CN106190814B - A kind of cardiac muscle cell manipulator - Google Patents

Abstract

The invention relates to a cardiomyocyte manipulator, which includes a piezoelectric ceramic controller, a precision motor controller, a precision motor, a piezoelectric ceramic group, an operating terminal, a display, a cell culture dish, an inverted microscope, a microelectrode array, a wire, and a phase-locked Amplifier, platform and main controller, the movement of the precision motor is controlled by the precision motor controller; the movement of the piezoelectric ceramic group is controlled by the piezoelectric ceramic controller; the operating end is located at the end position of the piezoelectric ceramic group; the display can simultaneously display the inverted microscope Take images; an inverted microscope is used to obtain the physical beating image information of the cells in the cell culture dish; the microelectrode array is located in the cell culture dish; the main controller is connected to control the precision motor controller, piezoelectric ceramic controller, lock-in amplifier, display and Inverted microscope. The operation end can vibrate synchronously with the cardiomyocytes, that is, keep the operation end and the surface of the cardiomyocytes relatively still, and reduce the influence caused by the beating of the cardiomyocytes.

Description

A cardiomyocyte manipulator

technical field

The invention relates to a cardiomyocyte manipulator, in particular to a manipulator that vibrates synchronously by performing algorithmic prediction based on detection of cardiomyocyte beating images and bioelectrical signals.

Background technique

Cardiomyocytes, also known as myocardial fibers, have striated involuntary muscles and have the ability to excite and contract. The primary culture of cardiomyocytes was cultured in vitro by tissue block method, enzyme digestion method and isolated heart perfusion method. During in vitro culture, cardiomyocytes often exhibit spontaneous beating, and the beating of cardiomyocytes in the same cell culture dish is often synchronized. When cardiomyocytes are beating, the beating frequency is generally not constant. In different environments, the beating frequency often changes, resulting in arrhythmia.

Currently, the manipulation of beating cells such as cardiomyocytes still relies on manual manipulation. Due to the pulsation phenomenon, the tip often causes a lot of scratches to the cells when manually touching the cells. When performing operations such as stripping, injecting, and transferring cardiomyocytes, damage to the cells is often caused due to the influence of the pulsation of the cardiomyocytes, and the efficiency is not high. Especially during injection, friction between the cell surface and the manipulation tip affects manipulation precision and cell viability. Relying on the human eye to judge the beating frequency and amplitude of cardiomyocytes is time-consuming and labor-intensive with low accuracy. In order to improve the accuracy and efficiency of the operation and avoid the influence caused by the pulsation and arrhythmia of the cardiomyocytes, an operator for operating the active cardiomyocytes is needed.

Contents of the invention

The purpose of the present invention is to provide a cardiomyocyte manipulator, realize the prediction of the beating frequency of cardiomyocytes, control the vibration of the manipulator at the same frequency, keep the end of the manipulator and the surface of the cardiomyocytes relatively static, and then move the manipulator on the basis of vibration , to achieve injection, stripping, transfer and other operations on cardiomyocytes.

In order to achieve the above object, the design idea of the present invention is:

The cardiomyocytes in the cell culture dish will beat autonomously and release weak bioelectrical signals at the same time. According to the principle of cardiomyocyte pulsation in physiology, after an action potential occurs in cardiomyocytes, the calcium ion channel opens, causing the movement of muscle fibers and forming the pulsation phenomenon of cardiomyocytes. The bioelectrical signal corresponds to the pulsation signal, that is, a bioelectrical signal is generated corresponding to a physical pulsation. First, move the operating end near the surface of the cardiomyocyte to be operated, adjust the vertical pointing to the edge of the cardiomyocyte, use an inverted microscope to collect the physical beating image of the cardiomyocyte to be operated, and obtain the physical beating frequency and amplitude of the cell edge contour on the focal plane. According to the collected physical pulsation signal, using algorithms such as RBF neural network, the predicted edge contour pulsation signal can be obtained on the basis of the known pulsation signal. Activating the piezo controller drives the piezo stack to vibrate so that the operating tip vibrates at a predicted frequency and amplitude. In this way, the end of the operation can vibrate at a certain frequency and amplitude, and maintain the maximum synchronization with the beating of the cardiomyocytes, so as to achieve the purpose of being relatively static, which is beneficial to the operation of stripping, injecting, and transferring the cardiomyocytes, and reduces the damage caused by the beating of the cardiomyocytes. Influenced by the measured bioelectrical signal, a calibration detection can be performed on the predicted beating signal.

The physical beating can be observed through an inverted microscope, and the cardiomyocyte contour image processing can be obtained. Bioelectrical signals can be detected with dedicated instrument circuits, such as lock-in amplifiers, Huygens bridges, etc. At the same time, the inverted microscope can calibrate the driving components such as piezoelectric ceramic group and precision motor, so as to improve the control accuracy of the operator's end.

According to above-mentioned inventive concept, the present invention adopts following technical scheme:

A cardiomyocyte manipulator, including a piezoelectric ceramic controller, a precision motor controller, a precision motor, a piezoelectric ceramic group, an operating terminal, a display, a cell culture dish, an inverted microscope, a microelectrode array, wires, a lock-in amplifier, and a platform and the main controller, the piezoelectric ceramic controller and the precision motor controller are installed on the platform, and the movement of the precision motor is controlled by the precision motor controller to realize the rapid movement of the operator; the piezoelectric ceramic group is installed on The precision motor is controlled by the piezoelectric ceramic controller to realize precise movement in three directions; the operating end is located at the end of the piezoelectric ceramic group, bent at a certain angle, and the cantilever is horizontal after installation; the display is installed on the On the platform, the captured image of the inverted microscope is displayed synchronously, which is convenient for the operator to observe; the inverted microscope is installed on the platform, and the platform has a transparent glass window at the position above the inverted microscope, and the bottom of the cell culture dish is transparent, placed on the platform on the transparent glass window; the microelectrode array is located in the cell culture dish, one end of the wire is connected to the microelectrode array, and the other end is connected to a lock-in amplifier; the main controller is respectively connected to a piezoelectric ceramic controller, a precision motor controller, a display , an inverted microscope and a lock-in amplifier.

In the main controller, process the collected image information, predict the beating frequency and amplitude of the cardiomyocytes, control the vibration frequency and amplitude of the piezoelectric ceramic group, and keep the operation terminal and the cardiomyocytes in synchronous vibration.

In the main controller, process and collect the frequency of bioelectrical signals, and predict the beating frequency of cardiomyocytes, obtain the predicted beating frequency calculated from the bioelectrical signal, and then correct the wrong frequency predicted by image processing, so as to ensure the prediction Beat rate accuracy.

Compared with the prior art, the present invention has the following obvious outstanding substantive features and significant technological progress:

The manipulator relies on the collected image information, uses algorithms such as RBF neural network to predict the beating frequency and amplitude of myocardial cells, controls the vibration frequency and amplitude of the piezoelectric ceramic group, and can keep the operation end and the myocardial cells in synchronous vibration, that is, maintain the operation end and the myocardium. The cell surface is relatively static, and has the advantages of high precision, compact structure, convenient operation, and little damage to cardiomyocytes.

Description of drawings

Fig. 1 is a schematic diagram of the composition of the system of the present invention.

Fig. 2 is a schematic diagram of the internal structure of the culture dish of the present invention.

Fig. 3 is a schematic diagram of beat signal prediction in the present invention.

Detailed ways

The present invention will be described in further detail below in conjunction with the accompanying drawings and preferred embodiments.

As shown in Figures 1 and 2, a cardiomyocyte manipulator includes a piezoelectric ceramic controller 1, a precision motor controller 2, a precision motor 3, a piezoelectric ceramic group 4, an operating terminal 5, a display 6, and a cell culture dish 7, inverted microscope 8, microelectrode array 9, wire 10, lock-in amplifier 11, platform 12 and master controller 13, described piezoelectric ceramic controller 1 and precision motor controller 2 are installed on the platform 12, and described precision The movement of the motor 3 is controlled by the precision motor controller 2 to realize the rapid movement of the operator; the piezoelectric ceramic group 4 is installed on the precision motor 3 and its movement is controlled by the piezoelectric ceramic controller 1 to realize the movement in three directions. Precise movement; the operating end 5 is located at the end of the piezoelectric ceramic group 4, bent at a certain angle, and the cantilever is horizontal after installation, and can be replaced with other operating equipment such as a syringe; the display 6 is installed on the platform 12, and the display is inverted synchronously The captured images of the microscope 8 are convenient for operators to observe; the inverted microscope 8 is installed on the platform 12, and the platform 12 has a transparent glass window at the position above the inverted microscope 8, and the bottom of the cell culture dish 7 is transparent and placed on the platform 12 on the transparent glass window; the inverted microscope 8 is used to obtain the physical beating image information of the cells in the cell culture dish 7; The lock-in amplifier 11; the main controller 13 is respectively connected to the piezoelectric ceramic controller 1, the precision motor controller 2, the display 6, the inverted microscope 8 and the lock-in amplifier 11.

The use process of the system of the present invention is as follows:

The active isolated cardiomyocytes are placed in the cell culture dish 7, and an appropriate amount of culture solution is added so that the microelectrode array 9 and the cardiomyocytes can be in stable contact.

First perform system calibration: turn on the inverted microscope 8 to observe the operation terminal 5, start the precision motor controller 2 and piezoelectric ceramic controller 1, and quickly move the operation terminal 5 to near the bottom of the cell culture dish 7 through the work of the precision motor 3, and then The piezoelectric ceramic group 4 moves the operating end 5 precisely to the bottom of the cell culture dish 7, and adjusts the inverted microscope 8 to observe the image of the operating end 5 in real time. When the shape of the operating end 5 changes in the field of view, it indicates that the operating end 5. Touch the bottom and bend, then lift the operating end 5, and repeat the process of moving down and touching the bottom. The average value was taken for multiple repetitions, and the height of the bottom 7 of the cell culture dish was recorded. Then, adjust the inverted microscope 8 to observe the shape of the top of the cardiomyocyte, and lift the operating end 5 to touch the top of the cardiomyocyte. When the shape of the top of the cardiomyocyte in the field of view is obviously different from that when it is not touched, it means that the operating end 5 has touched the cardiomyocyte At the top, the average value was repeated several times, and the height of the top of the cardiomyocyte was recorded.

Adjust the focal plane of the inverted microscope 8, find the surface position of the cardiomyocyte to be operated, and record the focal plane position after finding a suitable position. Then move the operation end 5 to the focal plane just now, and wait for the next operation. At this time, the surface on which cardiomyocytes are operated and the operating end 5 are on the same plane.

In this focal plane, use the inverted microscope 8 to acquire the image information of the edge contour of the cardiomyocytes, and in the main controller 13, process the image to obtain the predicted beat signal. As shown in Figure 3, the beating frequency and amplitude fitting process of cardiomyocytes is as follows:

1) Obtain a frame of image;

2) Gaussian filtering is performed on the image to remove image noise points;

3) Use Canny operator for edge detection;

4) Extracting detected edge profile information;

5) Calculate the edge contour information of the current image, including the inner area S of the contour and the length L of the contour;

6) Repeat steps 1)-5) to collect 1000 frames of data information;

7) construct RBF neural network, utilize data S and L to carry out fitting to heartbeat frequency and amplitude;

8) Obtain the predicted cardiomyocyte beating frequency and amplitude.

The predicted cardiomyocyte beating frequency and amplitude are transmitted to the piezoelectric ceramic controller 1, and the operating terminal 5 is controlled to vibrate at the predicted frequency and amplitude according to the geometric positional relationship between the piezoelectric ceramic group 4 and the operating terminal 5.

In this way, the operating end 5 keeps moving synchronously with the cardiomyocytes, that is, keeps relatively still. Minimal damage to cardiomyocytes occurs when manipulations are performed close to the surface of the cardiomyocytes. When performing operations such as injection on cardiomyocytes, it is only necessary to superimpose the required movement on the basis of the vibration of the operation end 5 .

The microelectrode array 9 in the cell culture vessel 7 can detect the bioelectrical signal of cardiomyocytes, which is transmitted to the lock-in amplifier 11 through the wire 10, thereby detecting the weak bioelectrical signal, which is amplified and transmitted to the main controller 13. In the main controller 13, for the frequency of the bioelectric signal, the RBF neural network algorithm is used to predict the beating frequency of cardiomyocytes, and the predicted beating frequency calculated from the bioelectric signal is obtained, and then the wrong frequency predicted by image processing can be corrected , so as to ensure the accuracy of the predicted beat frequency.

Claims (1)

1. A cardiomyocyte manipulator, characterized in that it includes a piezoelectric ceramic controller (1), a precision motor controller (2), a precision motor (3), a piezoelectric ceramic group (4), and an operating terminal (5) , monitor (6), cell culture dish (7), inverted microscope (8), microelectrode array (9), wire (10), lock-in amplifier (11), platform (12) and main controller (13), The piezoelectric ceramic controller (1) and the precision motor controller (2) are installed on the platform (12), and the movement of the precision motor (3) is controlled by the precision motor controller (2), so as to realize the rapid operation of the operator. Movement; the piezoelectric ceramic group (4) is installed on the precision motor (3), and its movement is controlled by the piezoelectric ceramic controller (1) to achieve precise movement in three directions; the operating end (5) is located at The end position of the piezoelectric ceramic group (4) is bent at a certain angle, and the cantilever is horizontal after installation; the display (6) is installed on the platform (12), and simultaneously displays the image taken by the inverted microscope (8), which is convenient for the operator to observe; The inverted microscope (8) is installed on the platform (12), and the platform (12) has a transparent glass window at the position above the inverted microscope (8), and the bottom of the cell culture dish (7) is transparent, placed on the platform ( 12) on the transparent glass window; the microelectrode array (9) is located in the cell culture dish (7), one end of the wire (10) is connected to the microelectrode array (9), and the other end is connected to a lock-in amplifier (11); the The main controller (13) is respectively connected with the piezoelectric ceramic controller (1), the precision motor controller (2), the display (6), the inverted microscope (8) and the lock-in amplifier (11).