Cell impedance measurement device and cell analysis system comprising same

Abstract

The present invention provides a cell impedance measurement device capable of measuring and collecting electrical data on cells and performing real-time cell analysis in a non-destructive manner, and a cell analysis system comprising same.

Description

Cell impedance measuring device and cell analysis system including the same

[0001] The present invention relates to a cell impedance measuring device and a cell analysis system including the same, and more specifically, to a cell impedance measuring device and a cell analysis system including the same capable of measuring and collecting electrical data of cells and performing real-time cell analysis in a non-destructive manner.

[0002] Cell analysis has established itself as an essential element in modern biological and medical research. In particular, accurately analyzing the state of cells—a critical biological process in which cells transform into mature forms with specific functions—plays a vital role in various studies and experiments, including disease diagnosis, the development of treatments, and the evaluation of drug efficacy.

[0003] Conventional cell analysis utilized methods such as microscopic observation and biochemical analysis, while cell differentiation evaluation employed methods including gene expression analysis, protein expression profiling, immunofluorescence, and flow cytometry. However, these methods not only made it difficult to monitor dynamic cellular changes in real time but also had limitations in terms of cell contamination, accuracy, cost, and time.

[0004] Therefore, there is a need to develop new devices and systems capable of monitoring and analyzing the state of cells more accurately and in real time, such as in cell culture, cell differentiation, and drug screening.

[0005] The technical problem that the present invention aims to solve is to provide a cell impedance measuring device capable of accurately measuring the electrical impedance of a cell in a non-destructive manner and generating various impedance data to enable real-time cell analysis.

[0006] Another technical problem that the present invention aims to solve is to provide a cell analysis system capable of accurately and conveniently analyzing the state of a cell in real time using impedance data for a cell generated by a cell impedance measurement device and a cell analysis model.

[0007] The technical problems of the present invention are not limited to those mentioned above, and other unmentioned technical problems will be clearly understood by a person skilled in the art from the description below.

[0008] A cell impedance measuring device according to one embodiment of the present invention for achieving the above technical problem comprises: a cell chip including a well formed with a receiving portion for culturing a cell or processing a cultured cell and a substrate having a plurality of electrodes formed thereon; an impedance measuring module that applies an alternating current signal to the cell chip to measure the electrical impedance of the cell; and a controller that controls the impedance measuring module and generates impedance data for the measured electrical impedance; wherein the cell chip, the impedance measuring module, and the controller are electrically connected to each other.

[0009] The above substrate is divided into a sensing area and a non-sensing area around the sensing area, and the receiving portion of the well can be placed on the sensing area.

[0010] The above electrode includes a common electrode and an operating electrode spaced apart from the common electrode, and the common electrode and the operating electrode may be formed at the lower part of the receiving portion of the well.

[0011] The substrate, the common electrode, and the working electrode may be formed of a transparent material.

[0012] Any one of the following nanomaterials may be deposited on the surface of the common electrode and the working electrode: gold nanoparticles (AuNPs), quantum dots (QDs), reduced graphene oxide (rGO), poly(3,4-ethylenediooxythiophene)-polystyrene sulfonate (PEDOT:PSS), polyaniline (PANI), carbon nanotubes (CNT), and multi-walled carbon nanotubes (MWCNT).

[0013] The above wells may be spaced apart in multiple directions vertically and horizontally on the substrate.

[0014] The impedance measurement module may include: a signal generator that generates the alternating current signal; a digital-to-analog converter (DAC) that converts the alternating current signal generated by the signal generator into an analog input signal and applies it to the plurality of electrodes corresponding to the well; and an analog-to-digital converter (ADC) that receives the analog response signal generated from the plurality of electrodes corresponding to the well, converts it into a digital signal, and transmits it to the controller.

[0015] It may further include a first signal amplifier that amplifies the analog input signal at the output of the digital-to-analog converter (DAC); and a second signal amplifier that amplifies the analog response signal at the output of the analog-to-digital converter (ADC).

[0016] The second signal amplifier may further include a multiplexer that includes a plurality of input channels and one output channel at the front end, and connects any one of the plurality of input channels to the output channel.

[0017] The above controller may include: a control unit that controls the frequency of the alternating current signal and the measurement time of the electrical impedance; and a data generation unit that receives information regarding the measured electrical impedance and generates the impedance data among resistive data, capacitive data, magnitude data, phase data, and combinations thereof.

[0018] The control unit controls the frequency of the AC signal to a single value during the measurement time, and the data generation unit can generate the impedance data for each time period.

[0019] The control unit controls the frequency of the AC signal to a plurality of different values ​​between a minimum frequency and a maximum frequency during the measurement time, and the data generation unit can generate the impedance data by time and / or frequency.

[0020] A cell analysis system according to an embodiment of the present invention for achieving the above technical problem comprises: a cell impedance measuring device; and a processor that analyzes the state of a cell using impedance data among resistive data, capacitive data, magnitude data, phase data, and combinations thereof generated by the cell impedance measuring device.

[0021] The processor may include a data storage unit that stores the impedance data; and a cell analysis unit that analyzes the state of a cell using the impedance data stored in the data storage unit and a cell analysis model.

[0022] The cell analysis system may further include a terminal device that displays the analysis results of the cell state analyzed by the processor.

[0023] According to the cell impedance measuring device of the present invention as described above, it is possible to accurately measure the electrical impedance of a cell in a non-destructive manner and generate various impedance data by controlling the frequency of the alternating current signal and the electrical impedance measurement time.

[0024] In addition, the cell analysis system of the present invention has the effect of accurately and conveniently analyzing the state of cells in real time, such as cell culture status, cell differentiation status, cell viability and toxicity evaluation, and optimal frequency for each cell, by utilizing impedance data generated from a cell impedance measurement device and a cell analysis model.

[0025] The effects of the present invention are not limited to those mentioned above, and other unmentioned effects will be clearly understood by a person skilled in the art from the description below.

[0026] FIG. 1 is a diagram illustrating the configuration of a cell analysis system according to one embodiment of the present invention.

[0027] FIG. 2 is a block diagram showing a cell analysis system according to one embodiment of the present invention.

[0028] FIG. 3 is a block diagram showing a cell impedance measuring device according to one embodiment of the present invention.

[0029] FIG. 4 is a diagram showing an example of a cell chip constituting the cell impedance measuring device of the present invention.

[0030] FIG. 5 is a diagram showing an example of a substrate constituting a cell chip among the cell impedance measuring devices of the present invention.

[0031] FIGS. 6(a) to 6(c) are drawings showing various embodiments of an impedance measurement module constituting a cell impedance measurement device of the present invention.

[0032] FIG. 7 is a flowchart showing the operation of a control unit according to an embodiment of the present invention.

[0033] FIGS. 8(a) to 8(d) are graphs illustrating a model for analyzing the state of a cell during a cell culture process in a processor according to one embodiment of the present invention.

[0034] FIGS. 9(a) and FIGS. 9(b) are graphs illustrating a model for analyzing the survival or lysis status of cells during the processing of cultured cells in a processor according to one embodiment of the present invention.

[0035] FIGS. 10 to 11 are graphs illustrating a model for analyzing the optimal frequency per cell by a processor according to one embodiment of the present invention.

[0036] FIGS. 12 and 13 are graphs illustrating a model for analyzing growth indicators by cell injection concentration by a processor according to one embodiment of the present invention.

[0037] FIGS. 14 and 15 are graphs illustrating a model for analyzing the state of undifferentiated cells or differentiated cells by a processor according to one embodiment of the present invention.

[0038] The advantages and features of the present invention and the methods for achieving them will become clear by referring to the embodiments described below in conjunction with the accompanying drawings. However, the present invention is not limited to the embodiments disclosed below but may be implemented in various different forms. These embodiments are provided merely to ensure that the disclosure of the present invention is complete and to fully inform those skilled in the art of the scope of the invention, and the present invention is defined only by the scope of the claims.

[0039] The terms used in this specification will be briefly explained, and the invention will be described in detail.

[0040] The terms used in this invention have been selected based on currently widely used general terms, taking into account their functions within the invention; however, these terms may vary depending on the intent of those skilled in the art, case law, the emergence of new technologies, etc. Additionally, in specific cases, terms have been arbitrarily selected by the applicant, and in such cases, their meanings will be described in detail in the relevant description of the invention. Therefore, the terms used in this invention should be defined not merely by their names, but based on their meanings and the overall content of the invention.

[0041] Throughout the specification, when a part is described as "comprising" a certain component, this means that, unless specifically stated otherwise, it does not exclude other components but may include additional components. Furthermore, terms such as "part," "module," and "unit" used in the specification refer to a unit that processes at least one function or operation and may be implemented as software, hardware components such as FPGAs or ASICs, or a combination of software and hardware. However, the terms "part," "module," and "unit" are not limited to software or hardware. "Part," "module," and "unit" may be configured to reside in an addressable storage medium or configured to operate one or more processors. Therefore, as an example, terms such as “part,” “module,” and “unit” include components such as software components, object-oriented software components, class components, and task components, as well as processes, functions, attributes, procedures, subroutines, segments of program code, drivers, firmware, microcode, circuits, data, databases, data structures, tables, arrays, and variables.

[0042] Below, embodiments of the present invention are described in detail with reference to the attached drawings so that those skilled in the art can easily implement the invention. Additionally, parts of the drawings that are irrelevant to the description are omitted to clearly explain the invention.

[0043] Terms including ordinal numbers, such as “first,” “second,” etc., may be used to describe various components, but the components are not limited by the terms. The terms are used solely for the purpose of distinguishing one component from another. For example, without departing from the scope of the present invention, the first component may be named the second component, and similarly, the second component may be named the first component. The term “and / or” includes a combination of multiple related items or any one of the multiple related items.

[0044]

[0045] FIG. 1 is a diagram illustrating the configuration of a cell analysis system according to one embodiment of the present invention, and FIG. 2 is a block diagram showing a cell analysis system according to one embodiment of the present invention.

[0046] Referring to FIGS. 1 and 2, a cell analysis system (1) according to one embodiment of the present invention includes a cell impedance measuring device (10) and a processor (20).

[0047] For example, the cell impedance system (1) may be implemented with a cell impedance measuring device (10) and a processor (20) as multiple objects. In this case, some of the series of processes for providing cell analysis may be performed in the cell impedance measuring device (10), and some of the remaining processes may be performed in the processor (20). Additionally, the cell impedance measuring device (10) and the processor (20) may exchange various data through a communication network. The communication network includes both wired and wireless communication networks. Wired communication networks include USC (Universe Serial Bus), PCI (Peripheral Component Interconnect), PCI-express, etc. Wired communication networks include Wireless LAN, Wi-Fi, Bluetooth, Zigbee, WFD (Wi-Fi Direct), UWB (Ultra Wideland), NFC (Near Field Communication), etc., but are not particularly limited as long as they are methods capable of exchanging various data.

[0048] As another example, the cell impedance system (1) can be implemented as a single object equipped with a processor (20) within a cell impedance measuring device (10). In this case, all processes for providing cell analysis can be performed in the cell impedance measuring device (10), and the cell impedance measuring device (10) can perform the role of the cell impedance system (1).

[0049] Below, we will focus on explaining the cell impedance system (1) implemented with multiple objects.

[0050]

[0051] FIG. 3 is a block diagram showing a cell impedance measuring device according to one embodiment of the present invention, FIG. 4 is a diagram showing one embodiment of a cell chip constituting the cell impedance measuring device of the present invention, FIG. 5 is a diagram showing one embodiment of a substrate constituting the cell chip of the cell impedance measuring device of the present invention, FIG. 6(a) to FIG. 6(c) are diagrams showing various embodiments of an impedance measuring module constituting the cell impedance measuring device of the present invention, and FIG. 7 is a flowchart showing the operation of a control unit according to one embodiment of the present invention.

[0052] Referring to FIG. 3, a cell impedance measuring device (10) according to one embodiment of the present invention includes a cell chip (100), an impedance measuring module (200), and a controller (300). At this time, the cell chip (100), the impedance measuring module (200), and the controller (300) are electrically connected to each other, and an alternating current signal is applied during the process of culturing the cell or processing the cultured cell to measure the electrical impedance of the cell and generate various impedance data for the measured electrical impedance.

[0053] Referring to FIG. 4, the cell chip (100) includes a substrate (120) having a well (110) for culturing cells or processing cultured cells and a plurality of electrodes (122, 124) formed thereon. In one example, the well (110) may be placed on the substrate (120), and the well (110) may be fixed by a heat compression method at 60 to 70°C and 30 to 30 psi on the substrate (120) to which a double-sided tape made of polyethylene, silicone, polyurethane, or polyester material is attached. However, it is not limited thereto, and the well (110) may be fixed on the substrate (120) by various methods.

[0054] The well (110) is configured for culturing cells or processing cultured cells, and is preferably made of a transparent biocompatible material. The biocompatible material may be a transparent plastic such as PDMS, PMMA, PET, or PC, but is not limited thereto and may be made of transparent glass, etc.

[0055] The well (110) may include a plate (112), a receiving section (114), and a partition (116). The plate (112) may be a flat plate with a rectangular cross-section, and the receiving section (114) may be a shape in which the top is open by a partition (116) formed on the plate (112) and the space inside is formed by being recessed to a predetermined depth. At this time, the horizontal cross-section of the receiving section (114) may be formed in various shapes, such as a square or a circle.

[0056] For example, the well (110) may be composed of a receiving portion (114) in which a space is formed inside by a partition wall (116) arranged vertically on one side of the plate (112).

[0057] As another example, the well (110) may have multiple receiving portions (114) arranged adjacently in rows (horizontal direction) and columns (vertical direction) by means of a partition (116) formed adjacent to one side of the plate (112). At this time, the number of receiving portions (114) can be determined in various ways, such as 4, 6, 8, 12, 16, 24, 48, 96, 128, etc.

[0058] As another example, the well (110) may have multiple receiving portions (114) spaced apart in rows (horizontal direction) and columns (vertical direction) by partitions (116) formed at predetermined intervals on one side of the plate (112). At this time, the number of receiving portions (114) can be varied, such as 4, 6, 8, 12, 16, 24, 48, 96, 128, etc.

[0059] The substrate (120) supports the cell chip (100) and simultaneously receives an alternating current signal from the impedance measurement module (200). The substrate (120) is preferably a non-conductive substrate made of a transparent material such as glass or plastic to optically monitor the cell culture or cell processing state.

[0060] Referring to FIG. 5, the substrate (120) may be divided into a sensing area (A) and a non-sensing area (B) surrounding the sensing area (B). The sensing area (A) refers to an area for measuring electrical impedance generated during the process of culturing cells in the well (110) or processing cultured cells. At this time, it is preferable for the accuracy of electrical impedance measurement that the receiving portion (114) of the well (110) is disposed on the sensing area (A), and the common electrode (122) and the operating electrode (124), which will be described later, are disposed below the receiving portion (114) of the well (110). The non-sensing area (B) refers to an area surrounding the sensing area (A), and the area other than the transmission line (126) and terminal pad (128), which will be described later, may be disposed insulated.

[0061] A plurality of electrodes are formed on the surface of the substrate (120), divided into a common electrode (122) and an operating electrode (124) spaced apart from the common electrode (122), and a transmission line (126) and a terminal pad (128) may be further formed together with the common electrode (122) and the operating electrode (124).

[0062] The common electrode (122) may include a body portion (122a) positioned in the center of the sensing area (A) and a recessed portion (122b) formed by a portion of the horizontal side of the body portion (122a) being recessed inward. At this time, it is preferable that the recessed portion (122b) be positioned at the lower part of the receiving portion (114) of the well (110) described above, and may be arranged in multiple numbers spaced apart at a certain interval from the horizontal side of the body portion (122a). Additionally, the recessed portion (122b) is preferably in the shape of a semicircular arc having a certain curvature, but is not limited thereto and may be in the shape of a square or triangle recessed inward.

[0063] The operating electrode (124) may be placed within a recess (122b) formed in the common electrode (122), and may be formed in various shapes depending on the shape of the recess (122b). For example, if the recess (122b) is a semicircular arc shape with a certain curvature, the operating electrode (124) may be a disk shape corresponding thereto.

[0064] The working electrode (124) is spaced apart from the common electrode (122) so that it can form an electrical impedance according to an alternating current signal applied by the impedance measurement module (200) described later, and more specifically, by being spaced apart from each other at the bottom of the receiving portion (114) of the well (100), it is possible to collect real-time electrical impedance changes resulting from cell culture or cell treatment in the receiving portion (114) of the well (100) in a non-destructive manner and perform analysis thereon.

[0065] The transmission line (126) serves to connect the common electrode (122) and the operating electrode (124) with the terminal pad (128), and the terminal pad (128) is connected to the signal generator (210) of the impedance measurement module (200) described later to receive an AC signal and can apply the AC signal to the common electrode (122) and the operating electrode (124) through the transmission line (126).

[0066] In the case where a plurality of recesses (122b) and a plurality of operating electrodes (124) are formed in a common electrode (122) as in one embodiment of the present invention (see FIG. 5), the transmission lines (126) connected to each operating electrode (124a, 124b, 124c, 12d) can be formed with different widths to be connected to the terminal pad (128), taking into account the coupling and parasitic capacitance generated by the adjacent transmission lines (126), and can also be formed with different spacing from the adjacent transmission lines (126) depending on the distance from the terminal pad (128). More specifically, it is preferable that the transmission line (126) connected to the operating electrode (124a) far from the terminal pad (128) is formed with a wide width, and the transmission line (126) is formed with a narrower width as it is connected to the operating electrode (124d) closer to the terminal pad (128). Additionally, it is preferable that the transmission line (126) connected to the operating electrode (124a) far from the terminal pad (128) has a wider gap with the adjacent transmission line (126) compared to the transmission line (126) connected to the operating electrode (124b) close to the terminal pad (128), thereby minimizing well variation and increasing the reliability of cell analysis. At this time, the width and length of the transmission line (126) can be determined according to the theoretical sheet resistance calculation value according to Equation 1.

[0067]

[0068] Here, R sheet ε represents the sheet resistance of the transmission line (126), ε represents the resistivity of the material of the transmission line (126), l represents the length of the current flow between the operating electrode (124d) and the terminal pad (128), w represents the width of the transmission line (126), and h represents the height of the transmission line (126).

[0069] A plurality of electrodes (122, 124), transmission lines (126), and terminal pads (128) can be formed as transparent, reflective, or semi-transparent types, and can simultaneously be formed of a biocompatible conductive material.

[0070] For example, a plurality of electrodes (122, 124), transmission lines (126), and terminal pads (128) may be formed by depositing a transparent metal oxide, such as ITO (indium tin oxide), IZO (indium zinc oxide), ZnO (zinc oxide), or ITZO (indium thin zinc oxide), on a substrate (120) and then patterning, or by depositing a transparent metal oxide and a photoresist together and then etching, or by a laser patterning method. In this case, the well (110) and the substrate (120), as well as the plurality of electrodes (122, 124), transmission lines (126), and terminal pads (128) formed on the substrate (120), are all formed from a transparent material, which has an advantageous effect for optically monitoring the cell culture or cell processing status.

[0071] Meanwhile, when the common electrode (122) and the working electrode (124) are formed of a transparent material, any one of the following nanomaterials may be deposited on the surface: gold nanoparticles (Au nanoparticles, AuNPs), quantum dot (QD) reduced graphene oxide (rGO), poly(3,4-ethylenedioxythiophene)-polystyrene sulfonate (PEDOT:PSS), polyaniline (PANI), carbon nanotubes (CNT), and multi-walled carbon nanotubes (MWCNT). In this case, it is preferable that the nanomaterial be a biocompatible conductive material, but the nanomaterial is not limited to the nanomaterials exemplified above as long as it is a biocompatible conductive material. As the interfacial impedance of the common electrode or working electrode is lowered through the above nanomaterial deposition, the sensitivity and accuracy of electrical impedance measurement can be further improved.

[0072] As another example, a plurality of electrodes (122, 124), transmission lines (126), and terminal pads (128) can be formed on a substrate (120) by printing metal nanowires, which are biocompatible materials such as chromium (Cr), gold (Au), and palladium (Pd). In this case, compared to forming the plurality of electrodes (122, 124), transmission lines (126), and terminal pads (128) all from a transparent material, there is an effect of improved productivity and cost-effectiveness.

[0073] Referring again to FIG. 3, the impedance measurement module (200) can measure the electrical impedance generated during the process of culturing cells or processing cultured cells by applying an alternating current signal to the cell chip (100). Specifically, when an alternating current signal is applied during the process of culturing cells in the well (110) of the cell chip (100), as the number of cells increases due to cell culture, the area covering the plurality of electrodes (122, 124) placed at the bottom of the well (100) increases, and the current is physically obstructed, causing the electrical impedance to increase. Conversely, when an alternating current signal is applied during the process of treating the cells cultured in the well (110) of the cell chip (100) with drugs, etc., as the number of cells decreases due to cell processing, the area covering the plurality of electrodes (122, 124) placed at the bottom of the well (100) decreases, causing the electrical impedance to decrease.

[0074] Referring to FIG. 6(a), in one embodiment of the present invention, the impedance measurement module (200) includes a signal generator (210), a digital-to-analog converter (220), and an analog-to-digital converter (230).

[0075] The signal generator (210) can generate an alternating current signal controlled by the controller (300) described later, and can be configured together with the controller (300) or provided as a separate power device and linked with the controller (300).

[0076] A digital-to-analog converter (220, digital-to-analog converter; DAC) can convert an alternating current signal generated from a signal generator (210) into an analog input signal and then apply it to a plurality of electrodes (122, 124) corresponding to each well (110). At this time, the analog input signal converted by the digital-to-analog converter (220) can be applied directly to the plurality of electrodes (122, 124) or can be applied to the plurality of electrodes (122, 124) after passing through an additional configuration at a later stage.

[0077] An analog-to-digital converter (230, analog-to-digital converter; ADC) can receive analog response signals generated from multiple electrodes (122, 124) corresponding to each well (110), convert them into digital signals, and then transmit them to a controller (300). At this time, the analog-to-digital converter (230) can receive the analog response signals generated from the multiple electrodes (122, 124) directly or receive them by passing through an additional configuration at the front end. Additionally, the digital signals converted by the analog-to-digital converter (230) can be transmitted directly to the controller (300) or transmitted to the controller (300) by passing through an additional configuration at the rear end.

[0078] Referring to FIG. 6(b), in another embodiment of the present invention, the impedance measurement module (200) may further include a first signal amplifier (242, amplifier; AMP) that amplifies an analog input signal at the output of a digital-to-analog converter (220) and a second signal amplifier (244) that amplifies an analog response signal at the output of an analog-to-digital converter (230). In this case, the first signal amplifier (242) or the second signal amplifier (244) may be a plurality of.

[0079] Referring to FIG. 6(c), in another embodiment of the present invention, the impedance measurement module (200) may further include a multiplexer (250) that includes a plurality of input channels and one output channel at the front end of the second signal amplifier (244) and connects one of the plurality of input channels to the output channel. In one embodiment of the present invention, when the cell chip (100) is composed of a plurality of wells (110), a plurality of electrodes (122, 124) are disposed at the bottom of the receiving portion (114) of each well (110), and the multiplexer (250) may receive all different analog response signals generated from the plurality of electrodes (122, 124) corresponding to each well (110) and selectively output them.

[0080] Referring again to FIG. 3, the controller (300) includes a control unit (310) and a data generation unit (320).

[0081] The control unit (310) can control the overall operation of the impedance measurement module (200) and can transmit the impedance data generated by the data generation unit (320) to the data storage unit (400) of the processor (20) described later (see FIG. 7). Specifically, the control unit (310) can control the frequency of the alternating current signal applied to the cell chip (100) through the impedance measurement module (200) and the measurement time of the electrical impedance.

[0082] The data generation unit (320) can generate impedance data by receiving information about the electrical impedance measured by the impedance measurement module (200). Specifically, the data generation unit (320) can generate impedance data among resistive data, capacitive data, magnitude data, phase data, and combinations thereof. More specifically, the data generation unit (32) can generate resistive data using the real part resistance value, generate capacitive data using the imaginary part reactance value, and generate magnitude data using both the real part resistance value and the imaginary part reactance value.

[0083] Additionally, the data generation unit (320) can generate impedance data for the cell and impedance data for the culture medium, respectively.

[0084] Additionally, the data generation unit (320) can generate different data depending on the control operation of the control unit (310).

[0085] For example, the control unit (310) can control the frequency of the alternating current signal to a single value during the time the impedance measurement module (200) measures the electrical impedance, and the data generation unit (320) can generate impedance data over time.

[0086] For example, when the cells cultured in the wells (110) of the cell chip (100) are all treated with the same or different drugs, the control unit (310) controls the frequency of the alternating current signal applied from the impedance measurement module (200) to each well (110) of the cell chip (100) to the same single value, and the data generation unit (320) can generate impedance data over time.

[0087] For example, when cells cultured in the wells (110) of the cell chip (100) are treated with different drugs, the control unit (310) controls the frequency of the alternating current signal applied from the impedance measurement module (200) to each well (110) of the cell chip (100) to a different single value, and the data generation unit (320) receives information about the electrical impedance generated in each well (110) from the impedance measurement module (200) and can generate impedance data for each time period.

[0088] For example, when undifferentiated cells cultured in the wells (110) of the cell chip (100) are treated with a differentiation inducer to induce them into differentiated cells, the control unit (310) controls the frequency of the alternating current signal applied from the impedance measurement module (200) to the wells (110) of the cell chip (100) to a low frequency having a single value of 10 kHz or less or a high frequency having a single value of 100 kHz or more, and the data generation unit (320) receives information about the electrical impedance generated in each well (110) from the impedance measurement module (200) and can generate time-dependent resistive data at low frequency or time-dependent capacitive data at high frequency, respectively.

[0089] As another example, the control unit (310) can control the frequency of the alternating current signal to a plurality of different values ​​between the minimum frequency and the maximum frequency during the time the impedance measurement module (200) measures the electrical impedance, and the data generation unit (320) can generate impedance data by time and / or frequency.

[0090] For example, if one wishes to find the optimal frequency for analyzing the state of a cell cultured in a well (110) of a cell chip (100), the control unit (310) sets the minimum frequency, maximum frequency, and number of frequencies to be measured between the minimum and maximum frequencies of the alternating current signal, calculates the frequency of the alternating current signal for each interval based on Equation 2, and then controls it with multiple different values, and the data generation unit (320) can generate impedance data by time and / or frequency.

[0091]

[0092] Here, frequency(n) represents the frequency of the AC signal in each section, min frequency is the minimum frequency of the AC signal, max frequency is the maximum frequency of the AC signal, step is the number of measured frequencies, and n represents the section.

[0093] Specifically, the control unit (310) can set the minimum frequency of the AC signal to 100Hz, the maximum frequency to 100kHz, and the number of measurement frequencies to 13, calculate the frequency of the AC signal for each section as shown in Table 1, and then control the AC signal based on the calculated frequency value for the corresponding section.

[0094] Section(n) Frequency 13 100 kHz 125 6.234 kHz 113 1.623 kHz 1017.783 kHz 910 kHz 85.623 kHz 73.162 kHz 61.778 kHz 51 kHz 45 62.3 Hz 33 16.2 Hz 21 77.8 Hz 1100 Hz

[0095] Additionally, the control unit (310) can control the frequency of the alternating current signal to multiple different values ​​between the minimum frequency and the maximum frequency during the time the impedance measurement module (200) measures the electrical impedance, and at the same time, control the measurement time differently for each frequency of the alternating current signal. This has the effect of improving the accuracy of the impedance data, which is generated as the required measurement time increases as the number of frequencies of the alternating current signal applied to the cell chip (100) increases and the frequency value decreases.

[0096] Thus, the cell impedance measuring device of the present invention can control the operation of the control unit (310) in various ways according to the purpose of cell impedance measurement, and the data generation unit (320) can generate different data according to the control operation of the control unit (310), thereby greatly improving the convenience of cell analysis.

[0097]

[0098] Referring again to FIGS. 1 and FIGS. 2, a cell analysis system (1) according to one embodiment of the present invention includes a cell impedance measuring device (10) and a processor (20). Since the cell impedance measuring device (10) has been described above, a detailed description thereof is omitted.

[0099] The processor (20) can analyze the state of a cell using resistive data, capacitive data, size data, phase data, and any combination thereof generated by the cell impedance measuring device (10), and may include a data storage unit (400) and a cell analysis unit (500).

[0100] The data storage unit (400) can store various impedance data generated by the data generation unit (320) within the controller (300) of the cell impedance measuring device (10). The impedance data stored in the data storage unit (400) includes not only any one of resistive data, capacitive data, magnitude data, phase data, and combinations thereof, but also impedance data generated by the data generation unit (320) according to the control operation of the control unit (310), i.e., impedance data by time and frequency. In addition, the data storage unit (400) can store the identification number of the well (110) of the cell chip (100) and the impedance measurement time, etc., that match each impedance data. Furthermore, the data storage unit (400) can store reference data regarding previously measured culture medium (culture solution), cells, drugs, etc.

[0101] The cell analysis unit (500) can analyze the state of a cell using impedance data stored in the data storage unit (400) and a cell analysis model. The term "cell analysis model" used in the present invention refers to an algorithm or set of algorithms for generating various graphs using impedance data (using additional reference data if necessary) and analyzing the state of a cell through the interpretation of the generated graphs, and the "cell analysis model" may be configured in the form of a software module.

[0102] FIGS. 8(a) to 8(d) are graphs illustrating a model for analyzing the state of a cell during a cell culture process in a processor according to one embodiment of the present invention.

[0103] Referring to FIGS. 8(a) to 8(d), the cell analysis unit (500) can generate a graph of changes in cell size data over time (see FIG. 8(a)), a graph of changes in cell resistance data over time (see FIG. 8(b)), a graph of changes in cell capacitance data over time (see FIG. 8(c)), a graph of changes in cell impedance index over time (see FIG. 8(d)), or a graph of changes in culture medium capacitance data over time (see FIG. 8(e)) using impedance data stored in the data storage unit (400) and a cell analysis model.

[0104] In addition, the cell analysis unit (500) can simultaneously analyze the cell culture state and the culture medium state over time using a cell analysis model and a graph of changes in cell size data over time, a graph of changes in cell resistance data over time, a graph of changes in cell capacitance data over time, a graph of changes in cell impedance index over time, or a graph of changes in culture medium capacitance data over time.

[0105] FIGS. 9(a) and FIGS. 9(b) are graphs illustrating a model for analyzing the survival or lysis status of cells during the processing of cultured cells in a processor according to one embodiment of the present invention.

[0106] Referring to FIGS. 9(a) and 9(b), the cell analysis unit (500) can generate a graph of changes in cell viability over time (see FIG. 9(a)) or a graph of changes in cell cytolysis over time (see FIG. 9(b)) using impedance data stored in the data storage unit (400) and a cell analysis model. At this time, the cell analysis unit (500) can set the state in which only cells exist without treating drugs, etc. in the well (110) of the cell chip (100) as viability 100 and cytolysis 0, and set the state in which there are no cells or all cells that have died as viability 0 and cytolysis 100, and then generate a graph of changes in cell viability over time or a graph of changes in cell cytolysis over time using the impedance data stored in the data storage unit (400).

[0107] In addition, the cell analysis unit (500) can analyze the state of the cell in various ways by extracting the time at which the cell lysis rate is 50% from the graph of changes in cell viability over time or the graph of changes in lysis rate as the KT50 (killing time 50) index, the CC50 (cytotoxicity concentration 50) which indicates the concentration at which a specific compound or drug reduces the cell viability to 50%, and the IC50 (inhibitory concentration 50) which indicates the concentration at which a specific compound or drug inhibits the cell function to 50%.

[0108] FIGS. 10 and 11 are graphs illustrating a model for analyzing the optimal frequency per cell by a processor according to one embodiment of the present invention.

[0109] Referring to FIGS. 10(a) to FIGS. 10(d), the cell analysis unit (500) can generate a graph of changes in cell size data by frequency (see FIG. 10(a)), a graph of changes in cell resistance data by frequency (see FIG. 10(b)), a graph of changes in cell capacitance data by frequency (see FIG. 10(c)), or a graph of changes in cell phase data by frequency (see FIG. 10(d)) using impedance data stored in the data storage unit (400) and a cell analysis model.

[0110] Meanwhile, the cell analysis unit (500) can additionally generate normalized impedance data based on impedance data at a specific time using impedance data stored in the data storage unit (400) and mathematical formula 3, and can additionally generate rate of change data of normalized impedance data using the generated normalized impedance data and mathematical formula 4.

[0111]

[0112] Here, Z(selected time) represents the impedance data value at a specific time.

[0113]

[0114] Here, Z(0) represents the impedance data value at the initial time point.

[0115] Referring to FIGS. 11(a) to FIGS. 11(c), the cell analysis unit (500) can generate a graph of magnitude data change by time and frequency using impedance data stored in the data storage unit (400), additionally generated normalized impedance data in the cell analysis unit (500), and a cell analysis model (see FIG. 11(a)), accumulate (see FIG. 11(b)), and then generate a normalized graph (see FIG. 11(c)) by normalizing the generated graph of magnitude data change by time and frequency.

[0116] In addition, the cell analysis unit (500) can identify the frequency with the highest rate of change of impedance magnitude (Ω) using normalized impedance data, rate of change data of normalized impedance data, and a cell analysis model, and thereby analyze the optimal frequency at which changes in electrical impedance are well detected for each characteristic of the cell. At this time, the cell analysis model has a time complexity Data-driven algorithms such as quick and heap sort algorithms can be used.

[0117] In addition, the cell analysis unit (500) can identify the frequency with the highest rate of change of impedance magnitude (Ω) using the generated normalization graph and cell analysis model, and thereby analyze the optimal frequency at which changes in electrical impedance are well detected for each characteristic of the cell. At this time, the cell analysis model may use image-based artificial intelligence algorithms such as a Convolutional Neural Network (CNN).

[0118] FIGS. 12 to 15 are graphs illustrating a model for analyzing growth indicators and the state of undifferentiated or differentiated cells according to cell amount at different cell injection concentrations by a processor according to one embodiment of the present invention.

[0119] Referring to FIG. 12(a) and FIG. 12(b), the cell analysis unit (500) can generate size data or capacitive data bar graphs at specific frequencies according to cell injection concentration and time, respectively, using impedance data stored in the data storage unit (400) and a cell analysis model.

[0120] In addition, the cell analysis unit (500) can analyze changes in cell growth during the culture time according to the initial cell injection concentration and the experimental period and drug treatment time according to the initial cell injection concentration using the generated bar graph and cell analysis model.

[0121] Referring to FIG. 13(a) and FIG. 13(b), the cell analysis unit (500) can generate, respectively, size data or capacitive data linear graphs at a specific frequency (100 kHz) according to cell injection concentration and time using impedance data stored in the data storage unit (400) and a cell analysis model.

[0122] Additionally, the cell analysis unit (500) can analyze the suitability of data used in the cell analysis model using the generated linear graph and the cell analysis model, and the cell analysis model can use the coefficient of determination of change amount (R-squared) and linear analysis, etc.

[0123] Referring to FIG. 14(a) and FIG. 14(b), the cell analysis unit (500) can generate a graph of resistance data change at a specific frequency (1 kHz) according to the culture time during which monocytes differentiate into MO-type macrophages (see FIG. 14(a)) or a graph of capacitance data change at a specific frequency (100 kHz) (see FIG. 14(b)).

[0124] In addition, the cell analysis unit (500) can analyze the state of monocytes having a floating phenotype and macrophages in the form of MO that are differentiated by stimulation with a differentiation-inducing substance and adsorbed to an electrode using the generated change graph and cell analysis model, and simultaneously analyze the state of monocytes with different phenotypes and macrophages differentiating into MO forms in real time.

[0125] Referring to FIG. 15(a) and FIG. 15(b), the cell analysis unit (500) can generate a graph of changes in resistance data at a specific frequency (1 kHz) according to the culture time in which monocytes differentiate into macrophages of the MO, M1, or M2 type (see FIG. 15(a)) or a graph of changes in capacitance data at a specific frequency (100 kHz) (see FIG. 15(b)).

[0126] In addition, the cell analysis unit (500) can analyze in real time the phenotype, morphological changes, polarization and maturation status, or changes in cell migration according to the phenotype in additional differentiation forms M1 or M2 following specific differentiation-inducing stimulating substances after monocytes have differentiated into MO-type macrophages, using the generated change graph and cell analysis model, and can also perform an analysis of erythrophagocytosis associated with the suppression of pro-inflammatory reactions, tissue repair, and promotion of anti-inflammatory cytokine production.

[0127] Meanwhile, the processor (20) may further include an analysis model learning unit (not shown) that machine learns the cell analysis model used in the cell analysis unit (500) through the analysis of characteristics of flow cytometry data for specific cells, immunofluorescence data for specific cells, or cell-specific impedance data, imaging analysis, and prediction modeling.

[0128] In addition, the above analysis model learning unit (not shown) can be used to calculate the unique impedance value for each cell by collecting impedance data stored in the data storage unit (400) and various graphs and analyzed data generated from the cell analysis unit (500) and learning. Through this, it is possible to predict toxicity assessment or simulate the timing of drug treatment according to cell growth stages, thereby reducing the experimental time by predicting the cell analysis results.

[0129] Additionally, the cell analysis system (1) may further include a terminal device (not shown) that displays the analysis results of the cell state analyzed by the processor (20).

[0130] The embodiments of the present invention described above are merely exemplary, and the scope of protection of the present invention may include various modifications and equivalents therefrom by those skilled in the art.

[0131] 1: Cell Analysis System

[0132] 10: Cell impedance measuring device

[0133] 20: Processor

[0134] 100: Cell Chip

[0135] 200: Impedance measurement module

[0136] 300: Controller

[0137] 400: Data storage unit

[0138] 500: Cell Analysis Department

Claims

1. A cell chip comprising a well formed with a receiving portion for culturing cells or processing cultured cells, and a substrate having a plurality of electrodes formed thereon; An impedance measurement module that measures the electrical impedance of the cell by applying an alternating current signal to the cell chip; and A controller that controls the impedance measurement module and generates impedance data for the measured electrical impedance; A cell impedance measuring device in which the cell chip, the impedance measuring module, and the controller are electrically connected to each other.

2. In Claim 1, The above substrate is divided into a sensing area and a non-sensing area around the sensing area, and A cell impedance measuring device, wherein the receiving portion of the above-mentioned well is disposed on the above-mentioned sensing area.

3. In Claim 2, The above electrode includes a common electrode and an operating electrode spaced apart from the common electrode, and A cell impedance measuring device in which the common electrode and the working electrode are formed at the lower part of the receiving portion of the well.

4. In Claim 3, A cell impedance measuring device in which the substrate, the common electrode, and the working electrode are formed of a transparent material.

5. In Claim 4, A cell impedance measuring device having a nanomaterial of any one of gold nanoparticles (AuNPs), quantum dots (QDs), reduced graphene oxide (rGO), poly(3,4-ethylenediooxythiophene)-polystyrene sulfonate (PEDOT:PSS), polyaniline (PANI), carbon nanotubes (CNT), and multi-walled carbon nanotubes (MWCNT) deposited on the surface of the common electrode and the working electrode.

6. In Claim 1, A cell impedance measuring device in which a plurality of wells are spaced apart in horizontal and vertical directions on the substrate.

7. In Claim 1, The above impedance measurement module is, A signal generator that generates the above alternating current signal; A digital-to-analog converter (DAC) that converts an AC signal generated by the signal generator into an analog input signal and applies it to the plurality of electrodes corresponding to the wells; and A cell impedance measuring device comprising: an analog-to-digital converter (ADC) that receives an analog response signal generated from a plurality of electrodes corresponding to the well, converts it into a digital signal, and transmits it to the controller.

8. In Claim 7, A first signal amplifier that amplifies the analog input signal at the output of the digital-to-analog converter (DAC); and A cell impedance measuring device further comprising a second signal amplifier that amplifies the analog response signal at the front end of the analog-to-digital converter (ADC).

9. In Claim 8, A cell impedance measuring device comprising a plurality of input channels and one output channel in the preceding stage of the second signal amplifier, and further comprising a multiplexer connecting any one of the plurality of input channels to the output channel.

10. In Claim 1, The above controller is, A control unit that controls the frequency of the above alternating current signal and the measurement time of the above electrical impedance; and A cell impedance measuring device comprising: a data generation unit that receives information regarding the measured electrical impedance and generates impedance data among resistive data, capacitive data, magnitude data, phase data, and combinations thereof.

11. In Claim 10, The above control unit controls the frequency of the AC signal to a single value during the measurement time, and The above data generation unit is a cell impedance measuring device that generates the above impedance data over time.

12. In Claim 10, The above control unit controls the frequency of the AC signal to a plurality of different values ​​between a minimum frequency and a maximum frequency during the measurement time, and A cell impedance measuring device in which the data generating unit generates the impedance data by time or / and frequency.

13. A cell impedance measuring device of any one of claims 1 to 12; and A cell analysis system comprising: a processor that analyzes the state of a cell using resistive data, capacitive data, magnitude data, phase data, and any combination thereof, generated by the cell impedance measuring device.

14. In Claim 13, The above processor is, A data storage unit for storing the above impedance data; and A cell analysis system comprising: a cell analysis unit that analyzes the state of a cell using the impedance data stored in the data storage unit and a cell analysis model.

15. In Claim 13, A cell analysis system further comprising: a terminal device for displaying the analysis results of the cell state analyzed by the processor.

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