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

Abstract

The invention relates to an impedance biosensor and a bio-impedance detection analysis method. The impedance biosensor comprises a microprocessor, an impedance detection module and a microelectrode module. The microprocessor controls the impedance detection module so that acquisition and processing analysis of impedance of the microelectrode module are realized. The impedance detection module is provided with an impedance detection chip, a high-pass filter, a voltage follower, an I-V buffer and a range adjuster and realizes fast and accurate measurement. The impedance biosensor has the advantages of low cost, wide frequency band, high precision, quantitative analysis and on-site detection.

Description

A kind of impedance biosensor and bio-impedance determination method

Technical field

The present invention relates to bio-impedance detection technique field, particularly relate to a kind of impedance biosensor and bio-impedance determination method.

Background technology

Food security and animal epidemic are one of most outstanding problems of facing of China.Current, food security accident is in the multiple phase, and highly pathogenic animal epidemic also happens occasionally.Pathogenic microorganism examination is the key of food security and Field of Animal Epidemic Disease Control, because existing common detection methods cannot realize field quick detection usually, therefore in the urgent need to development pathogenic microorganism Fast Detection Technique.

Impedance biosensor is a kind of new bio detection technique, the advantage such as have that detection speed is very fast, sensitivity is higher and operation is simpler, has obtained the concern of the field researchers such as agricultural, food, environment and health and enterprise.Forefathers' research shows: its accuracy of detection, usually between large-scale experiment room analytical instrument (as real-time fluorescence quantitative PCR) and conventional method for quick (as colloidal gold strip), is expected to the detection realizing the simple, quick of pathogenic microorganism and low cost.At present, impedance biosensor is made up of impedance biochip and electric impedance analyzer usually, impedance analysis has generally been come by large-scale impedance analysis instrument, and U.S. Agilent, Solartron company of Britain, German Zahner company all have developed impedance analysis instrument or electrochemical workstation; In addition, some small-sized impedance detection devices have also been studies have reported that, as the impedance detection device in people's independent developments such as Ying Yibin (authorizing public CN 203310795 U), Hu Yaohua (authorizing public CN 203241371 U).The hand-held impedance detection device of Ying Yibin adds pre-process between simulating signal and detecting electrode, improves the precision and stability of impedance detection.The hand-held impedance detection device of Hu Yaohua can realize multiple goal and measure, and has warning function.

But existing small-sized impedance detection device exists, and frequency band is narrow, detection speed and the problem such as precision is on the low side, function is simpler.

Summary of the invention

The technical problem to be solved in the present invention is to provide a kind of impedance biosensor, realizes quick and precisely measurement and the analysis of bio-impedance within the scope of broad frequency band.

For solving the problems of the technologies described above, the invention discloses a kind of impedance biosensor, 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.

Preferably, 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.

Preferably, 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.

In this step, the measurement of described impedance measurement chip obtains real part and the imaginary part (sexadecimal) of impedance, and give described microprocessor by transmitting measured values, first described microprocessor obtains real part and the imaginary part of the reality of the impedance of microprocessor module according to the conversion formula in product description corresponding to described impedance measurement chip, formula is below utilized to ask for electro transfer resistance afterwards

$R=R_s+\frac{R_{\mathrm{et}}}{{\left(\mathrm{\ω}{R}_{\mathrm{et}}{c}_{\mathrm{dl}}\right)}^2+1}.$

In formula, R obtains value of real part, R for measuring under specific angle frequencies omega _etfor the electro transfer resistance of equivalent electrical circuit, R _sfor the solution resistance of equivalent electrical circuit, C _dlfor the electric double layer capacitance of equivalent electrical circuit.

Preferably, 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.

Preferably, 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.

Preferably, 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.

Preferably, 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.

Preferably, 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.

A kind of bio-impedance determination method, 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 above-mentioned impedance biosensor measurement to obtain the electro transfer increased resistance value of described microelectrode module, and utilize described relational model, find corresponding objective microbe concentration.

Preferably, 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.

The described relational model of above-mentioned structure described electro transfer increased resistance value and objective microbe concentration specifically comprises the following steps:

S1, the detection system that microelectrode is formed is converted into described equivalent electrical circuit, and calculates the impedance of described equivalent electrical circuit, and as shown by the equation:

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

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 described equivalent electrical circuit.

S2, the capacitive reactance calculating described electric double layer capacitance are:

$X_c=\frac{1}{\mathrm{j\ω}{c}_{\mathrm{dl}}},\mathrm{\ω}=2\mathrm{\πf}$

Wherein, C _dlfor the capacitance of described electric double layer capacitance, f is frequency, and ω is angular frequency.

S3, ask for the real part R of the impedance of described equivalent electrical circuit:

$R=R_s+\frac{R_{\mathrm{et}}}{{\left(\mathrm{\ω}{R}_{\mathrm{et}}{c}_{\mathrm{dl}}\right)}^2+1}$

Real part is converted into:

(R _s-R)(1+ω ²C _dl ²R _et ²)+R _et＝0

S4, measure and obtain the value of value of real part R and frequency values ω described in three groups, be designated as (R respectively ₁, ω ₁), (R ₂, ω ₂) and (R ₃, ω ₃), can system of equations be obtained:

$\left\{\begin{array}{c}{f}_1(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})=(R_s-R_1)(1+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_1}^2)+R_{\mathrm{et}}=0;\\ f_2(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})=(R_s-R_2)(1+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_2}^2)+R_{\mathrm{et}}=0;\\ f_3(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})=(R_s-R_3)(1+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_3}^2)+R_{\mathrm{et}}=0;\end{array}\right.$

S5, introducing vector

$f(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})=\left(\begin{array}{c}{f}_1(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})\\ f_2(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})\\ f_3(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})\end{array}\right),m=\left(\begin{array}{c}{R}_{\mathrm{et}}\\ R_s\\ C_{\mathrm{dl}}\end{array}\right);$

Formula in described step S4 is expressed as

f(m)＝0

S6, introduce go down the hill factor lambda, draw following computing formula:

$m_{k+1}=m_k-\mathrm{\λ}\frac{f\left(m_k\right)}{f^{\′}\left(m_k\right)},$

Wherein:

λ＝2 ^-n,n＝0,1,2......

$\begin{array}{c}{f}^{,}\left(m_k\right)=\left(\begin{array}{c}\frac{{\∂f}_1\left(m\right)}{{\∂R}_{\mathrm{et}}},\frac{{\∂f}_1\left(m\right)}{{\∂R}_s},\frac{{\∂f}_1\left(m\right)}{{\∂C}_{\mathrm{dl}}}\\ \frac{{\∂f}_2\left(m\right)}{{\∂R}_{\mathrm{et}}},\frac{{\∂f}_2\left(m\right)}{{\∂R}_s},\frac{{\∂f}_2\left(m\right)}{{\∂C}_{\mathrm{dl}}}\\ \frac{{\∂f}_3\left(m\right)}{{\∂R}_{\mathrm{et}}},\frac{{\∂f}_3\left(m\right)}{{\∂R}_s},\frac{{\∂f}_3\left(m\right)}{{\∂C}_{\mathrm{dl}}}\end{array}\right)\\ =\left(\begin{array}{c}2(R_s-R_1)R_{\mathrm{et}}{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_1}^2+\mathrm{1,1}+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_1}^2,2(R_s-R_1)C_{\mathrm{dl}}{R_{\mathrm{et}}}^2{{\mathrm{\ω}}_1}^2\\ 2(R_s-R_2)R_{\mathrm{et}}{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_2}^2+\mathrm{1,1}+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_2}^2,2(R_s-R_2)C_{\mathrm{dl}}{R_{\mathrm{et}}}^2{{\mathrm{\ω}}_2}^2\\ 2(R_s-R_3)R_{\mathrm{et}}{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_3}^2+\mathrm{1,1}+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_3}^2,2(R_s-R_3)C_{\mathrm{dl}}{R_{\mathrm{et}}}^2{{\mathrm{\ω}}_3}^2\end{array}\right)\end{array}$

According to newton's higher education park, solve described electro transfer resistance R _et, described electric double layer capacitance C _dland described solution resistance R _sapproximate value;

The difference of S7, 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, according to objective microbe concentration now, set up the relational model of described electro transfer increased resistance value and objective microbe concentration.

Technique scheme tool of the present invention has the following advantages: the present invention produces a low-frequency clock by ARM9 processor timer internal T0, be broad frequency band (100Hz-100kHz) by the unifrequency in existing small-sized impedance detection device or narrow-band extension of detecting capability, for the electrode polarization caused by DC offset voltage difference in existing apparatus and the problem such as impedance measurement is inaccurate, the present invention is by a Hi-pass filter, direct current biasing difference between the input signal of a voltage follower and an I-V impact damper elimination microelectrode module and output signal, make the DC offset voltage in whole signal chains constant in VDD/2, automatically regulate accurate for impedance extended detection range to 100 Ω-100k Ω by Stepwise calibration and range, and by adopting equivalent electrical circuit to ask for the very positively related electro transfer resistance with objective microbe concentration, replace the impedance magnitude that forefathers adopt, set up the relational model with objective microbe concentration, carry out the quantitative test of objective microbe.In addition, mostly existing apparatus is to utilize single-chip microcomputer to develop, fairly simple on interpersonal interactive interface, and the present invention adopts ARM9 processor to develop the simpler and more intuitive touch display screen human-computer interaction interface of display of operation; In a word, impedance biosensor of the present invention have that detection speed is very fast, cost is lower, precision is higher, frequency band is wider, the advantage such as quantitative test and Site Detection.

Accompanying drawing explanation

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

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

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

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

Fig. 5 is the process flow diagram utilizing device of the present invention to carry out biological detection.

Embodiment

Below in conjunction with drawings and Examples, the specific embodiment of the present invention is described in further detail.Following examples for illustration of the present invention, but are not used for limiting the scope of the invention.

The invention discloses a kind of impedance biosensor, as shown in Figure 1, described impedance biosensor comprises microelectrode module 1, microprocessor 2 and impedance measurement module 3, and biological adsorption is in described microelectrode module 1 thus change the impedance of described microelectrode module 1; Described microprocessor 2 controls acquisition and the Treatment Analysis that described impedance measurement module 3 completes the impedance of described microelectrode module 1.

As shown in Figure 3, described impedance measurement module 3 comprises: impedance measurement chip 3.2, for measuring the impedance (obtaining real part and the imaginary part of impedance under different frequency) of described microelectrode module 1, described impedance measurement chip 3.2 is connected with described microprocessor 2, and described microprocessor 2 provides control signal for described impedance measurement chip 3.2; Hi-pass filter 3.4, its input end is connected with the pumping signal output terminal of described impedance measurement chip 3.2, for filtering low undesired signal; Voltage follower 3.5, its input end connects the output terminal of described Hi-pass filter 3.4, its output terminal connects the input end of described microelectrode module 1, 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 3.6, its input end connects the output terminal of described microelectrode module 1, its output terminal is connected described impedance measurement chip 3.2 by feedback resistance RFB1 with RFB2, current signal for described microelectrode module being exported converts voltage signal to and is input to described impedance measurement chip 3.2, and the impedance obtaining described microelectrode module is processed by described impedance measurement chip 3.2, pass to described microprocessor; And range adjuster 3.1, its input end connects the output terminal of described microelectrode module 1, and its output terminal connects the output terminal of described I-V impact damper 3.6, for automatically selecting suitable measurement shelves for described impedance measurement chip 3.2.

Described I-V impact damper 3.6 with the connected mode of described impedance measurement chip 3.2 is: the output terminal of described I-V impact damper 3.6 is connected to the VIN pin of described impedance measurement chip 3.2 by feedback resistance RFB1, the output terminal of described I-V impact damper 3.6 is connected to the RFB pin of described impedance measurement chip 3.2 by feedback resistance RFB1 and feedback resistance RFB2.

The feedback resistance (RF1, RF2, RF3, RF4) of four different resistances that described range adjuster 3.1 comprises four signal relays (3.1.1,3.1.2,3.1.3,3.1.4) and connects from described four signal relays respectively, described signal relay is connected with described microprocessor four I/O mouths (GPA1, GPA2, GPA3, GPA4) respectively, realizes the range of described microprocessor 2 to described range adjuster 3.1 and automatically regulates.

Described impedance measurement module also comprises: voltage stabilizer 3.3, and it is connected with described impedance measurement chip 3.2, for providing stable 3V voltage for described impedance measurement chip 3.2; Touch display module, for testing result display (namely corresponding with described electro transfer resistance microorganism concn) and optimum configurations, comprises sample number, survey frequency is arranged, time of repose is arranged, threshold value arranges.

Described microprocessor 2 adopts TQ2440 core board, plate is integrated with an ARM9 processor chips S3C2440, and described microprocessor 2 with the connected mode of described impedance measurement chip 3.2 is: the timer internal T0 pin TOUT0 of described microprocessor 2 is connected with the external clock pin MCLK of described impedance measurement chip 3.2; The I of described microprocessor 2 ²c interface communication clock pin I ²cSCL and I ²c interface communication data pin I ²cSDA respectively with the I of 3.2, described impedance measurement core ²c interface clock pins SCL and I ²c interface data pin SDA connects.

Described microprocessor 2 comprises analytical calculation unit, it obtains electro transfer resistance according to the impedance of described microprocessor module, build and store the relational model of described electro transfer increased resistance value and corresponding objective microbe concentration, and according to described relational model, in conjunction with the impedance of measuring, obtain corresponding objective microbe concentration, described analytical calculation unit comprises model construction subelement, the detection system of microelectrode module composition is converted into equivalent electrical circuit by it, as shown in Figure 2, and calculate the impedance of described equivalent electrical circuit, its real part is asked for according to the impedance of described equivalent electrical circuit, combine afterwards to measure and obtain described value of real part and corresponding frequency values, described newton's higher education park is utilized to ask for electro transfer impedance, 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.

As shown in Figure 4, described microelectrode module 1 comprises substrate 1.1 and is positioned at the first gold electrode 1.2, second gold electrode 1.3, first pad 1.4 and the second pad 1.5 in described substrate 1.1; Described first gold electrode 1.2 is connected with described first pad 1.4, and described second gold electrode 1.3 is connected with described second pad 1.5; Described first gold electrode 1.2 is combined to form with identical spacing parallel connection with the finger electrode of the second gold electrode 1.3 by multiple same size, and the finger electrode of described first gold electrode 1.2 and the second gold electrode 1.3 intersects mutually; The surface of the finger electrode of described first gold electrode 1.2 and the second gold electrode 1.3 is all modified with the bio-identification material of objective microbe.Described first pad 1.4 and the second pad 1.5 are respectively as the input end of described microelectrode module and output terminal.

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 microprocessor; 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; PC, for described USB module, serial port module and JTAG model calling, realize communication and debugging.

The invention also discloses a kind of bio-impedance determination method, first the relational model of electro transfer increased resistance value and objective microbe concentration is built, comprise the following steps: 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 signal 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, utilize the relational model of described electro transfer increased resistance value and objective microbe concentration, analyze and obtain corresponding objective microbe concentration, afterwards, utilize described relational model, in conjunction with the described electro transfer resistance measured, find corresponding objective microbe concentration.

Further, the described relational model building described electro transfer increased resistance value and objective microbe concentration specifically comprises the following steps:

S1, the detection system that microelectrode is formed is converted into described equivalent electrical circuit, and calculates the impedance of described equivalent electrical circuit, and as shown by the equation:

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

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.

S2, the capacitive reactance calculating described electric double layer capacitance are:

$X_c=\frac{1}{\mathrm{j\ω}{c}_{\mathrm{dl}}},\mathrm{\ω}=2\mathrm{\πf}$

Wherein, C _dlfor the capacitance of described electric double layer capacitance, f is frequency, and ω is angular frequency.

S3, ask for the real part R of described equivalent electrical circuit:

$R=R_s+\frac{R_{\mathrm{et}}}{{\left(\mathrm{\ω}{R}_{\mathrm{et}}{c}_{\mathrm{dl}}\right)}^2+1}.$

Real part can be converted into:

(R _s-R)(1+ω ²C _dl ²R _et ²)+R _et＝0

S4, measure and obtain the value of value of real part R and frequency values ω described in three groups, be designated as (R respectively ₁, ω ₁), (R ₂, ω ₂) and (R ₃, ω ₃), can system of equations be obtained:

$\left\{\begin{array}{c}{f}_1(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})=(R_s-R_1)(1+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_1}^2)+R_{\mathrm{et}}=0;\\ f_2(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})=(R_s-R_2)(1+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_2}^2)+R_{\mathrm{et}}=0;\\ f_3(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})=(R_s-R_3)(1+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_3}^2)+R_{\mathrm{et}}=0;\end{array}\right.$

S5, introducing vector

$f(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})=\left(\begin{array}{c}{f}_1(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})\\ f_2(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})\\ f_3(R_{\mathrm{et}},R_s,C_{\mathrm{dl}})\end{array}\right),m=\left(\begin{array}{c}{R}_{\mathrm{et}}\\ R_s\\ C_{\mathrm{dl}}\end{array}\right);$

Formula in described step S4 is expressed as

f(m)＝0

S6, introduce go down the hill factor lambda, obtain following computing formula:

$m_{k+1}=m_k-\mathrm{\λ}\frac{f\left(m_k\right)}{f^{\′}\left(m_k\right)},$

Wherein:

λ＝2 ^-n,n＝0,1,2......

$\begin{array}{c}{f}^{,}\left(m_k\right)=\left(\begin{array}{c}\frac{{\∂f}_1\left(m\right)}{{\∂R}_{\mathrm{et}}},\frac{{\∂f}_1\left(m\right)}{{\∂R}_s},\frac{{\∂f}_1\left(m\right)}{{\∂C}_{\mathrm{dl}}}\\ \frac{{\∂f}_2\left(m\right)}{{\∂R}_{\mathrm{et}}},\frac{{\∂f}_2\left(m\right)}{{\∂R}_s},\frac{{\∂f}_2\left(m\right)}{{\∂C}_{\mathrm{dl}}}\\ \frac{{\∂f}_3\left(m\right)}{{\∂R}_{\mathrm{et}}},\frac{{\∂f}_3\left(m\right)}{{\∂R}_s},\frac{{\∂f}_3\left(m\right)}{{\∂C}_{\mathrm{dl}}}\end{array}\right)\\ =\left(\begin{array}{c}2(R_s-R_1)R_{\mathrm{et}}{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_1}^2+\mathrm{1,1}+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_1}^2,2(R_s-R_1)C_{\mathrm{dl}}{R_{\mathrm{et}}}^2{{\mathrm{\ω}}_1}^2\\ 2(R_s-R_2)R_{\mathrm{et}}{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_2}^2+\mathrm{1,1}+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_2}^2,2(R_s-R_2)C_{\mathrm{dl}}{R_{\mathrm{et}}}^2{{\mathrm{\ω}}_2}^2\\ 2(R_s-R_3)R_{\mathrm{et}}{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_3}^2+\mathrm{1,1}+{R_{\mathrm{et}}}^2{C_{\mathrm{dl}}}^2{{\mathrm{\ω}}_3}^2,2(R_s-R_3)C_{\mathrm{dl}}{R_{\mathrm{et}}}^2{{\mathrm{\ω}}_3}^2\end{array}\right)\end{array}$

According to newton's higher education park, solve described electro transfer resistance R _et, described electric double layer capacitance C _dland described solution resistance R _sapproximate value;

The difference of S7, 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, according to objective microbe concentration now, set up the relational model of described electro transfer increased resistance value and objective microbe concentration.

Further, described microprocessor 2 is the microprocessors based on ARM9 kernel, is integrated with abundant Resources on Chip, comprises memory portion, clock part and power unit.

Further, a kind of high precision impedance transformation integrated circuit AD5933 or AD5934 that described impedance measurement chip 3.2 adopts AD company to release, sheet is integrated with frequency generator and analog to digital converter.

Further, the model of impedance measurement chip 3.2 is AD5933 or AD5934, the model of voltage stabilizer 3.3 is ADR433, the model of voltage follower 3.5 is 1/2AD8606, the model of I-V impact damper 3.6 is 1/2AD8606, four signal relay 3.1.1, 3.1.2, 3.1.3 G6E-134P-US is with the model of 3.1.4, six feedback resistance (RFB1, RFB2, RF1, RF2, RF3 and RF4) and four biasing resistor (R1, R2, R3 and R4) precision resistance of equal service precisions 0.1%, by four I/O mouth GPA1 of microprocessor 2, GPA2, GPA3 and GPA4 controls.

Fig. 5 is the process flow diagram of a kind of impedance biosensor biological detection of a preferred embodiment of the present invention.Described first electrode 1.2, second electrode 1.3 is all modified with the bio-identification material (for antibody) of objective microbe on the surface: be first adsorbed on the surface of described first electrode 1.2, second electrode 1.3 by electrostatic and hydrophobic effect by albumin A, antibody is fixed on described first electrode 1.2, second electrode 1.3 by the Fc section generation specific binding of the immunoglobulin (Ig) of recycling albumin A and antibody, finally utilizes bovine serum albumin to close residual binding site and avoids nonspecific reaction.During enforcement, first carry out impedance measurement, obtained the electro transfer resistance R contrasted by the Analysis of Equivalent Circuit of described electrode _etc, afterwards the sample drop comprising objective microbe is added on the electrodes, described objective microbe by catch by the antibody on described electrode 1.2,1.3 surface, after recycling PBS cleaning, drip redox probe ([Fe (CN) ₆] ^3-/4-) potential electrode impedance, the electro transfer resistance R of sample is obtained by the Analysis of Equivalent Circuit of described electrode _ets, ask for R _etswith R _etcdifference △ R _et, then by △ R _etrelational model with objective microbe concentration, quantitatively detects objective microbe.

The electro transfer resistance R that table 1 obtains for utilizing bioimpedance analysis method of the present invention _etthe electro transfer resistance ZR that value and existing Impedance Analysis software Zsimpwin obtain _etthe comparison of value.The present invention is analyzed the experimental data that described electrode gathers at bio-modification and objective microbe testing process (albumin A is fixed, antibody coupling, bovine serum albumin BSA are closed and object is caught) respectively, and the equivalent electrical circuit that Zsimpwin software application is identical carries out simulation process to experimental data and obtains electro transfer impedance Z R _etthe present invention extracts 15 data from different frequency from experimental data, i.e. 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), again from low, and respectively choose one in high frequency and consist of one group of data, frequency and the real part of often organizing data process according to the method described above, namely utilize Newton-decline method to solve, 5 electro transfer resistance values can be obtained; Finally carry out caving area, get middle 3 values and average as electro transfer resistance R _et.Visible, the result of bioimpedance analysis method of the present invention and Zsimpwin has good consistance.

The results contrast of table 1 bioimpedance analysis of the present invention method and existing impedance analysis software

The problems such as the frequency band existed for existing small-sized impedance detection device is narrow, accuracy of detection is on the low side, function is simpler, the 125kHz low-frequency clock that the present invention adopts ARM9 processor (S3C2440) timer internal to produce, solves the LF-response problem of AD5933 or AD5934.For the electrode polarization caused by DC offset voltage difference in existing impedance detection device and the problem such as impedance measurement is inaccurate, the present invention is by a Hi-pass filter, direct current biasing difference between the input signal of a voltage follower and an I-V impact damper elimination microelectrode module and output signal, , make the DC offset voltage in whole signal chains constant in VDD/2, automatically regulated by Stepwise calibration and range and impedance detection scope is extended to 100 Ω-100k Ω, the resistance of all biasing resistors and the equal service precision 0.1% of feedback resistance is to reduce inaccuracy, effectively improve measuring accuracy, and by adopting equivalent electrical circuit to ask for the very positively related electro transfer resistance with objective microbe concentration, replace the impedance magnitude that forefathers adopt, set up the relational model with objective microbe concentration, carry out the quantitative test of objective microbe.In addition, mostly existing apparatus is to utilize single-chip microcomputer to develop, fairly simple on interpersonal interactive interface, and the present invention adopts ARM9 processor to develop the simpler and more intuitive touch-screen human-computer interaction interface of display of operation.

Last it is noted that above embodiment is only in order to illustrate technical scheme of the present invention, be not intended to limit; Although with reference to previous embodiment to invention has been detailed description, those of ordinary skill in the art is to be understood that: it still can be modified to the technical scheme described in foregoing embodiments, or carries out equivalent replacement to wherein portion of techniques feature; And these amendments or replacement, do not make the essence of appropriate technical solution depart from the spirit and scope of various embodiments of the present invention technical scheme.

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.