega;</span>}{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{\mathrm{<span\; class="notranslate">\; c</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span>}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; .</span>$

In the formula, R is the real part value measured at a specific angular frequency ω, R _et is the electron transfer resistance of the equivalent circuit, R _s is the solution resistance of the equivalent circuit, and C _dl is the electric double layer capacitance of the equivalent circuit.

Preferably, the range regulator;

The range adjuster includes four signal relays and four feedback resistors of different resistances connected to the four signal relays respectively, the signal relays are respectively connected to the four I/O ports of the microprocessor, The automatic selection of the range of the range adjuster by the microprocessor is realized.

Preferably, the impedance measurement module also includes:

a voltage regulator, which is connected to the impedance measurement chip and used to provide a stable voltage for the impedance measurement chip;

The touch display module is used for displaying test results and parameter setting, wherein the parameters set in the parameter setting include sample number, measurement frequency, resting time, and threshold.

Preferably, the connection mode between the I-V buffer and the impedance measurement chip is:

The output end of the I-V buffer is connected to the input terminal VIN pin of the impedance measurement chip through the feedback resistor RFB1, and the output end of the I-V buffer is connected to the impedance measurement chip through the feedback resistor RFB1 and the feedback resistor RFB2. RFB pin.

Preferably, the microelectrode module includes a substrate, and a first gold electrode, a second gold electrode, a first pad and a second pad located on the substrate;

The first gold electrode is connected to the first pad, and the second gold electrode is connected to the second pad; both the first gold electrode and the second gold electrode are composed of a plurality of finger electrodes of the same size formed in parallel and parallel at the same pitch, and the finger electrodes of the first gold electrode and the second gold electrode cross each other;

The surfaces of the finger electrodes of the first gold electrode and the second gold electrode are decorated with biological recognition materials of target microorganisms.

Preferably, the impedance biosensor also includes:

A USB module, connected to the microprocessor, for performing information access operations with the microprocessor;

A serial port module, connected to the microprocessor, for communicating with the microprocessor;

JTAG module, is connected with described microprocessor, is used for testing described processor;

a storage module, connected to the microprocessor, for storing data in the microprocessor;

The power supply module is connected with the microprocessor, the impedance measurement module and the storage module, and is used for supplying power to the microprocessor, the impedance measurement module and the storage module.

A bioimpedance detection and analysis method, said method comprising the following steps:

Construct the relationship model between the change value of electron transfer resistance and the concentration of target microorganisms: convert the detection system composed of microelectrode modules into an equivalent circuit, calculate the impedance of the equivalent circuit, and then calculate the actual value based on the impedance of the equivalent circuit. part, combined with the real part of the measured impedance of the microelectrode module and the corresponding angular frequency, use the Newton down-hill algorithm to obtain the electron transfer resistance, and calculate the difference between the electron transfer resistance of the microelectrode module and the microelectrode module at this time The difference of the electron transfer resistance when adsorbing the target microorganism is used as the change value of the electron transfer resistance, and the relationship model between the change value of the electron transfer resistance and the concentration of the target microorganism is established;

The change value of the electron transfer resistance of the microelectrode module is measured by using the above-mentioned impedance biosensor, and the corresponding target microorganism concentration is found by using the relationship model.

Preferably, the impedance of the equivalent circuit is:

$\mathrm{<span\; class="notranslate">\; Z</span>}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; +</span>\frac{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{\mathrm{<span\; class="notranslate">\; x</span>}}_{\mathrm{<span\; class="notranslate">\; c</span>}}}{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; x</span>}}_{\mathrm{<span\; class="notranslate">\; c</span>}}}$

Wherein R _et is the electron transfer resistance of the equivalent circuit, X _c is the capacitive reactance of the electric double layer capacitance of the equivalent circuit, and R _s is the solution resistance of the equivalent circuit.

The above-mentioned construction of the relationship model between the change value of the electron transfer resistance and the target microorganism concentration specifically includes the following steps:

S1. Convert the detection system formed by the microelectrode into the equivalent circuit, and calculate the impedance of the equivalent circuit, and as shown in the formula:

$\mathrm{<span\; class="notranslate">\; Z</span>}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; +</span>\frac{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{\mathrm{<span\; class="notranslate">\; x</span>}}_{\mathrm{<span\; class="notranslate">\; c</span>}}}{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; x</span>}}_{\mathrm{<span\; class="notranslate">\; c</span>}}}$

Wherein R _et is the electron transfer resistance of the equivalent circuit, X _c is the capacitive reactance of the electric double layer capacitance of the equivalent circuit, and R _s is the solution resistance of the equivalent circuit.

S2, calculating the capacitive reactance of the electric double layer capacitor as:

${\mathrm{<span\; class="notranslate">\; x</span>}}_{\mathrm{<span\; class="notranslate">\; c</span>}}<span\; class="notranslate">\; =</span>\frac{\mathrm{<span\; class="notranslate">\; 1</span>}}{\mathrm{<span\; class="notranslate">\; j\&omega;</span>}{\mathrm{<span\; class="notranslate">\; c</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; \&omega;</span>}<span\; class="notranslate">\; =</span>\mathrm{<span\; class="notranslate">\; 2</span>}\mathrm{<span\; class="notranslate">\; \&pi;f</span>}$

Wherein, C _dl is the capacitance value of the electric double layer capacitor, f is the frequency, and ω is the angular frequency.

S3. Calculate the real part R of the impedance of the equivalent circuit:

$\mathrm{<span\; class="notranslate">\; R</span>}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; +</span>\frac{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}{{<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; \&omega;</span>}{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{\mathrm{<span\; class="notranslate">\; c</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span>}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1</span>}}$

Convert the real part to:

(R _s -R)(1+ω ² C _dl ² R _et ² )+R _et ＝0

S4, measure and obtain the value of three groups of said real part value R and frequency value ω, record as (R ₁ , ω ₁ ), (R ₂ , ω ₂ ) and (R ₃ , ω ₃ ) respectively, and the equation system can be obtained :

$\left\{\begin{array}{c}{\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span><span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; 1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; =</span>\mathrm{<span\; class="notranslate">\; 0</span>}<span\; class="notranslate">\; ;</span>\\ {\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span><span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; 1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; =</span>\mathrm{<span\; class="notranslate">\; 0</span>}<span\; class="notranslate">\; ;</span>\\ {\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span><span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; 1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; =</span>\mathrm{<span\; class="notranslate">\; 0</span>}<span\; class="notranslate">\; ;</span>\end{array}\right.$

S5, introduce vector

$\mathrm{<span\; class="notranslate">\; f</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span>\left(\begin{array}{c}{\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span>\\ {\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span>\\ {\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span>\end{array}\right)<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; =</span>\left(\begin{array}{c}{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}\\ {\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}\\ {\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}\end{array}\right)<span\; class="notranslate">\; ;</span>$

The formula in the step S4 is expressed as

f(m)=0

S6. Introduce the downhill factor λ to obtain the following calculation formula:

${\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}}<span\; class="notranslate">\; -</span>\mathrm{<span\; class="notranslate">\; \&lambda;</span>}\frac{\mathrm{<span\; class="notranslate">\; f</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}}<span\; class="notranslate">\; )</span>}{{\mathrm{<span\; class="notranslate">\; f</span>}}^{<span\; class="notranslate">\; \&prime;</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}}<span\; class="notranslate">\; )</span>}<span\; class="notranslate">\; ,</span>$

in:

λ=2- ⁿ ,n=0,1,2...

$\begin{array}{c}{\mathrm{<span\; class="notranslate">\; f</span>}}^{<span\; class="notranslate">\; ,</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span>\left(\begin{array}{c}\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}\\ \frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}\\ \frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}\end{array}\right)\\ <span\; class="notranslate">\; =</span>\left(\begin{array}{c}\mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1,1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}\\ \mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1,1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}\\ \mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1,1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}\end{array}\right)\end{array}$

According to Newton's down-hill algorithm, the approximate values of the electron transfer resistance R _et , the electric double layer capacitance C _dl and the solution resistance R _s are solved;

S7. Calculate the difference between the electron transfer resistance of the microelectrode module and the electron transfer resistance when the microelectrode module does not adsorb the target microorganism at this time, as the change value of the electron transfer resistance, and according to the concentration of the target microorganism at this time, establish the The relationship model between the change value of the electron transfer resistance and the concentration of the target microorganism is described.

The technical scheme of the present invention has the following advantages: the present invention generates a low-frequency clock through the internal timer T0 of the ARM9 processor, and expands the single-frequency or narrow-band detection in the existing small impedance detection device to a wider frequency band (100Hz-100kHz) , aiming at the problems of inaccurate electrode polarization and impedance measurement caused by DC bias voltage difference in existing devices, the present invention eliminates the input signal and The DC bias difference between the output signals keeps the DC bias voltage in the entire signal chain constant at VDD/2, and the precise detection range of impedance is extended to 100Ω-100kΩ through segment correction and automatic range adjustment, and by using equivalent The circuit obtains the electron transfer resistance that is really related to the concentration of target microorganisms, and replaces the impedance amplitude used by the predecessors to establish a relationship model with the concentration of target microorganisms for quantitative analysis of the target microorganisms. In addition, most existing devices are developed using single-chip microcomputers, and the human-interaction interface is relatively simple. The present invention adopts the ARM9 processor to develop a touch screen human-computer interaction interface with simpler operation and more intuitive display; in a word, the impedance of the present invention Biosensors have the advantages of fast detection speed, low cost, high precision, wide frequency band, quantitative analysis and on-site detection.

Description of drawings

Fig. 1 is the structural representation of a kind of impedance biosensor of the present invention;

Fig. 2 is the circuit diagram of equivalent circuit among the present invention;

Fig. 3 is the circuit diagram of impedance measurement module among the present invention;

Fig. 4 is the structural representation of microelectrode module among the present invention;

Fig. 5 is a flowchart of biological detection using the device of the present invention.

Detailed ways

The specific implementation manners of the present invention will be further described in detail below in conjunction with the accompanying drawings and examples. The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention.

The invention discloses an impedance biosensor. As shown in FIG. 1, the impedance biosensor includes a microelectrode module 1, a microprocessor 2, and an impedance measurement module 3. The impedance of the microelectrode module 1; the microprocessor 2 controls the impedance measurement module 3 to complete the acquisition, processing and analysis of the impedance of the microelectrode module 1.

As shown in Figure 3, described impedance measurement module 3 comprises: impedance measurement chip 3.2, is used for measuring the impedance of described microelectrode module 1 (obtains the real part and the imaginary part of impedance under different frequencies), and described impedance measurement chip 3.2 Connect with described microprocessor 2, described microprocessor 2 provides control signal for described impedance measurement chip 3.2; High-pass filter 3.4, its input end is connected with the excitation signal output end of described impedance measurement chip 3.2, for Filter low-frequency interference signals; voltage follower 3.5, its input terminal is connected to the output terminal of the high-pass filter 3.4, and its output terminal is connected to the input terminal of the microelectrode module 1 for adjusting the excitation signal output by the impedance measurement chip DC bias voltage, thereby eliminating the DC component difference between the output terminal and the input terminal of the microelectrode module; I-V buffer 3.6, its input terminal is connected to the output terminal of the microelectrode module 1, and its output terminal is fed back Resistors RFB1 and RFB2 are connected to the impedance measurement chip 3.2, and are used to convert the current signal output by the microelectrode module into a voltage signal and input it to the impedance measurement chip 3.2, and process the impedance measurement chip 3.2 to obtain the The impedance of the microelectrode module is passed to the microprocessor; and the range adjuster 3.1, its input terminal is connected to the output terminal of the microelectrode module 1, and its output terminal is connected to the output terminal of the I-V buffer 3.6 for An appropriate measurement file is automatically selected for the impedance measurement chip 3.2.

The connection mode between the I-V buffer 3.6 and the impedance measurement chip 3.2 is: the output terminal of the I-V buffer 3.6 is connected to the VIN pin of the impedance measurement chip 3.2 through the feedback resistor RFB1, and the I-V buffer 3.6 The output terminal of is connected to the RFB pin of the impedance measuring chip 3.2 through the feedback resistor RFB1 and the feedback resistor RFB2.

The range regulator 3.1 includes four signal relays (3.1.1, 3.1.2, 3.1.3, 3.1.4) and four feedback resistors (RF1, RF1, RF2, RF3, RF4), the signal relays are respectively connected with four I/O ports (GPA1, GPA2, GPA3, GPA4) of the microprocessor to realize the control of the microprocessor 2 to the range adjuster 3.1 Automatic range adjustment.

The impedance measurement module also includes: a voltage stabilizer 3.3, which is connected with the impedance measurement chip 3.2, and is used to provide a stable 3V voltage for the impedance measurement chip 3.2; a touch display module, used for detection result display (that is, with Microbial concentration corresponding to the electron transfer resistance) and parameter settings, including sample number, measurement frequency setting, resting time setting, threshold setting, etc.

Described microprocessor 2 adopts TQ2440 core board, integrated an ARM9 processor chip S3C2440 on the board, and the connection mode of described microprocessor 2 and described impedance measuring chip 3.2 is: the internal timing of described microprocessor 2 The device T0 pin TOUT0 is connected with the external clock pin MCLK of the impedance measuring chip 3.2; the I ² C interface communication clock pin I ² CSCL of the microprocessor 2 and the I ² C interface communication data pin I ² CSDA They are respectively connected to the I ² C interface clock pin SCL and the I ² C interface data pin SDA of the impedance measuring core 3.2 slices.

The microprocessor 2 includes an analysis calculation unit, which obtains the electron transfer resistance according to the impedance of the microprocessor module, constructs and stores the relationship model between the change value of the electron transfer resistance and the corresponding target microorganism concentration, and according to the The relational model, combined with the measured impedance, obtains the corresponding target microbial concentration; the analysis and calculation unit includes a model construction subunit, which converts the detection system formed by the microelectrode module into an equivalent circuit, as shown in Figure 2, and calculates the According to the impedance of the equivalent circuit, the real part is obtained according to the impedance of the equivalent circuit, and then the real part value and the corresponding frequency value are obtained in combination with the measurement, and the electron transfer impedance is obtained by using the Newton down-hill algorithm, and the calculated The difference between the electron transfer resistance of the microelectrode module and the electron transfer resistance when the microelectrode module does not adsorb target microorganisms is used as the change value of the electron transfer resistance, and the relationship between the change value of the electron transfer resistance and the concentration of the target microorganism is used The model is used to analyze and obtain the corresponding target microbial concentration.

As shown in Figure 4, the microelectrode module 1 includes a base 1.1, and a first gold electrode 1.2, a second gold electrode 1.3, a first pad 1.4 and a second pad 1.5 located on the base 1.1; The first gold electrode 1.2 is connected to the first pad 1.4, and the second gold electrode 1.3 is connected to the second pad 1.5; the first gold electrode 1.2 and the second gold electrode 1.3 are made of multiple identical The finger electrodes of the same size are formed in parallel parallel with the same pitch, and the finger electrodes of the first gold electrode 1.2 and the second gold electrode 1.3 cross each other; the finger electrodes of the first gold electrode 1.2 and the second gold electrode 1.3 The surface is decorated with biological recognition materials of target microorganisms. The first pad 1.4 and the second pad 1.5 serve as the input end and the output end of the microelectrode module respectively.

The impedance biosensor also includes: a USB module, connected to the microprocessor, for performing information access operations with the microprocessor; a serial port module, connected to the microprocessor, for communicating with the microprocessor The processor communicates; the JTAG module is connected with the microprocessor for testing the microprocessor; the memory module is connected with the microprocessor for storing data in the microprocessor; the power supply module, connected with the microprocessor, impedance measurement module and memory module, for powering the microprocessor, impedance measurement module and memory module; PC, for connecting with the USB module, serial port module and The JTAG module is connected to realize communication and debugging.

The invention also discloses a bio-impedance detection and analysis method. Firstly, the relationship model between the change value of electron transfer resistance and the concentration of target microorganisms is constructed, which includes the following steps: converting the detection system composed of micro-electrode modules into an equivalent circuit, and calculating the The impedance of the equivalent circuit, and then obtain the real part according to the impedance of the equivalent circuit, then combine the real part of the measured impedance signal and the corresponding angular frequency, use Newton's down-hill algorithm to obtain the electron transfer resistance, and calculate the current The difference between the electron transfer resistance of the microelectrode module and the electron transfer resistance when the microelectrode module does not adsorb the target microorganism is used as the change value of the electron transfer resistance, and the relationship model between the change value of the electron transfer resistance and the concentration of the target microorganism is used, The corresponding target microbial concentration is obtained through analysis; then, the corresponding target microbial concentration is found by using the relationship model combined with the measured electron transfer resistance.

Further, constructing the relationship model between the change value of the electron transfer resistance and the target microorganism concentration specifically includes the following steps:

S1. Convert the detection system formed by the microelectrode into the equivalent circuit, and calculate the impedance of the equivalent circuit, and as shown in the formula:

$\mathrm{<span\; class="notranslate">\; Z</span>}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; +</span>\frac{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{\mathrm{<span\; class="notranslate">\; x</span>}}_{\mathrm{<span\; class="notranslate">\; c</span>}}}{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; x</span>}}_{\mathrm{<span\; class="notranslate">\; c</span>}}}$

Wherein R _et is the electron transfer resistance of the equivalent circuit, X _c is the capacitive reactance of the electric double layer capacitance of the equivalent circuit, and R _s is the solution resistance of the equivalent circuit.

S2, calculate the capacitive reactance of the electric double layer capacitor as:

${\mathrm{<span\; class="notranslate">\; x</span>}}_{\mathrm{<span\; class="notranslate">\; c</span>}}<span\; class="notranslate">\; =</span>\frac{\mathrm{<span\; class="notranslate">\; 1</span>}}{\mathrm{<span\; class="notranslate">\; j\&omega;</span>}{\mathrm{<span\; class="notranslate">\; c</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; \&omega;</span>}<span\; class="notranslate">\; =</span>\mathrm{<span\; class="notranslate">\; 2</span>}\mathrm{<span\; class="notranslate">\; \&pi;f</span>}$

Wherein, C _dl is the capacitance value of the electric double layer capacitor, f is the frequency, and ω is the angular frequency.

S3, obtaining the real part R of the equivalent circuit:

$\mathrm{<span\; class="notranslate">\; R</span>}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; +</span>\frac{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}{{<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; \&omega;</span>}{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{\mathrm{<span\; class="notranslate">\; c</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span>}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; .</span>$

The real part can be transformed into:

(R _s -R)(1+ω ² C _dl ² R _et ² )+R _et ＝0

S4, measure and obtain the value of three groups of said real part value R and frequency value ω, record as (R ₁ , ω ₁ ), (R ₂ , ω ₂ ) and (R ₃ , ω ₃ ) respectively, and the equation system can be obtained :

$\left\{\begin{array}{c}{\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span><span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; 1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; =</span>\mathrm{<span\; class="notranslate">\; 0</span>}<span\; class="notranslate">\; ;</span>\\ {\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span><span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; 1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; =</span>\mathrm{<span\; class="notranslate">\; 0</span>}<span\; class="notranslate">\; ;</span>\\ {\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span><span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; 1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; +</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; =</span>\mathrm{<span\; class="notranslate">\; 0</span>}<span\; class="notranslate">\; ;</span>\end{array}\right.$

S5, introduce vector

$\mathrm{<span\; class="notranslate">\; f</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span>\left(\begin{array}{c}{\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span>\\ {\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span>\\ {\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; ,</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}<span\; class="notranslate">\; )</span>\end{array}\right)<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; =</span>\left(\begin{array}{c}{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}\\ {\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}\\ {\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}\end{array}\right)<span\; class="notranslate">\; ;</span>$

The formula in the step S4 is expressed as

f(m)=0

S6. Introduce the downhill factor λ to obtain the following calculation formula:

${\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; =</span>{\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}}<span\; class="notranslate">\; -</span>\mathrm{<span\; class="notranslate">\; \&lambda;</span>}\frac{\mathrm{<span\; class="notranslate">\; f</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}}<span\; class="notranslate">\; )</span>}{{\mathrm{<span\; class="notranslate">\; f</span>}}^{<span\; class="notranslate">\; \&prime;</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}}<span\; class="notranslate">\; )</span>}<span\; class="notranslate">\; ,</span>$

in:

λ=2- ⁿ ,n=0,1,2...

$\begin{array}{c}{\mathrm{<span\; class="notranslate">\; f</span>}}^{<span\; class="notranslate">\; ,</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; m</span>}}_{\mathrm{<span\; class="notranslate">\; k</span>}}<span\; class="notranslate">\; )</span><span\; class="notranslate">\; =</span>\left(\begin{array}{c}\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}\\ \frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}\\ \frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}}<span\; class="notranslate">\; ,</span>\frac{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; f</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; (</span>\mathrm{<span\; class="notranslate">\; m</span>}<span\; class="notranslate">\; )</span>}{{<span\; class="notranslate">\; \&PartialD;</span>\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}\end{array}\right)\\ <span\; class="notranslate">\; =</span>\left(\begin{array}{c}\mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1,1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 1</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}\\ \mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1,1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 2</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}\\ \mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; +</span>\mathrm{<span\; class="notranslate">\; 1,1</span>}<span\; class="notranslate">\; +</span>{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}<span\; class="notranslate">\; ,</span>\mathrm{<span\; class="notranslate">\; 2</span>}<span\; class="notranslate">\; (</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; the\; s</span>}}<span\; class="notranslate">\; -</span>{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}<span\; class="notranslate">\; )</span>{\mathrm{<span\; class="notranslate">\; C</span>}}_{\mathrm{<span\; class="notranslate">\; dl</span>}}{{\mathrm{<span\; class="notranslate">\; R</span>}}_{\mathrm{<span\; class="notranslate">\; et</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}{{\mathrm{<span\; class="notranslate">\; \&omega;</span>}}_{\mathrm{<span\; class="notranslate">\; 3</span>}}}^{\mathrm{<span\; class="notranslate">\; 2</span>}}\end{array}\right)\end{array}$

According to Newton's down-hill algorithm, solve the electron transfer resistance R _et , the approximate value of the electric double layer capacitance C _dl and the solution resistance R _s ;

S7. Calculate the difference between the electron transfer resistance of the microelectrode module and the electron transfer resistance when the microelectrode module does not adsorb the target microorganism at this time, as the change value of the electron transfer resistance, and according to the concentration of the target microorganism at this time, establish the The relationship model between the change value of the electron transfer resistance and the concentration of the target microorganism is described.

Further, the microprocessor 2 is a microprocessor based on ARM9 core, which integrates abundant on-chip resources, including a memory part, a clock part and a power supply part.

Further, the impedance measurement chip 3.2 adopts a high-precision impedance conversion integrated circuit AD5933 or AD5934 released by AD Company, and a frequency generator and an analog-to-digital converter are integrated on the chip.

Furthermore, the model of the impedance measurement chip 3.2 is AD5933 or AD5934, the model of the voltage regulator 3.3 is ADR433, the model of the voltage follower 3.5 is 1/2AD8606, the model of the I-V buffer 3.6 is 1/2AD8606, and the four signal relays 3.1 .1, 3.1.2, 3.1.3 and 3.1.4 models are all G6E-134P-US, six feedback resistors (RFB1, RFB2, RF1, RF2, RF3 and RF4) and four bias resistors (R1, R2, R3 and R4) all use precision resistors with a precision of 0.1%, controlled by four I/O ports GPA1, GPA2, GPA3 and GPA4 of the microprocessor 2.

Fig. 5 is a flow chart of an impedance biosensor biological detection in a preferred embodiment of the present invention. Both the surfaces of the first electrode 1.2 and the second electrode 1.3 are modified with biorecognition materials of target microorganisms (taking antibodies as an example): first, protein A is adsorbed on the first electrode 1.2 and the second electrode through electrostatic and hydrophobic interactions. On the surface of 1.3, use protein A to specifically bind to the Fc segment of the immunoglobulin of the antibody to immobilize the antibody on the first electrode 1.2 and the second electrode 1.3, and finally use bovine serum albumin to block the remaining binding sites Avoid non-specific reactions. During implementation, the impedance measurement is first performed, and the electron transfer resistance R _etc of the control is obtained through the equivalent circuit analysis of the electrode, and then the sample containing the target microorganism is dropped on the electrode, and the target microorganism will be detected by the electrode. 1.2, 1.3 Captured by the antibody on the surface, and then washed with PBS, the redox probe ([Fe(CN) ₆ ] ^3-/4- ) was added dropwise to measure the electrode impedance, which was obtained by the equivalent circuit analysis of the electrode For the electron transfer resistance R _ets of the sample, calculate the difference △R _et between R _ets and R _etc , and then use the relationship model between △R _et and the concentration of target microorganisms to quantitatively detect the target microorganisms.

Table 1 is a comparison of the electron transfer resistance R _et value obtained by using the biological impedance analysis method of the present invention and the electron transfer resistance ZR _et value obtained by the existing impedance spectrum analysis software Zsimpwin. The present invention compares and analyzes the experimental data collected by the electrodes in the process of biological modification and target microorganism detection (protein A immobilization, antibody coupling, bovine serum albumin BSA sealing and target capture), Zsimpwin software uses the same equivalent circuit The experimental data is simulated to obtain the electron transfer impedance ZR _et . The present invention extracts 15 data from different frequencies from the experimental data, namely low frequency (100, 100 ± 5Hz and 100 ± 10Hz), intermediate frequency (5kHz, 5 ± 0.25kHz and 5±0.5kHz) and high frequency (85kHz, 85±1kHz and 85±2kHz), and then select a group of data from each of the low, middle and high frequency, and the frequency and real part of each group of data follow the above method For processing, that is, to use Newton's downhill method to solve, five electron transfer resistance values can be obtained; finally, the bubble method is used to sort, and the average value of the middle three values is taken as the electron transfer resistance R _et . It can be seen that the biological impedance analysis method of the present invention is in good agreement with the results of Zsimpwin.

Table 1 The biological impedance analysis method of the present invention compares with the result of existing impedance analysis software

Aiming at the problems of narrow frequency band, low detection accuracy and relatively simple functions in existing small impedance detection devices, the present invention adopts the 125kHz low-frequency clock generated by the internal timer of ARM9 processor (S3C2440) to solve the low-frequency response of AD5933 or AD5934 question. Aiming at the problems of electrode polarization and inaccurate impedance measurement caused by DC bias voltage difference in the existing impedance detection device, the present invention eliminates the input signal of the microelectrode module through a high-pass filter, a voltage follower and an I-V buffer and the DC bias difference between the output signal, so that the DC bias voltage in the entire signal chain is constant at VDD/2, and the impedance detection range is extended to 100Ω-100kΩ through segmental correction and automatic range adjustment, all bias resistors Both the resistor and the feedback resistor use a resistor with a precision of 0.1% to reduce the inaccuracy, which effectively improves the measurement accuracy, and obtains the electron transfer resistance that is truly related to the target microbial concentration by using an equivalent circuit, instead of the impedance amplitude used by the predecessors , to establish a relationship model with the concentration of target microorganisms, and carry out quantitative analysis of target microorganisms. In addition, most existing devices are developed by single-chip microcomputers, and the human-interaction interface is relatively simple. The present invention adopts the ARM9 processor to develop a touch-screen human-computer interaction interface with simpler operation and more intuitive display.

Finally, it should be noted that: the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that: it can still be Modifications are made to the technical solutions described in the foregoing embodiments, or equivalent replacements are made to some of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the various embodiments of the present invention.

Claims (10)

1. an impedance biosensor, is characterized in that, described impedance biosensor comprises microprocessor, impedance measurement module and microelectrode module, and objective microbe to be adsorbed in described microelectrode module thus to change the impedance of described microelectrode module; Impedance measurement module described in described Microprocessor S3C44B0X completes the acquisition of the impedance of described microelectrode module, and carries out Treatment Analysis to described impedance and obtain objective microbe concentration;

Described impedance measurement module comprises:

Impedance measurement chip, for measuring the impedance of described microelectrode module, described impedance measurement chip is connected with described microprocessor, and described microprocessor provides control signal for described impedance measurement chip;

Hi-pass filter, its input end is connected with the pumping signal output terminal of described impedance measurement chip, for filtering low undesired signal;

Voltage follower, its input end connects the output terminal of described Hi-pass filter, its output terminal connects the input end of described microelectrode module, the DC offset voltage of the pumping signal exported for regulating described impedance measurement chip, thus eliminate the DC component difference between the output terminal of described microelectrode module and input end;

I-V impact damper, its input end connects the output terminal of described microelectrode module, its output terminal connects input pin VIN and RFB of described impedance measurement chip by feedback resistance, current signal for described microelectrode module being exported converts voltage signal to and is input to described impedance measurement chip, and by described impedance measurement chip, the described voltage signal process received is obtained the impedance of described microelectrode module, then the impedance of described microelectrode module is passed to described microprocessor;

And range adjuster, its input end connects the output terminal of described microelectrode module, and its output terminal connects the output terminal of described I-V impact damper, for providing multiple measurement shelves for described impedance measurement chip.

2. impedance biosensor according to claim 1, is characterized in that, described microprocessor adopts ARM9 processor, and the connected mode of described ARM9 processor and described impedance measurement chip is:

The pin TOUT0 of the timer internal T0 of described ARM9 processor is connected with the external clock pin MCLK of described impedance measurement chip, for described impedance measurement chip provides clock signal; The I of described ARM9 processor ²c interface clock pins I ²cSCL and I ²c interface data pin I ²cSDA respectively with the I of described impedance measurement chip ²c interface clock pins SCL and I ²c interface data pin SDA connects.

3. impedance biosensor according to claim 2, it is characterized in that, described microprocessor comprises analytical calculation unit, the impedance of its described microelectrode module obtained according to the measurement of described impedance measurement chip obtains the electro transfer resistance of now described microelectrode module, and the difference of electro transfer resistance when calculating the non-adsorbed target microorganism of now the electro transfer resistance of described microelectrode module and described microelectrode module, as electro transfer increased resistance value, utilize the relational model of described electro transfer increased resistance value and objective microbe concentration, analyze and obtain corresponding objective microbe concentration,

Described analytical calculation unit comprises model construction subelement, the detection system of described microelectrode module composition is converted into equivalent electrical circuit by it, and calculate the impedance of described equivalent electrical circuit, its real part is asked for again according to the impedance of described equivalent electrical circuit, afterwards in conjunction with real part and the corresponding angular frequency of the impedance of measurement, newton's higher education park is utilized to ask for electro transfer resistance; The difference of electro transfer resistance when asking for described electro transfer resistance now and the non-adsorbed target microorganism of described microelectrode module, as electro transfer increased resistance value, sets up the relational model of described electro transfer increased resistance value and objective microbe concentration.

4. impedance biosensor according to claim 3, is characterized in that, described range adjuster;

The feedback resistance of four different resistances that described range adjuster comprises four signal relays and connects from described four signal relays respectively, described signal relay is connected with four I/O mouths of described microprocessor respectively, realizes the automatic selection of described microprocessor to the range of described range adjuster.

5. impedance biosensor according to claim 4, is characterized in that, described impedance measurement module also comprises:

Voltage stabilizer, it is connected with described impedance measurement chip, for providing stable voltage for described impedance measurement chip;

Touch display module, for testing result display and optimum configurations, the parameter arranged in wherein said optimum configurations comprises sample number, survey frequency, time of repose, threshold value.

6. impedance biosensor according to claim 5, is characterized in that, the connected mode of described I-V impact damper and described impedance measurement chip is:

The output terminal of described I-V impact damper is connected to the input end VIN pin of described impedance measurement chip by feedback resistance RFB1, the output terminal of described I-V impact damper is connected to the RFB pin of described impedance measurement chip by feedback resistance RFB1 and feedback resistance RFB2.

7. impedance biosensor according to claim 6, is characterized in that, described microelectrode module comprises substrate and is positioned at described suprabasil first gold electrode, the second gold electrode, the first pad and the second pad;

Described first gold electrode is connected with described first pad, and described second gold electrode is connected with described second pad; Described first gold electrode is combined to form with identical spacing parallel connection with the finger electrode of the second gold electrode by multiple same size, and the finger electrode of described first gold electrode and the second gold electrode intersects mutually;

The surface of the finger electrode of described first gold electrode and the second gold electrode is all modified with the bio-identification material of objective microbe.

8. impedance biosensor according to claim 7, is characterized in that, described impedance biosensor also comprises:

USB module, is connected with described microprocessor, for carrying out Information Access operation with described microprocessor;

Serial port module, is connected with described microprocessor, for carrying out communication with described microprocessor;

JTAG module, is connected with described microprocessor, for testing described processor;

Reservoir module, is connected with described microprocessor, for storing the data in described microprocessor;

Power module, is connected with described microprocessor, impedance measurement module and reservoir module, for powering for described microprocessor, impedance measurement module and reservoir module.

9. a bio-impedance determination method, is characterized in that, said method comprising the steps of:

Build the relational model of electro transfer increased resistance value and objective microbe concentration: the detection system of microelectrode module composition is converted into equivalent electrical circuit, and calculate the impedance of described equivalent electrical circuit, its real part is asked for again according to the impedance of described equivalent electrical circuit, afterwards in conjunction with real part and the corresponding angular frequency of the impedance of the microelectrode module of measurement, newton's higher education park is utilized to ask for electro transfer resistance, and the difference of electro transfer resistance when calculating the non-adsorbed target microorganism of now the electro transfer resistance of described microelectrode module and described microelectrode module, as electro transfer increased resistance value, set up the relational model of described electro transfer increased resistance value and objective microbe concentration,

Utilize the impedance biosensor measurement described in any one of claim 1 to 8 to obtain the electro transfer increased resistance value of described microelectrode module, and utilize described relational model, find corresponding objective microbe concentration.

10. method according to claim 9, is characterized in that, the impedance of described equivalent electrical circuit is:

$Z=R_s+\frac{R_{\mathrm{et}}{X}_c}{R_{\mathrm{et}}+X_c}$

Wherein R _etfor the electro transfer resistance of described equivalent electrical circuit, X _cfor the capacitive reactance of the electric double layer capacitance of described equivalent electrical circuit, R _sfor the solution resistance of equivalent electrical circuit.