Bacterial endotoxin detection analyzer

Abstract

The invention discloses a bacterial endotoxin detection analyzer, and particularly relates to the technical field of biological detection, and the bacterial endotoxin detection analyzer comprises a detection instrument, an MCU microcontroller, a color development detection module, a position sensing module, a motor driving module, an automatic temperature control module, a data transmission and processing system and upper computer software. The MCU coordinates the operation of each module, the color development detection module is combined with an electrochemical impedance spectroscopy and an aptamer biosensor to quantify the endotoxin concentration, and the other modules are respectively responsible for sample frame positioning, sample positioning, temperature control, data interaction, human-computer interaction and the like. The technical effects and advantages are remarkable: various stable voltages can be efficiently converted, and the system stability is improved; high-sensitivity and high-specificity signal acquisition and concentration quantification are realized, and the detection accuracy is enhanced; signal jitter is eliminated, motor out-of-step stalling is prevented, and reliability is improved; the temperature fluctuation is maintained between-0.2 DEG C and + 0.2 DEG C, so that the biochemical reaction is stable and repeatable.

Description

Technical Field

[0001] This invention relates to the field of biological detection technology, and more specifically, to a bacterial endotoxin detection and analysis instrument. Background Technology

[0002] Bacterial endotoxins are lipopolysaccharide and protein complexes found in the cell walls of Gram-negative bacteria, with lipid A being the main toxic component. Endotoxins are released upon bacterial death or autolysis. Large amounts of endotoxins entering the bloodstream can cause fever, microcirculatory disturbances, endotoxic shock, and disseminated intravascular coagulation, potentially leading to multiple organ failure and septic shock, with a high mortality rate. Endotoxins are closely associated with various infectious diseases; disease progression is often accompanied by an increase in endotoxin levels, while disease remission is often accompanied by a decrease in endotoxin levels. Therefore, rapid detection of endotoxin levels in blood and organs can provide a reference for the diagnosis and prognosis of related clinical diseases.

[0003] While existing Gram-negative bacterial endotoxin detection technologies have broad application prospects, they suffer from the following core defects and problems: existing detection instruments often rely on simple voltage regulation circuits, which cannot efficiently convert between various working voltages; traditional colorimetric detection methods are easily affected by environmental noise and lack dynamic correction mechanisms, resulting in inaccurate impedance signal acquisition and low sensitivity; existing position detection modules often use a single sensor, which is prone to signal jitter and has weak noise resistance, causing sample rack positioning errors and affecting the accuracy and efficiency of automated sample processing.

[0004] Therefore, a bacterial endotoxin detection and analysis instrument is proposed to address the above problems. Summary of the Invention

[0005] In order to overcome the above-mentioned defects of the prior art, the present invention provides a bacterial endotoxin detection and analysis instrument to solve the problems mentioned in the background art.

[0006] To achieve the above objectives, the present invention provides the following technical solution: a bacterial endotoxin detection and analysis instrument, comprising a detection instrument body, characterized in that: the detection instrument consists of an MCU microcontroller, a colorimetric detection module, a position sensing module, a motor drive module, an automatic temperature control module, a data transmission and processing system, and host computer software.

[0007] The MCU (Microcontroller Unit) serves as the core control unit of the testing instrument, responsible for coordinating the operation of various functional modules.

[0008] The colorimetric detection module uses electrochemical impedance spectroscopy combined with an aptamer biosensor to quantify endotoxin concentration by measuring impedance changes.

[0009] The position sensing module is used to accurately determine the movement position of the sample holder, ensuring the accuracy of the detection process;

[0010] The motor driving module controls the stepping motor to realize accurate positioning of the sample and prevent the phenomenon of losing step and locking.

[0011] The automatic temperature control module realizes local accurate temperature control through a distributed micro temperature control unit.

[0012] The data transmission and processing system realizes reliable data interaction between the upper computer and the lower computer through serial communication.

[0013] The upper computer software provides a human-computer interaction interface and supports data storage, analysis and report generation.

[0014] Preferably, the MCU microcontroller integrates a voltage conversion unit and a multi-stage voltage stabilization circuit, which is used to convert the externally input 12V DC power into stable working voltages required by each functional module of the detection instrument. The multi-stage voltage stabilization circuit includes a Buck-Boost topology structure and a linear voltage stabilizer.

[0015] Preferably, the multi-stage voltage stabilization circuit can convert the input 12V voltage into a series of stable voltages with different amplitudes, including +12V, -12V, +10V, -10V, 5V and 3.3V, for providing stable and accurate voltages to each module in the detection instrument.

[0016] Preferably, the color development detection module includes a detection unit and a signal acquisition unit. The detection unit uses aptamer probe sequence-specific capture of bacterial endotoxin to trigger a color development reaction, and the microfluidic chip array is fixed on the surface. The inside of the microfluidic chip array is integrated with a nano-porous gold electrode to generate a measurable impedance signal in response to changes in charge distribution.

[0017] Preferably, the signal acquisition unit includes a lock-in amplifier circuit and a dynamic baseline correction algorithm. The lock-in amplifier circuit converts the impedance signal into a digital signal and inputs it into the dynamic baseline correction algorithm. The expression of the dynamic baseline correction algorithm is as follows:

[0018]

[0019] wherein, represents the corrected impedance change amount; represents the original impedance value at time point t; represents the size of the sliding window, with a value range of 10-100; represents the sampling time interval, with a value range of 0.1-1ms; represents the dynamic noise baseline. The signal acquisition unit is connected to the detection unit, real-time collects the electrochemical impedance change signal generated by the color development reaction, and converts it into endotoxin concentration data. The MCU microcontroller calculates the quantitative relationship between the impedance change amount and the endotoxin concentration in real time.

[0020] Preferably, the position sensing module is composed of multiple sets of position sensors and corresponding circuits, the position sensing module adopts photoelectric or mechanical switch sensors to accurately determine the motion position of the sample holder by outputting high and low level signals; the supporting signal processing circuit includes a voltage conversion chip and a signal conditioning circuit, the voltage conversion chip is used to convert the non-standard level signal output by the position sensor into a stable signal matched with the logic level of the master control unit, and the signal conditioning circuit is used to eliminate signal jitter and improve noise suppression capability, each set of the position sensor is independently configured with a signal processing channel, and multiple sets of the position sensors output level signals to the master control unit through independent signal processing channels to realize multi-position synchronous detection of the sample holder.

[0021] Preferably, the motor driving module is used to receive a single-chip microcomputer GPIO port control signal and output a motor driving current to meet the driving requirements of the stepper motor. In software, the motor accelerates and decelerates from the starting speed to the target speed at a constant acceleration, which can prevent the motor from losing steps and blocking at start and stop due to out-of-step and overshoot. This module not only provides stable motor driving function, but also has precise motor control capability, ensuring accurate positioning and operation of the sample during detection.

[0022] Preferably, the automatic temperature control module is composed of multiple independent temperature-controlled microfluidic channels, the microfluidic channels integrate thin film platinum resistance temperature sensors and micro thermoelectric elements to realize local temperature accurate regulation, the automatic temperature control module maintains temperature fluctuation within-0.2℃-+0.2℃ based on the data in the microfluidic channels using an adaptive temperature control algorithm, and the expression of the adaptive temperature control algorithm is as follows:

[0023]

[0024] wherein, represents output power; represents current temperature deviation; represents temperature deviation change rate; represents fluid thermal inertia; represents rule consequent function; represents the activation strength of the kth fuzzy rule; represents fuzzy rule activation strength; represents adaptive gain coefficient; represents instantaneous power compensation, and the expression is sampled every 2 seconds to dynamically update coefficient, so as to adapt to reagent switching.

[0025] Preferably, the data transmission and processing system realizes the communication between the upper computer and the lower computer through the serial port RS232 communication protocol, the lower computer sends the data collected by the single-chip microcomputer to the upper computer, the upper computer calls the equivalent circuit fitting algorithm for processing after checking the received impedance data, and the expression of the equivalent circuit fitting algorithm is:

[0026]

[0027] wherein, represents the impedance response function in the complex frequency domain; d represents the zero-frequency resistance component; represents the high-frequency asymptotic slope control term; K represents the number of terms; represents the time constant of the system dynamic characteristic; represents the residual vector, the high-confidence parameter is fixed through the equivalent circuit fitting algorithm, and the remaining parameters are iteratively optimized.

[0028] Preferably, the upper computer software stores and analyzes the impedance data, supports equivalent circuit fitting and concentration calibration curve generation, and generates a concentration calibration curve and a statistical report, and supports data export.

[0029] The technical effects and advantages of the present application are as follows:

[0030] 1. Compared with the prior art, the bacterial endotoxin detection analyzer can convert 12V input into multiple stable working voltages through the voltage conversion unit, ensure that each functional module can reliably operate under accurate voltage, solve the problem of unstable power supply, and improve the overall stability of the system.

[0031] 2. Compared with the prior art, the bacterial endotoxin detection analyzer can use the aptamer probe to specifically capture endotoxin and generate an impedance signal through the colorimetric detection module, simultaneously corrects the background noise in real time, realizes high-sensitivity and high-specificity signal acquisition and concentration quantification, solves the problems of large detection signal noise and insufficient real-time processing, and enhances detection accuracy.

[0032] 3. Compared with the prior art, the bacterial endotoxin detection analyzer can eliminate signal jitter and improve noise suppression capability through the position sensing module, realizes multi-position synchronous detection and accurate positioning of the sample holder, and through the motor driving module, can control the stepper motor at a constant acceleration, prevent the out-of-step and locked-rotor phenomenon in the starting and stopping stages, ensure accurate positioning of the sample and operation continuity, solve the problems of motor control out-of-step and locked-rotor, and improve the reliability of the system.

[0033] 4. Compared with existing technologies, this bacterial endotoxin detection and analysis instrument can dynamically compensate for changes in fluid heat capacity through an automatic temperature control module, maintaining the temperature fluctuation of the microfluidic channel within the range of -0.2℃ to +0.2℃, solving the problem of large temperature control fluctuations, and ensuring the stability and repeatability of biochemical reactions. Attached Figure Description

[0034] Figure 1 This is an overall system block diagram of the present invention.

[0035] Figure 2 This is a schematic diagram of the outer casing of the present invention.

[0036] Figure 3 This is a diagram of the internal structure of the present invention.

[0037] Figure 4 This is a structural diagram of the sample holder moving device of the present invention.

[0038] Figure 5 This is a schematic diagram of the overall circuit board of the present invention.

[0039] Figure 6 This is a flowchart of the display detection process of the present invention.

[0040] Figure 7 This is a flowchart of the lower-level machine software operation of the present invention.

[0041] Figure 8 This is a flowchart of the serial communication process of the present invention.

[0042] The attached figures are labeled as follows: 1. Support frame; 2. Guide rail assembly; 3. Equipment base; 4. Testing instrument; 5. Sample holder; 6. Motor assembly; 7. Position sensor; 11. Microfluidic chip array circuit and interface; 12. Drive interface; 13. Stepper motor drive interface; 14. Power input and filtering circuit; 15. Nanoporous gold electrode interface; 16. Position sensor interface; 17. Data output interface. Detailed Implementation

[0043] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0044] Example 1

[0045] As attached Figures 1 to 8The bacterial endotoxin detection and analysis instrument shown includes a main body of the detection instrument, characterized in that: the detection instrument consists of an MCU microcontroller, a colorimetric detection module, a position sensing module, a motor drive module, an automatic temperature control module, a data transmission and processing system, and host computer software.

[0046] The MCU (Microcontroller Unit) serves as the core control unit of the testing instrument, responsible for coordinating the operation of various functional modules.

[0047] The colorimetric detection module uses electrochemical impedance spectroscopy combined with an aptamer biosensor to quantify endotoxin concentration by measuring impedance changes.

[0048] The position sensing module is used to accurately determine the movement position of the sample holder, ensuring the accuracy of the detection process;

[0049] The motor drive module controls the stepper motor to achieve precise sample positioning and prevent step loss and stalling.

[0050] The automatic temperature control module achieves precise local temperature control through distributed micro temperature control units;

[0051] The data transmission and processing system achieves reliable data interaction between the host computer and the slave computer through serial communication;

[0052] The host computer software provides a human-computer interaction interface and supports data storage, analysis, and report generation.

[0053] Specifically, such as Figure 2 As shown, the instrument casing of this invention is entirely 3D printed from resin material, possessing excellent structural strength and lightweight characteristics, making it suitable for laboratory and field portable operation. The casing has a cuboid structure, with overall dimensions of 25cm (length) × 20cm (width) × 28cm (height), making it compact, space-saving, and easy to deploy in various working environments.

[0054] like Figure 3 As shown, the overall structure of this invention is stable and compact, consisting of multiple core functional components, including a support frame 1, a guide rail assembly 2, an equipment base 3, a testing instrument 4, a sample rack 5, a motor assembly 6, and a position sensor 7. An integrated connection frame is also embedded inside the device, integrating key electronic units such as a power interface, an interaction interface, a control motherboard, a temperature control module, a colorimetric detection module, a signal acquisition module, and sensor modules. This device uses a 12V DC power supply for unified power supply, providing stable energy to all modules of the entire system through the power interface. The interaction interface uses RS232 serial communication to achieve bidirectional communication with the host computer, enabling the issuance of control commands and the uploading of data.

[0055] like Figure 6The diagram shows the overall circuit board of this invention. The circuit board has a compact structure and high functional integration, serving as the core control module for automated detection of bacterial endotoxins. The circuit board includes multiple functional interfaces and drive circuits: a microfluidic chip array circuit and interface 11, a drive interface 12, a stepper motor drive interface 13, a power input and filtering circuit 14, a nanoporous gold electrode interface 15, a position sensor interface 16, and a data output interface 17. The microfluidic chip array drive section uses a PT4115 constant current or DRV8870 constant voltage solution; the stepper motor drive module uses a TMC2660 chip, precisely controlling the movement of the microplate tray via the SPI protocol; the power input terminal is equipped with filtering and isolation circuits to stably distribute 12V power to each functional area; and a position sensor interface and a data output interface are also provided for real-time status feedback and communication with the host computer.

[0056] Based on Example 1, the solution in Example 1 will be further described in detail below with reference to the specific working method, such as... Figures 1 to 8 As shown below, see details:

[0057] In a preferred embodiment, the MCU microcontroller integrates a voltage conversion unit and a multi-stage voltage regulator circuit to convert the externally input 12V DC power supply into a stable operating voltage required by the various functional modules of the testing instrument. The multi-stage voltage regulator circuit includes a Buck-Boost topology and a linear regulator. The multi-stage voltage regulator circuit can convert the input 12V voltage into a series of stable voltages of different amplitudes, including +12V, -12V, +10V, -10V, 5V, and 3.3V, to provide stable and accurate voltages to the various modules in the testing instrument.

[0058] Specifically, the MCU microcontroller, as the core control unit of the testing instrument, is responsible for coordinating the operation of various functional modules. The control motherboard is the core unit of the entire system, including a power interface, an RS232 serial communication interface, a signal acquisition interface, a microcontroller, a light source controller, a motor controller, and a voltage conversion module. The microcontroller uses an STM32F103VET6 chip, based on an ARM Cortex-M3 core, with a main frequency of 72MHz. It has abundant GPIO, ADC, USART, PWM, and other interface resources, offering powerful performance and stable reliability. Through the GPIO ports, the microcontroller can read signals from various sensors and coordinate the operation of actuators such as the light source and stepper motor, completing various operation commands issued by the host computer. Furthermore, the control motherboard adopts a modular design with a clear software architecture, separating the underlying drivers from the upper-level communication protocols, facilitating maintenance and expansion. The overall design, while ensuring efficient data acquisition and precise control, provides stable, flexible, and highly integrated hardware support for the entire testing system.

[0059] The voltage conversion module boasts high adaptability and stability. Through a precise voltage regulation circuit design, it can convert the input 12V voltage into a series of stable voltages of different amplitudes, including but not limited to +12V, -12V, +10V, -10V, 5V, and 3.3V. This multi-stage voltage output design not only ensures that each module within the detector receives the precise voltage required by it, but also enhances the compatibility and reliability of the entire system. To adapt to different operating environments and power supply conditions, the voltage conversion module also features a wide input voltage range, maintaining stable output even under conditions of significant power fluctuations, ensuring the normal operation of the detector.

[0060] In a preferred embodiment, the colorimetric detection module includes a detection unit and a signal acquisition unit. The detection unit uses aptamer probe sequences fixed on the surface of a microfluidic chip array to specifically capture bacterial endotoxins and trigger a colorimetric reaction. The microfluidic chip array integrates nanoporous gold electrodes that generate measurable impedance signals in response to changes in charge distribution. The signal acquisition unit includes a lock-in amplifier circuit and a dynamic baseline correction algorithm. The lock-in amplifier circuit converts the impedance signal into a digital signal and then inputs it into the dynamic baseline correction algorithm. The expression of the dynamic baseline correction algorithm is as follows:

[0061]

[0062] in, This indicates the amount of impedance change after correction; This represents the initial impedance value at time point t; This indicates the size of the sliding window, with a value ranging from 10 to 100. This indicates the sampling time interval, with a value ranging from 0.1 to 1 ms. The signal acquisition unit is connected to the detection unit to represent the dynamic noise baseline. It acquires the electrochemical impedance change signal generated by the colorimetric reaction in real time and converts it into endotoxin concentration data. The MCU microcontroller calculates the quantitative relationship between the impedance change and the endotoxin concentration in real time.

[0063] Specifically, the detection unit uses a chip array constructed using standard microfluidic fabrication technology. Specific aptamer probe sequences are fixed to its surface by chemical bonding. When Gram-negative bacterial endotoxins in the sample are captured, a colorimetric reaction is triggered.

[0064] This microfluidic chip integrates nanoporous gold electrodes. The porous structure increases the surface area, and the chip generates a measurable impedance signal in real time by responding to changes in charge distribution through electrochemical impedance spectroscopy. This converts the biological reaction into an electrical signal. The signal acquisition unit is designed as a lock-in amplifier circuit, based on the principle of phase-sensitive detection. This circuit amplifies the impedance signal and filters out high-frequency noise, then converts the analog signal into a digital signal. The digital signal is input to a dynamic baseline correction algorithm. This algorithm uses a sliding window mechanism, with the window size N adjustable from 10 to 100. The dynamic noise baseline is calculated in real time on the MCU microcontroller, and the original impedance value is corrected through an expression to eliminate the influence of environmental drift. The principle behind this design is to ensure that the impedance change reflects only the endotoxin reaction, rather than background interference.

[0065] The MCU microcontroller performs quantitative relationship calculations in real time based on the corrected impedance change. The impedance change is mapped to endotoxin concentration data through a preset calibration curve. This method significantly improves detection sensitivity and specificity. The specific capture of the aptamer probe reduces false positives, the nanoporous electrode enhances the signal response, and lock-in amplification and dynamic baseline correction jointly suppress noise, achieving high-precision real-time monitoring.

[0066] In a preferred embodiment, the position sensing module consists of multiple sets of position sensors and corresponding circuits. The position sensing module uses photoelectric or mechanical switch sensors to accurately determine the movement position of the sample rack by outputting high and low level signals. The supporting signal processing circuit includes a voltage conversion chip and a signal conditioning circuit. The voltage conversion chip is used to convert the non-standard level signals output by the position sensors into stable signals that match the logic level of the main control unit. The signal conditioning circuit is used to eliminate signal jitter and improve noise suppression capability. Each set of position sensors is independently configured with a signal processing channel. Multiple sets of position sensors output level signals to the main control unit through independent signal processing channels to realize synchronous detection of multiple positions of the sample rack.

[0067] Specifically, the position sensing module includes position sensors for aligning with the sample holder. The position sensors accurately determine the movement position of the sample holder by outputting high and low level signals. When the sample holder reaches the corresponding position, the position sensor sends a high-level signal back to the MCU, and the stepper motor stops rotating to facilitate subsequent detection operations. Each set of position sensors is independently configured with a signal processing channel, which outputs a level signal to the main control unit, enabling synchronous detection of multiple positions on the sample holder.

[0068] In a preferred embodiment, the motor drive module receives control signals from the microcontroller's GPIO port and outputs motor drive current to meet the driving requirements of the stepper motor. By incorporating a trapezoidal acceleration / deceleration algorithm in the software, the motor accelerates and decelerates from its starting speed to the target speed with a constant acceleration, preventing step loss and stalling due to overshoot during startup and shutdown. This module not only provides stable motor drive functionality but also possesses precise motor control capabilities, ensuring accurate positioning and operation of the sample during the testing process.

[0069] Specifically, such as Figure 4 As shown, the sample holder moving device is the key execution unit for moving the sample holder during the detection process. Moving the sample holder aligns different sample areas with the detection unit. This structure mainly includes a stepper motor, coupling, transmission mechanism, slide assembly, guide rail, and fixed bracket. The stepper motor is controlled by a microstepping high-precision driver, and its output shaft is connected to the transmission assembly via a coupling. The coupling effectively compensates for axial and radial errors, reduces vibration, and improves system stability. The stepper motor is mounted on the side fixed bracket, driving the ball screw to rotate. The screw meshes with the slider, enabling the sample holder to move smoothly along the linear guide rail. Both the guide rail and slider are selected from high-rigidity, low-friction models, possessing advantages such as low noise, wear resistance, and high load-bearing capacity, making them suitable for long-term continuous operation. In terms of software, a trapezoidal acceleration / deceleration algorithm is used to accelerate and decelerate the motor from the starting speed to the target speed with a constant acceleration, preventing motor step loss and stalling due to step loss and overshoot during startup and shutdown.

[0070] In a preferred embodiment, the automatic temperature control module comprises multiple independently temperature-controlled microfluidic channels. Each microfluidic channel integrates a thin-film platinum resistance temperature sensor and a miniature thermopile to achieve precise local temperature control. Based on data from the microfluidic channels, the automatic temperature control module employs an adaptive temperature control algorithm to maintain temperature fluctuations between -0.2℃ and +0.2℃. The expression for the adaptive temperature control algorithm is as follows:

[0071]

[0072] in, Indicates output power; Indicates the current temperature deviation; Indicates the rate of change of temperature deviation; Indicates the thermal inertia of a fluid; Represents the consequent function of the rule; This represents the activation strength of the k-th fuzzy rule; Indicates the activation strength of the fuzzy rule; Indicates the adaptive gain coefficient; This represents instantaneous power compensation, measured every 2 seconds using the expression. Dynamic updates The coefficient is used to adapt to reagent switching.

[0073] Specifically, each channel is precision-machined from a chemically resistant transparent polymer, with a thin-film platinum resistance temperature sensor and a miniature thermopile integrated into its inner wall. The thin-film platinum resistance is directly deposited on the inner surface of the microchannel using a sputtering process, sensing fluid temperature changes in real time and feeding the data back to the MCU microcontroller. The miniature thermopile achieves precise heating or cooling in localized areas of the channel through the thermal effect of semiconductor materials. This module regulates the temperature based on an adaptive temperature control algorithm. First, the thin-film platinum resistance monitors the current temperature deviation and the rate of change of the deviation in real time, serving as the antecedent input for the fuzzy rules. Simultaneously, the fluid thermal inertia parameters are sampled every 2 seconds to dynamically update the thermal capacity coefficient in the consequent function of the rules. The algorithm's output power consists of two parts: the decoupling enhancement output term calculates the steady-state power based on the activation intensity of the fuzzy rules, and the fluid thermal capacity dynamic compensation term corrects the instantaneous power compensation required for changes in thermal inertia caused by reagent switching in real time through partial differential operations, thereby quickly responding to sudden changes in fluid thermal characteristics.

[0074] The MCU precisely drives the micro thermopile based on power calculations, forming a closed-loop temperature control system within the microfluidic channels. This ensures that each channel independently maintains its target temperature with fluctuations strictly controlled between -0.2℃ and +0.2℃. This design eliminates the hysteresis error of traditional external temperature measurement through in-situ integrated sensors. The adaptive algorithm compensates for the thermal inertia of the fluid in real time, overcoming the temperature instability caused by reagent changes. Independent temperature control of multiple channels avoids cross-interference, ultimately providing a highly stable temperature environment for the colorimetric reaction and significantly improving detection repeatability and accuracy.

[0075] In a preferred embodiment, the data transmission and processing system uses the RS232 serial communication protocol to realize communication between the host computer and the slave computer. The slave computer sends data collected by the microcontroller to the host computer. After verifying the received impedance data, the host computer calls an equivalent circuit fitting algorithm for processing. The expression of the equivalent circuit fitting algorithm is:

[0076]

[0077] in, d represents the impedance response function in the complex frequency domain; d represents the zero-frequency resistance component. This represents the high-frequency asymptotic slope control term; K represents the number of terms. The time constant represents the dynamic characteristics of the system. The residual vector is represented by the equivalent circuit fitting algorithm, which fixes the high-confidence parameters and iteratively optimizes the remaining parameters.

[0078] Specifically, the system uses a four-wire RS232 serial communication protocol to connect the lower-level machine and the upper-level machine. The lower-level machine sends the acquired raw impedance data packets in real time through the USART interface of the STM32 microcontroller. Each data packet contains a timestamp, channel identifier, and impedance amplitude and phase information. After receiving the data, the upper-level machine first performs CRC check and frame integrity check. If the check fails, it requests retransmission to ensure data reliability.

[0079] The impedance data that has passed verification is input into an equivalent circuit fitting algorithm for processing. The innovative implementation principle of this algorithm is—based on the complex frequency domain impedance response function model, high-confidence parameters such as the zero-frequency resistance component d and the high-frequency asymptotic slope control term e are fixed first. These parameters are predetermined during the system calibration stage; subsequently, the residual vector is processed... and system dynamic characteristics time constant Iterative optimization is performed, and the error between the measured impedance and the model fitting is minimized through a nonlinear optimization algorithm, thereby accurately analyzing the impedance change characteristics caused by the endotoxin reaction.

[0080] In a preferred embodiment, the host computer software stores and analyzes impedance data, supports equivalent circuit fitting and concentration calibration curve generation, generates concentration calibration curves and statistical reports, and supports data export.

[0081] Specifically, the host computer software provides a user-friendly interface that greatly simplifies the operation process. Users can easily set parameters through an intuitive graphical interface, such as setting the temperature, selecting the detection area, and configuring the detection mode. These settings can be completed with simple clicks and drags, without requiring complex steps or in-depth professional knowledge.

[0082] Furthermore, the software provides a real-time feedback mechanism, allowing users to monitor the progress and status of experiments at any time, ensuring that the experiments are executed precisely according to the preset parameters. After the experiment, the software can automatically collect and organize the data, providing users with a detailed experimental report, including charts, statistical analysis, and conclusions and suggestions. The host computer software is not only responsible for storing and analyzing the acquired raw signals, but also integrates various data processing algorithms, such as data filtering, baseline calibration, curve fitting, and noise suppression, to ensure the accuracy, stability, and repeatability of the output results.

[0083] The working process of this invention is as follows: First, the user starts the detection program through the host computer software. The MCU microcontroller then activates the integrated power system to convert the external 12V input into a multi-level stable voltage of ±12V, ±10V, 5V and 3.3V for the use of the entire system. Then, the motor drive module executes the trapezoidal acceleration and deceleration algorithm to drive the sample rack to move. At the same time, the position sensing module feeds back the position level signal of the sample rack to the MCU in real time through multiple independent signal processing channels.

[0084] When the sample reaches the detection position, the automatic temperature control module immediately starts the adaptive temperature control algorithm, which regulates the temperature of the microfluidic channel by decoupling and strengthening the output and the fluid heat capacity dynamic compensation term, and dynamically updates the fluid thermal inertia parameters every 2 seconds to maintain a constant temperature of ±0.2℃; then the aptamer probe of the colorimetric detection module specifically captures the endotoxin to trigger the colorimetric reaction, and the integrated nanoporous gold electrode responds to the charge change to generate the original impedance signal.

[0085] Next, the signal acquisition unit converts the signal through a lock-in amplifier and executes a dynamic baseline correction algorithm, outputting noise-reduced impedance in real time with a 10-100 point sliding window and a 0.1-1ms interval. Then, the MCU uploads the data to the host computer via RS232 protocol, and the host computer calls the equivalent circuit fitting algorithm to analyze the impedance characteristics. Finally, the endotoxin concentration calibration curve and statistical report are automatically generated, and the data is exported. The above is the working principle of this bacterial endotoxin detection and analysis instrument.

Claims

1. A bacterial endotoxin detection and analysis instrument, comprising a detection instrument, characterized in that: The detection instrument consists of an MCU microcontroller, a colorimetric detection module, a position sensing module, a motor drive module, an automatic temperature control module, a data transmission and processing system, and host computer software. The MCU (Microcontroller Unit) serves as the core control unit of the testing instrument, responsible for coordinating the operation of various functional modules. The colorimetric detection module uses electrochemical impedance spectroscopy combined with an aptamer biosensor to quantify endotoxin concentration by measuring impedance changes. The position sensing module is used to accurately determine the movement position of the sample holder, ensuring the accuracy of the detection process; The motor drive module controls the stepper motor to achieve precise sample positioning and prevent step loss and stalling. The automatic temperature control module achieves precise local temperature control through distributed micro temperature control units; The data transmission and processing system achieves reliable data interaction between the host computer and the slave computer through serial communication; The host computer software provides a human-computer interaction interface and supports data storage, analysis, and report generation.

2. The bacterial endotoxin detection and analysis instrument according to claim 1, characterized in that: The MCU microcontroller integrates a voltage conversion unit and a multi-stage voltage regulator circuit to convert the externally input 12V DC power supply into a stable operating voltage required by the various functional modules of the testing instrument. The multi-stage voltage regulator circuit includes a Buck-Boost topology and a linear regulator.

3. The bacterial endotoxin detection and analysis instrument according to claim 2, characterized in that: The multi-stage voltage regulator circuit can convert the input 12V voltage into a series of stable voltages of different amplitudes, including +12V, -12V, +10V, -10V, 5V and 3.3V, to provide stable and accurate voltages to the various modules in the testing instrument.

4. The bacterial endotoxin detection and analysis instrument according to claim 1, characterized in that: The colorimetric detection module includes a detection unit and a signal acquisition unit. The detection unit uses aptamer probe sequences fixed on the surface of a microfluidic chip array to specifically capture bacterial endotoxins and trigger a colorimetric reaction. The microfluidic chip array integrates nanoporous gold electrodes that generate measurable impedance signals in response to changes in charge distribution.

5. The bacterial endotoxin detection and analysis instrument according to claim 4, characterized in that: The signal acquisition unit includes a lock-in amplifier circuit and a dynamic baseline correction algorithm. The lock-in amplifier circuit converts the impedance signal into a digital signal and then inputs it into the dynamic baseline correction algorithm. The expression of the dynamic baseline correction algorithm is as follows: ， in, This indicates the amount of impedance change after correction; This represents the initial impedance value at time point t; This indicates the size of the sliding window, with a value ranging from 10 to 100. This indicates the sampling time interval, with a value ranging from 0.1 to 1 ms. The signal acquisition unit is connected to the detection unit to represent the dynamic noise baseline. It acquires the electrochemical impedance change signal generated by the colorimetric reaction in real time and converts it into endotoxin concentration data. The MCU microcontroller calculates the quantitative relationship between the impedance change and the endotoxin concentration in real time.

6. The bacterial endotoxin detection and analysis instrument according to claim 1, characterized in that: The position sensing module consists of multiple sets of position sensors and corresponding circuits. The position sensing module uses photoelectric or mechanical switch sensors to accurately determine the movement position of the sample rack by outputting high and low level signals. The supporting signal processing circuit includes a voltage conversion chip and a signal conditioning circuit. The voltage conversion chip is used to convert the non-standard level signals output by the position sensors into stable signals that match the logic level of the main control unit. The signal conditioning circuit is used to eliminate signal jitter and improve noise suppression capability. Each set of position sensors is independently configured with a signal processing channel. Multiple sets of position sensors output level signals to the main control unit through independent signal processing channels to realize synchronous detection of multiple positions of the sample rack.

7. The bacterial endotoxin detection and analysis instrument according to claim 1, characterized in that: The motor drive module is used to receive control signals from the GPIO port of the microcontroller and output motor drive current to meet the driving requirements of the stepper motor. In terms of software, it combines a trapezoidal acceleration and deceleration algorithm so that the motor accelerates and decelerates from the starting speed to the target speed with a constant acceleration.

8. The bacterial endotoxin detection and analysis instrument according to claim 1, characterized in that: The automatic temperature control module consists of multiple independently temperature-controlled microfluidic channels. Each microfluidic channel integrates a thin-film platinum resistance temperature sensor and a miniature thermopile to achieve precise local temperature regulation. Based on data from the microfluidic channels, the automatic temperature control module employs an adaptive temperature control algorithm to maintain temperature fluctuations between -0.2℃ and +0.2℃. The expression for the adaptive temperature control algorithm is as follows: ， in, Indicates output power; Indicates the current temperature deviation; Indicates the rate of change of temperature deviation; Indicates the thermal inertia of a fluid; Represents the consequent function of the rule; This represents the activation strength of the k-th fuzzy rule; Indicates the activation strength of the fuzzy rule; Indicates the adaptive gain coefficient; This represents instantaneous power compensation, measured every 2 seconds using the expression. Dynamic updates The coefficient is used to adapt to reagent switching.

9. The bacterial endotoxin detection and analysis instrument according to claim 1, characterized in that: The data transmission and processing system uses the RS232 serial communication protocol to achieve communication between the host computer and the slave computer. The slave computer sends data collected by the microcontroller to the host computer. After verifying the received impedance data, the host computer calls the equivalent circuit fitting algorithm for processing. The expression of the equivalent circuit fitting algorithm is: ， in, d represents the impedance response function in the complex frequency domain; d represents the zero-frequency resistance component. This represents the high-frequency asymptotic slope control term; K represents the number of terms. The time constant represents the dynamic characteristics of the system. The residual vector is represented by the equivalent circuit fitting algorithm, which fixes the high-confidence parameters and iteratively optimizes the remaining parameters.

10. A bacterial endotoxin detection and analysis instrument according to claim 1, characterized in that: The host computer software stores and analyzes impedance data, supports equivalent circuit fitting and concentration calibration curve generation, generates concentration calibration curves and statistical reports, and supports data export.

Images