Device for taking electrochemical measurements, and associated probe and bioreactor

Abstract

Disclosed is a device (100) for taking electrochemical measurements with a view to measuring physico-chemical quantities and / or concentrations of molecules in a solution (300), the device comprising: - a set (13) of sensors arranged on a carrier (11), the set comprising at least one amperometric sensor (134, 134a, 134b) and at least one OCP sensor (131, 131a, 131b, 132, 132a, 132b) for taking measurements of open-circuit potential, the amperometric sensor comprising a working electrode (1341, 1341a, 1341b), a reference electrode (1342, 1342a, 1342b) and a counter-electrode (1343, 1343a, 1343b), the OCP sensor comprising a reference electrode (1312, 1312a, 1312b, 1322, 1322a, 1322b) and a working electrode (1311, 1311a, 1311b, 1321, 1321a, 1321b); - an electronic device (10) for acquiring and processing measurement signals delivered by the sensors, the electronic device further comprising at least one circuit (103) for controlling the bias of the electrodes of the sensors.

Description

[0001] 43825

[0002] 1

[0003] DESCRIPTION TITLE: Electrochemical measurement device, probe and associated bioreactor

[0004] technical field

[0005] The present invention belongs to the technical field of electro-chemical measurement and relates more particularly to a device for measuring electrochemical quantities and / or concentrations of molecules (analytes, dissolved gases, etc.) in a solution contained in a bioreactor.

[0006] It finds applications particularly in the field of bioproduction.

[0007] State of the art

[0008] Multi-sensor probes are available for various applications, such as monitoring water quality in natural streams, swimming pools, wastewater treatment plants, and industrial effluents. These probes can integrate several types of sensors, sometimes to measure up to four distinct parameters, for example, pH, conductivity, dissolved oxygen, and temperature, using a remote reader.

[0009] Adapting multi-parameter measurement systems in bioreactors poses several difficulties, including size, cost, and sterilization.

[0010] Document EP3514223A1 presents, for example, a probe for a bioreactor equipped with several sensors co-integrated on the same support to perform a measurement of three electrochemical quantities.

[0011] When measurements are taken with sensors whose electrodes are close together and possibly mounted on the same support, the problem of potential interference between the electrodes of different sensors arises. In particular, interference can be caused by the different types of transducers used. Thus, when a measurement is taken on one sensor, depending on the type of transduction used, this measurement can interfere with a second sensor located nearby. To mitigate this, the different measurements can be taken sequentially. For example, a measurement is taken using an amperometric sensor, and then, once this measurement is complete, a measurement is taken using another sensor.This sequencing, combined with the generally required potential stabilization time of certain measurement electrodes, can induce an unacceptable response time for some applications, particularly when monitoring a bioprocess.

[0012] Presentation of the invention

[0013] The present invention aims to overcome all or part of the drawbacks presented above, and proposes an improved electro-chemical measurement device for measuring physico-chemical quantities and concentrations of molecules in a solution.

[0014] To this end, the present invention relates to an electrochemical measurement device for measuring physicochemical quantities and / or concentrations of molecules in a solution, the device comprising:

[0015] - a set of sensors arranged on a support, said set comprising at least one amperometric sensor and at least one "OCP" sensor for open circuit potential measurements, the amperometric sensor comprising a working electrode, a reference electrode and a counter electrode and the OCP sensor comprising a reference electrode and a working electrode;

[0016] - an electronic device for acquiring and processing measurement signals from said sensors, the electronic device further comprising at least one control circuit for the polarization of the electrodes of said sensors, the control circuit being configured to:

[0017] As a preliminary step to a measurement sequence, apply the following to the amperometric sensor:

[0018] a VamperoWE polarization potential on the working electrode;

[0019] a bias potential VamperoRE on the reference electrode, with VamperoRE = VamperoWE - Vbias where Vbias is a voltage differential between the working electrode and the reference electrode;

[0020] then, perform k times (where k is a non-zero integer) a sequence of measurements comprising steps consisting of:

[0021] During an amperometric measurement phase, acquire measurement signals from the amperometric sensor while maintaining the VamperoWE polarization potential on the working electrode and maintaining the VamperoRE polarization potential on the reference electrode; and

[0022] during an OCP measurement phase, acquire measurement signals from the OCP sensor while polarizing the OCP sensor by applying a polarization potential VpolOCP dependent on the polarization potential VamperoCE of the counter electrode of the amperometric sensor on the reference electrode of the OCP sensor, the polarization potential VamperoRE on the reference electrode and the polarization potential VamperoWE on the working electrode of the amperometric sensor, being maintained during the measurement phase on the OCP sensor.

[0023] By maintaining this polarization of the amperometric sensor during measurement phases distinct from those during which the sensor is in use, an equilibrium of ionic species in the medium is maintained, and measurements can be performed sequentially without the time required for electrode stabilization. This allows for a measurement sequence while minimizing interference and the time required for these measurements.

[0024] Advantageously, the counter electrode of the amperometric sensor is coupled to the reference electrode of the OCP sensor and / or the bias potential VpolOCP is substantially equal to the bias potential VamperoCE of the counter electrode of the amperometric sensor or to a measured value Vmes from the counter electrode and close to VamperoCE, with Vmes = VamperoCE + Vdelta, where Vdelta is a voltage differential between -100 mV and +100 mV.

[0025] Advantageously, the sensor set further includes at least one EIS sensor for electrochemical impedance spectroscopic measurements, the EIS sensor comprising a first working electrode and a second working electrode, the control circuit being configured in this case so that, during said sequence and during an EIS measurement phase during which measurement signals are acquired from the EIS sensor:

[0026] apply a bias potential VpolEIS on the second working electrode of the EIS sensor, and apply on the first working electrode of the EIS sensor a bias potential equal to the sum of a DC component equal to VpolEIS + Voffset, with Voffset a DC voltage between -1000 mV and +1000 mV, preferably zero, and an AC component, while maintaining the bias potential VamperoRE on the reference electrode and maintaining the bias potential VamperoWE on the working electrode of the amperometric sensor.

[0027] According to a particular feature of the invention, the polarization potential VpolEIS is equal to the polarization potential VamperoCE of the counter electrode of the amperometric sensor or equal to the polarization potential VpolOCP of the reference electrode of the OCP sensor, or equal to the highest potential between said potential of the counter electrode of the amperometric sensor and said potential of the reference electrode of the OCP sensor.

[0028] According to another particular feature of the invention, after the EIS measurement phase and prior to another measurement phase, the polarization of the first working electrode of the EIS sensor and the polarization of the second working electrode of the EIS sensor are interrupted.

[0029] Advantageously, the VamperoRE polarization potential on the reference electrode, the VamperoWE polarization potential on the working electrode and the VamperoCE polarization potential on the counter electrode of the amperometric sensor are maintained throughout the measurement sequence.

[0030] Advantageously, the sensor set includes one or more amperometric sensors to measure an oxygen level, at least one pair of OCP sensors to measure a carbon dioxide level and / or to measure a hydrogen potential, and at least two EIS sensors to measure a conductivity and impedance of the solution.

[0031] Advantageously, at least one amperometric sensor and at least one OCP sensor have respective reference electrodes of equal surface area.

[0032] Advantageously, the support is a printed circuit board, and a block of electrically insulating material is disposed on one face of the printed circuit board between the electrodes of the second sensor and the electrodes of the amperometric sensor.

[0033] Advantageously, the support is a printed circuit board comprising a first face and a second face opposite the first face, the electrodes of the OCP sensor being distributed on the first face, the electrodes of the amperometric sensor being arranged on the second face, and when the device is equipped with an EIS sensor, the electrodes of said EIS sensor are arranged on the second face. According to a particular feature of the invention, the support is in particular a printed circuit board and said electronic device is arranged on a second support distinct from said support, in particular a second printed circuit board distinct from the first board.

[0034] Another object of the invention is an electrochemical measuring probe for measuring physicochemical quantities and concentrations of molecules in a solution, comprising a device as described above, said probe comprising:

[0035] - a probe housing head intended to be placed outside the solution;

[0036] - an oblong-shaped probe housing body intended to be placed in the solution.

[0037] The electronic device can then be integrated into the probe housing head, and the probe housing body is intended to accommodate at least a portion of a support for said sensor assembly, the probe housing head and / or the probe housing body being provided with at least one mechanical coupling element to allow a removable assembly of the housing head and the housing body into a probe housing.

[0038] Advantageously, the probe body includes a fixing structure, in particular a thread, to allow the probe body to be assembled onto a receiving port, in particular one equipped with an additional thread.

[0039] Another object of the invention is an electrochemical measurement system for measuring physicochemical quantities and concentrations of molecules in a solution, said system comprising:

[0040] - an electro-chemical measurement probe as described above; - a bioreactor equipped with a chamber intended to accommodate said solution and a port for receiving a probe housing body arranged between an external volume and an internal volume of the chamber, in particular a port according to the PG 13.5 standard, the probe housing body extending into said chamber when the probe housing is assembled with the bioreactor.

[0041] Presentation of the drawings

[0042] The figures are provided for illustrative purposes only to aid understanding of the invention without limiting its scope. The various elements may be represented schematically and are not necessarily to the same scale. Throughout the figures, identical or equivalent elements are given the same numerical reference.

[0043] This is illustrated as follows:

[0044] [Fig. 1]: a perspective view of an electro-chemical measuring probe, according to one embodiment of the invention;

[0045] [Fig. 2]: an exploded and cross-sectional view of an electro-chemical measurement system, according to one embodiment of the invention;

[0046] [Fig. 3]: a perspective view of an electrochemical measuring probe housing head;

[0047] [Fig. 4A]: a perspective view of an embodiment of an electronic device for the acquisition and processing of measurement signals from sensors of an electro-chemical measurement device and for controlling the polarization of the electrodes of the sensors of this electro-chemical measurement device;

[0048] [Fig. 4B]: an electrical diagram of the electronic device;

[0049] [Fig. 5A]: a view of a first face of the sensor support of the electro-chemical measurement device, according to a first embodiment;

[0050] [Fig. 5B]: a view of the first face of the sensor support of the electro-chemical measurement device, according to the first embodiment;

[0051] [Fig. 5C]: a view of a first face of the sensor support of the electro-chemical measurement device, according to a second embodiment;

[0052] [Fig. 5D]: a cross-sectional view of the sensor support of the electrochemical measurement device, according to a third embodiment;

[0053] [Fig. 6]: a perspective view of an electrical connection means between the sensor support and the electronic device of the electro-chemical measuring device; [Fig. 7A], [Fig. 7B] and [Fig. 7C]: views of different embodiments of a mechanical connection means between the probe housing head and a probe housing body of the electro-chemical measuring probe;

[0054] [Fig. 8A]: an electrical diagram of the assembly of an amperometric sensor of the electro-chemical measurement device, according to an embodiment of the invention;

[0055] [Fig. 8B]: an electrical diagram of the assembly of a sensor for open-circuit potential measurements of the electrochemical measuring device, according to an embodiment of the invention.7

[0056] [Fig. 8C]: an electrical diagram of the assembly of a sensor for electrochemical impedance spectroscopic measurements of the electrochemical measurement device, according to an embodiment of the invention.

[0057] [Fig. 9]: a diagram of the main phases of a measurement sequence, according to one embodiment of the invention.

[0058] [Fig. 10]: an electrical diagram of a sensor assembly, according to an embodiment of the invention.

[0059] [Fig. 11 A]: a timing diagram corresponding to a first mode of polarization of the sensors prior to and during a measurement sequence.

[0060] [Fig. 1 IB]: a timing diagram corresponding to a second mode of polarization of the sensors prior to and during a measurement sequence.

[0061] Detailed description of implementation methods

[0062] The present invention aims, in particular, to provide an improved electrochemical measurement device for measuring physicochemical quantities and the concentrations of molecules in a solution. The electrochemical measurement device comprises a set of several sensors with different transduction modes, integrated on a single support, and an electronic device for controlling the polarization of the sensor measurement electrodes, acquiring and processing the measurement signals from the sensors.

[0063] Advantageously, the sensor support and the electronic device can be arranged on separate elements that can be removably connected and assembled together. The sensor support can thus be designed as a single-use element, while the electronic device can be reused. In one embodiment, the electrochemical measurement device is integrated into a probe to facilitate its use.

[0064] Figure 1 shows a perspective view of an example of an electrochemical measuring probe 200 according to an embodiment of the invention. The electrochemical measuring probe 200 mainly comprises a probe housing head 20 and a probe housing body.

[0065] The probe housing head 20 is designed to accommodate an electronic device 10 as described above, an embodiment of which will be shown in more detail in connection with Figure 4A and Figure 4B. The probe housing body 21 is designed to accommodate at least a portion of a sensor support 11 for a sensor array 13. The body has an elongated ob shape, for example, a cylinder. The housing body 21 can be made of a biocompatible material, preferably having: low release properties, preferably insensitive to sterilization by gamma irradiation and / or moist heat drying and / or acid cleaning, particularly with hydrochloric acid, without this being a limitation to the present invention. Preferably, the housing body 21 is made of a material that is approved by the U.S. Food and Drug Administration (FDA).The housing body 21 can be made, for example, of a material such as polypropylene, 316L steel, polyetheretherketone (PEEK), or polytetrafluoroethylene (PTFE). In one particular embodiment, the housing body 21 is made of molded polypropylene.

[0066] The probe housing head 20 and the probe housing body 21 are each equipped with a mechanical connection means or mechanical coupling element, several embodiments of which are shown in Figures 7A to 7C. The coupling element(s) allow for a removable assembly of the head and the body. In other words, once assembled, the probe housing head 20 and the housing body 21 can be separated again into two distinct elements.

[0067] The probe housing body 21 may be further provided with a mounting structure 212 to allow its assembly onto a port of a container for a solution or medium to be studied, in particular a bioreactor. The mounting structure 212 is preferably a thread. Furthermore, to ensure a seal between the probe housing body 21 and the bioreactor port, and to maintain the sterility of the interior of the bioreactor to which the probe 200 is attached after screwing, the body 21 may include an O-ring 213 positioned at the mounting structure 212. The O-ring 213 is made, for example, of a material such as biocompatible silicone, ethylene propylene diene monomer (EPDM), polytetrafluoroethylene (PTFE), or fluorocarbon-based elastomers (FKM).

[0068] Because the sensor assembly 13 can be separated from the electronic device 10, the constraints related to the sterilization cycle are no longer imposed on the sensor assembly 13 and the electronic device 10. The electronic device 10 no longer necessarily needs to be sterilized by gamma irradiation or moist heat, for example. This reduces the risk of damage to sensitive components on the electronic device.

[0069] Furthermore, since the conditioning of the electrochemical and physical measurement signals is performed within the electronic device 10, the electronic device 10 can be reused as it can be easily detached from the sensor assembly. This possibility of detachment also facilitates the reconfiguration of the electronic device to allow for association and potential calibration with another set of sensors having different characteristics. Thus, a single electronic device 10 can be used with different sensor assemblies 13 and therefore with different probe housings 21.

[0070] Figure 2 shows an exploded and cross-sectional view of an example of an electrochemical measurement system 400 incorporating a probe as described previously. The system 400 comprises a bioreactor 500, containing a solution 300 to be studied, and the electrochemical measurement probe 200 attached to the bioreactor 500.

[0071] The bioreactor 500 consists of a housing 50 equipped with a port 51 for the probe housing body 21. The arrangement is such that the probe housing body 21 extends partly into an external volume 53 of the housing and partly into an internal volume 52 of said housing. The probe housing head 21 is located entirely within the external volume 53. The port 51 can, for example, be a port conforming to standard PG 13.5. In the illustrated embodiment, the port 51 is positioned on a cover 55 of the housing 50 of the bioreactor 500.

[0072] Figure 2 shows the through-hole design of the sensor support, which extends along the entire length and inside the probe housing 20 and is oriented such that one end 12 of the support and the sensor assembly 13 are located outside the housing. End 12 is intended for electrical connection to the electronic device 10. The sensor assembly 13 is intended to be immersed in the solution 300.

[0073] Furthermore, in order to achieve a complete seal between the inner volume 51 and the solution 300 on the one hand, and the outer volume 53 on the other, a filling material such as a resin (not shown here) can be injected into the probe housing body 21 to fill the volume between said body and the sensor support 11.10

[0074] The probe housing head 20 and the electronic device 10 can be designed so as to be contained within a volume of less than 100 cm³ 3 in order not to interfere with the integration of probe 200 on bioreactor 500.

[0075] As can be seen in Figure 3, the electronic device 10 can be at least partially realized on an electronic board, for example of PCB type (for "Printed Circuit Board").

[0076] Figure 3 shows a perspective view of the housing head 20 of the electrochemical measuring probe. The housing head 20 includes an output port 202, for example, to transmit all measurements typically as digital signals to a controller for a bioprocess in solution 300.

[0077] In a particular embodiment of the invention, the output port 202 comprises or is associated with at least one element among a wired output, a Universal Asynchronous Receiver Transmitter (UART), a synchronous serial data bus (Serial Peripheral Interface, SPI), a bidirectional half-duplex synchronous serial bus (Inter-Integrated Circuit, I2C), a CAN (Control Area Network) bus, or according to a communication protocol (for example based on ISO / IEC / IEEE 8802-3, Ethernet).Alternatively or in combination, the output port 202 uses a wireless communication technology, for example according to the Bluetooth® standard, the ZigBee® protocol, the Wi-Fi™ protocol to be able to exchange signals with an automaton typically providing at least one function of control and / or automation of measurement procedures and / or control of experimental parameters, and / or data management and / or user interface.

[0078] Figure 4A and Figure 4B respectively represent a perspective view and an equivalent diagram of an example of the aforementioned electronic device 10, integrated into the probe housing head 20 and configured to perform an on-board potentiostat function.

[0079] The electronic device 10, arranged here on an electronic board 101, includes, in particular, a system-on-chip 103. Besides processing signals from the measurement sensors, the system-on-chip 103 is configured, in particular, to apply a specific sequence to the polarization signals of the measurement electrodes. An example of such a sequence will be given later. It allows measurements to be taken on different sensors, typically sequentially, in order to...

[0080] 11

[0081] The goal is to limit interference between sensors while also limiting the sequence duration and, consequently, the total time required to perform a series of measurements. In one particular embodiment, the 103 system-on-chip can be equipped with an ADuCM355 chip from Analog Devices. The 103 system-on-chip includes a microcontroller, associated with one or more memories, and can be equipped with, or associated with, a signal conditioning circuit for signals to or from the sensors. Such a conditioning circuit can consist of amplifier(s) and one or more analog-to-digital converters (ADCs), as well as one or more digital-to-analog converters (DACs).

[0082] In the specific embodiment shown in Figures 4A-4B, the electronic device 10 is provided with an interface 105, for example, a JTAG (Joint Test Action Group) type interface for debugging and testing. Such an interface 105 can also be used to program the system-on-chip 103.

[0083] A first analog conditioning circuit 106 interfaces between the system-on-chip 103 on the one hand, and on the other hand, a measurement stage and / or OCP-type sensors 131a, 131b, 132a, 132b intended for open-circuit potential measurements, for example, to measure pH or CO2. This conditioning circuit 106 typically performs one or more of the following functions: amplification, filtering, attenuation, and signal shaping.

[0084] A second analog conditioning circuit 107 is also provided in this particular example. This circuit 107 interfaces between the system-on-chip 103 and a measurement stage and / or at least one temperature sensor or probe. This second conditioning circuit 107 typically performs one or more of the following functions: amplification, filtering, attenuation, and shaping. Temperature measurement can be performed, for example, using a platinum resistance temperature detector (RTD), such as a PT100 probe.

[0085] In this embodiment, the electronic device 10 can also be equipped with a power supply management module 108, an input / output management module 102 connected to the output port 202, and an indicator light 109. The power supply management module 108 of the electrochemical measurement device 100 can, for example, be configured to receive an input voltage 43825

[0086] 12

[0087] Vin continuous between 10 V and 36 V, and to convert said voltage, typically into voltages of 5 V and 3.3 V.

[0088] For example, the indicator light 109 is in the form of a light-emitting diode (LED). The indicator light 109 allows the operating status of the electrochemical measuring device 100 to be visually monitored by means of a color code and / or a flashing sequence.

[0089] The sensor input port 111 allows a connection to be established with the sensor assembly mounted on another support. This input port can be, for example, a so-called "high-density" connector, meaning one with a high connection capacity, for example, on the order of twenty connections, in a small space. In one particular embodiment, the sensor input port 111 could be of the MECT-110-01-MD-RA1 type. In another example, the sensor input port 111 could be of the micro HDMI type. Generally, a port type that provides a good equivalent connection density and is compatible with repeated use is preferred. For this reason, connections made of a material and a configuration suitable for frequent and repeated use are provided, for example, gold connections.The sensor input port 111 is intended to receive the end 12 of the sensor support 11, this end having additional tracks (not shown here) to the connections of said port.

[0090] Advantageously, the electronic device 10 can be reused and configured according to measurement needs and the sensors with which it is to be associated. The electronic device 10 is therefore adaptable for monitoring different types of bioreactions.

[0091] As previously stated, in an electrochemical device 100 as implemented according to the invention, the set of electrochemical measurement sensors 13 can be arranged on a support 11 separate from that of the electronic device 10 for processing and controlling the polarization of the measurement electrodes.

[0092] The sensor array 13 mounted on such a support typically includes at least one amperometric sensor and at least one second sensor, for performing measurements different from those of the amperometric sensor. This second sensor may be an "OCP" (Open Circuit Potential) sensor for performing open-circuit potential measurements or an "EIS" (Electrochemical Impedance Spectroscopy) sensor for performing 43825

[0093] 13

[0094] Electrochemical impedance spectroscopic measurements are provided. Advantageously, one or more amperometric sensors, at least one OCP sensor, and at least one EIS sensor are provided on the same support 11. The amperometric sensor(s) may be designed to measure oxygen levels. The OCP sensor(s) may be dedicated to measuring carbon dioxide and / or hydrogen potential (pH). The EIS sensor(s) may be designed to measure conductivity and / or impedance.

[0095] Figures 5A, 5B, 5C, and 5D represent, respectively, views of the first face of a sensor support according to a first embodiment (Figures 5A and 5B), a view of the first face according to a second embodiment (Figure 5C), and a cross-sectional view of the sensor support according to a third embodiment (Figure 5D). In these examples, the sensor support is in the form of a PCB (Printed Circuit Board) on which metal electrodes are arranged.

[0096] In this example, sensor set 13 includes:

[0097] Four sensors called "OCP", 131a, 131b, 132a and 132b, for open circuit potential measurements;

[0098] Two sensors called "EIS", 133a and 133b, for electrochemical impedance spectroscopic measurements;

[0099] Two amperometric sensors 134a and 134b; and

[0100] A platinum resistance thermometer 135.

[0101] Each OCP 131a-b and 132a-b sensor comprises a reference electrode (1312a-b and 1322a-b, respectively) and a working electrode (131la-b and 1321a-b, respectively). In one particular embodiment, a first pair of OCP 131la-b sensors is used to measure carbon dioxide, and a second pair of OCP 132a and 132b sensors is used to measure the pH of solution 300. For CO2 measurement, the electrodes can be, for example, of the Severinghaus type and include a CO2-permeable membrane. The CO2 measurement is obtained indirectly from the measurement of pH variations in solution 300, these variations being induced by changes in the CO2 concentration in said solution.

[0102] For pH measurement, the electrodes can, for example, be made of a metal oxide, such as iridium, tungsten, tantalum or ruthenium, but also 43825

[0103] 14

[0104] pH-sensitive polymers such as polyaniline (PANI) or ion-selective electrodes (also known as Ion Selective Electrodes, ISE). Each EIS 133a-b sensor here comprises a first working electrode 1331a-b and a second working electrode 1332ab.

[0105] The EIS 133a and 133b sensors can be designed to measure the conductivity and impedance of solution 300, particularly over a frequency band from 20 kHz to 200 kHz. For conductivity measurement, the electrodes can, for example, be made of carbon-based materials such as graphene (graphite, nanotubes, etc.) or diamond-like carbon layers (DLC).

[0106] Each amperometric sensor 134a-b here comprises a working electrode 1341a-b, a reference electrode 1342a-b and a counter electrode 1343a-b, for example platinum-based.

[0107] A reference electrode is defined as an electrode that provides a stable and constant potential, this potential serving as a reference point for measuring the potential of a working electrode. The reference electrode is made of a material such as silver (Ag) or silver chloride (AgCl).

[0108] A working electrode is defined as an electrode at which an electrochemical reaction of interest takes place. The potential of the working electrode is measured relative to the reference electrode, and the current flowing through it is measured using an ammeter. The working electrode is typically the electrode at which the analyte is oxidized or reduced.

[0109] The term "counter electrode" refers to an electrode that complements the amperometric sensor. In a particular embodiment of the invention, the counter electrode is connected to the working electrode by means of an ammeter, and the counter electrode carries an electric current in the opposite direction to that of the working electrode. The counter electrode is made from a material inert to the electrochemical reaction, such as platinum or graphite.

[0110] The counter electrode 1343 ab is dimensioned with a larger surface area than the working electrode 1341a-b. As explained previously, the main role of the counter electrode 1343 ab is to allow current to flow through it, and not through the reference electrode 1342a-b. The counter electrode 1343 ab thus limits current leakage to the other electrodes. Par43825

[0111] 15

[0112] elsewhere, the reference electrodes 1312a-b, 1322a-b and 1342a-b advantageously have equal respective surfaces so that one of said electrodes does not prevail over others, which could lead to measurement errors.

[0113] In a particular embodiment of the invention, the amperometric sensors 134a and 134b are intended to measure an oxygen level in solution 300.

[0114] The platinum resistance thermometer 135, on the other hand, is preferably a PT100 probe.

[0115] Figure 5B shows conductive tracks 119 of the printed circuit board.

[0116] According to one embodiment, the sensor assembly 13 can be fabricated by screen printing to produce low-cost, miniaturized electrochemical sensors using a well-established industrial process. As an alternative to the specific example shown, other sensor integration methods can be used. For example, sensors with electrodes in a glass tube can be employed.

[0117] Furthermore, the examples of electrochemical quantities and concentrations given in this embodiment are not exhaustive. Other types of sensors can be used to measure the concentration of glucose, potassium, or hormones in solution 300.

[0118] The arrangement of sensors shown in Figure 5A and Figure 5B is a non-limiting example.

[0119] Figure 5C represents the first face lia, according to a second embodiment, where we find conductive tracks and electrodes corresponding to the sensors just described.

[0120] One of the criteria for choosing the arrangement of sensors relative to each other is to avoid the risk of electrical interference. Therefore, it is preferable for the electrodes of the different sensors to be sufficiently far apart to avoid distorting the measurements. Within a single sensor, for example, an inter-electrode distance of between 1 mm and 5 mm is chosen, while between two different sensors, a greater inter-electrode distance is preferred, which could be, for example, on the order of a centimeter to limit this interference.

[0121] An alternative solution to these embodiments, shown in Figure 5D, is to position at least one block 14 of electrically insulating material, for example polymer, between different sensors.43825

[0122] 16

[0123] Another alternative solution (not shown here) is to position one or more sensors on one face of a support and one or more other sensors on an opposite face of the same support. For example, the OCP sensors 131a-b and 132a-b are thus arranged on a first face 1a of the sensor support 11, while the EIS sensors 133a-b and the amperometric sensors 134a-b are arranged, for example, on a second face opposite the first face.

[0124] Figure 6 represents a perspective view of the implementation of the electrical connection means 111 between the sensor support 11 and the electronic device 10 of the electro-chemical measuring device 100.

[0125] As mentioned previously, the electrical connection between the sensor support 11 and the electronic device 10 can be ensured by a high-density electrical connection means 111, comprising, for example, about twenty connections, these connections being, for example, gold-plated. Furthermore, the electrical connection is itself ensured at the end 12 of the sensor support by adding supplementary tracks 121 that are sufficiently wide and exhibit good mechanical resistance to the regular and repetitive connection and disconnection of the electrochemical measurement device 100, said tracks preferably being gold-plated themselves.

[0126] Figure 7A, Figure 7B and Figure 7C represent different examples of assembly modes between a probe housing head 20 and a probe housing body 21 in separable elements as described previously.

[0127] The mechanical connection can be achieved here by means of two complementary coupling elements that cooperate to create a removable assembly between the probe housing head 20 and the probe housing body 21. These elements can, for example, be in the form of:

[0128] - a retaining mechanism located on the head of the probe housing 20; and

[0129] - of a stop located on the body of the probe housing 21.

[0130] This mechanical support is preferably implemented without requiring the use of tools.

[0131] Furthermore, the mechanical retention provides sufficient resistance to pull-out to ensure the re-establishment of contact between all sensitive elements and also to guarantee the preservation of the sterility of the portion of the probe located inside chamber 50 of the bioreactor 500.43825

[0132] 17

[0133] For each embodiment, the cooperation between the probe housing head 20 and the probe housing body 21 can be facilitated by means of a groove or keying feature and a linear guide system (not shown here) to allow correct positioning and ensure a preferably 0° inclination between the electrical connection means 111 and the complementary tracks 121 so that there are no mechanical stresses during regular and repetitive connections and disconnections of the electrochemical measurement device 100.

[0134] Preferably, the guide system has a length of at least 16 mm.

[0135] In the embodiment shown in Figure 7A, the mechanical connection is achieved by a retaining mechanism 201a comprising a plurality of spring-loaded plungers that fit into a chamfered stop 211a. For example, the spring-loaded plungers of the retaining mechanism 201a have a retaining force of between 300 g and 500 g to ensure easy insertion of the probe housing body 21 into the probe housing head 20, while also providing the necessary support between said body and said head for measurements and during handling of the bioreactor 500. During insertion, the orientation of the probe housing body 21 within the probe housing head 20 is achieved by means of a pin-end screw 20 Id fixed in said head and adapted to slide in a groove (not shown here), said groove being formed along a portion of said body that is inserted into said head.The combination of the pin-end screw 20 Id with the groove then prevents rotation between the probe housing head 20 and the probe housing body 21.

[0136] In the embodiment shown in Figure 7B, the mechanical connection is made by means of buttons 201b which can be moved in translation and which, when pressure is applied, release a flexible attachment structure 211b, for example in the form of a hook, said attachment structure being housed in a plurality of cavities 202b when the probe housing body 21 is inserted into the probe body head 20.As with the embodiment shown in Figure 7A, during insertion, the orientation of the probe housing body 21 in the probe housing head 20 is achieved by means of a pin-end screw (not shown here) fixed in said head and being able to slide in a groove (not shown here), said groove being made along a part of said body which is inserted into said head, this combination of the pin-end screw 20 Id with the groove preventing rotation between the probe housing head 20 and the probe housing body 21.43825.

[0137] 18

[0138] In the embodiment shown in Figure 7C, the removable assembly is achieved by means of a screw 201c, for example a set screw with a pointed tip, which fits into a stop 211c having the shape of a chamfer. The clamping force of the screw 201c ensures that the probe housing head 20 is held securely to the probe housing body 21. As before, during insertion, the orientation of the probe body 21 within the probe housing head 20 is achieved by means of a pin-end screw 201e fixed in said head and adapted to slide in a groove 201f, said groove being formed along a portion of said body which is inserted into said head. This combination of the pin-end screw 201e with the groove prevents rotation between the probe housing head 20 and the probe housing body 21.

[0139] An electrochemical device equipped with a set of several closely spaced sensors, preferably arranged on the same support, requires special control of the polarization of the electrodes of the different sensors in order to both limit interference and limit the total measurement time.

[0140] An example of the implementation of such control will now be described in connection with figures 8A-8C as well as figure 9.

[0141] Figure 8A, Figure 8B and Figure 8C represent respectively an example of an amperometric measurement module, an OCP measurement module and an EIS measurement module, of the electro-chemical measurement device 100. These modules are obtained by combining stages 1030A, 1030B, 1030C of the electronic device 10 formed for example in particular by the system on chip 103 described previously with sensors such as those mentioned above.

[0142] Polarization signals Spoln, Spol, Spoh, Spobi, Spob2 from the electrodes are transmitted here via digital-to-analog converters, DACs, 1031, 1033 and 1035, respectively connected to the amperometric sensor 134, the OCP sensor 131 and the EIS sensor 133. A plurality of analog-to-digital converters, ADCs, 1032, 1034 and 1036, respectively connected to the amperometric sensor 134, the OCP sensor 131 and the EIS sensor 133, are intended to receive analog output signals from said sensors 134, 131, 133.

[0143] To adapt the input and output signals, a plurality of amplifiers, in particular operational amplifiers 1041, 1042, 1061, 1062, 1063, 1045, 1046, are used here.43825

[0144] 19

[0145] A VamperoWE bias potential, dedicated to the biasing of the electrodes of amperometric sensors, is used here. In one particular embodiment, the VamperoWE bias potential is equal to 0.6 V.

[0146] Figure 8A illustrates the connection of an amperometric sensor 134 to a stage 1030a of the electronic device 10, which is dedicated to controlling the polarization of the electrodes of the amperometric sensor 134 and processing the signals from this sensor in order to perform an amperometric measurement. Thus, at the output of the converter 1031, the biasing potential VamperoWE is applied to permanently bias the working electrode 1341 of the amperometric sensor 134, while a second biasing potential VamperoRE is applied to bias the reference electrode 1342, and a third biasing potential VamperoCE is applied to permanently bias the counter electrode 1343, both during and outside the amperometric measurement phases. The working electrode 1341 is connected to the inverting input of a transimpedance amplifier 1042, as well as to the output of this amplifier.The non-inverting input of the transimpedance amplifier 1042 is connected to the DAC 1031, the input to which the biasing potential VamperoWE is applied. The potential of the working electrode 1341 and the potential of the reference electrode 1342 are maintained at VamperoWE and VamperoRE respectively during and outside of the amperometric measurement phases, and in particular during measurement phases performed on the other sensor(s) to maintain the ionic stability necessary for amperometric measurement during the various measurements, the ionic stabilization time being long, from thirty seconds to several minutes in the case of amperometric measurements.During and outside the amperometric measurement phases, a voltage differential Vbias of a few hundred mV (particularly -400 mV) is applied, such that VamperoRE = VamperoWE - Vbias, in order to generate a current between the working electrode 1341 and the counter electrode 1343 in order to carry out the amperometric measurements.

[0147] At the output of the transimpedance amplifier 1042, we obtain a voltage V1042 which is notably a function of the current generated according to the oxygen concentration in the solution 300. The voltage V1042 is then transmitted to the processing system on chip 103 by means of the CAN 1032.

[0148] Figure 8B illustrates the connection of an OCP 131 sensor to a 1030b stage of the electronic device 10, which is dedicated in particular to the polarization control of the 43825

[0149] 20

[0150] electrodes of sensor 131 and signal processing from this sensor 131 in order to perform an OCP measurement. Thus, at the output of converter 1033, a bias potential VpolOCP, corresponding to the bias potential VamperoCE of the counter electrode 1343 of the amperometric sensor 134, is applied to the non-inverting input of an operational amplifier 1061 here mounted as a follower or functioning as a "buffer". The inverting input and output of amplifier 1061 are connected to the reference electrode 1312 of the OCP sensor 131. Here, at least for the duration of an OCP measurement phase, the image polarization potential of VamperoCE is applied to the reference electrode 1312 of the OCP sensor 131 in order to limit any interference due to the passage of current linked to the permanent polarization of the working electrode 1341 and the reference electrode 1342 of the amperometric sensor 134.In this example, the working electrode 1311 of the OCP sensor is connected to the non-inverting input of a second operational amplifier 1062, mounted as a follower, and whose inverting input and output of the amplifier 1062 are connected to an ADC 1034.

[0151] At the output of amplifiers 1062 and 1063, a potential difference voltage Vio63 is obtained which is dependent on a molecular concentration f(x), for example of CO2, or on a pH level f(x) in solution 300. Advantageously, the working electrode 1311 is placed in high impedance, for example on the order of 50 MOhm, to limit any current that could pass through the working electrode 1311. In a particular embodiment of the invention, and outside the OCP measurement phases, the OCP sensor 131 is not polarized and the working electrode 1311 as well as the reference electrode 1312 can be left floating.

[0152] In another particular embodiment of the invention, and outside the OCP measurement phases, the OCP sensor 131 is polarized by maintaining a VpolOCP polarization on the reference electrode 1312.

[0153] Figure 8C is used to illustrate the connection of an EIS 133 sensor to a stage 1030c of the electronic device 10 dedicated in particular to the control of the polarization of the electrodes of the EIS 133 sensor and to the processing of signals from this sensor 133 in order to carry out an "EIS" measurement.

[0154] At the output of the CNA 1035, a polarization potential VpolEIS, as close as possible to VamperoCE, is applied to polarize the two working electrodes 1331, 1332. During the EIS measurement phase, 133143825 is superimposed on the first working electrode.

[0155] 21

[0156] a sinusoidal excitation signal of the "sliding sine" type defined by the function sin(cot), with co = 2nf, the angular frequency of the signal, and fia the frequency of the signal between 20 kHz and 200 kHz, with a DC component equal to VpolEIS + Voffset, with Voffset a DC voltage between -1000 mV and +1000 mV, preferably zero.

[0157] In a particular embodiment of the invention, where the DC offset voltage Voffset is not zero, the electrochemical measuring device 100 is suitable for use, throughout the monitoring of the bioproduction of solution 300 or, alternatively, on an ad hoc basis, to perform studies of corrosion phenomena as well as studies of reactions on the surfaces of the working electrodes 1331 and 1332, such as can be observed during electrodeposition or electropolymerization processes. A feedback resistor 1047 is inserted between the inverting input and the output of the transimpedance amplifier 1046.

[0158] At the output of the 1046 transimpedance amplifier, a voltage V1046 is obtained, which is a function of the impedance and conductivity of the medium at frequencies f between 20 kHz and 200 kHz. The voltage V1046 is defined by the expression

[0159] Vio46 = B x sin(cot + 0), where B is an amplitude parameter and 0 is the phase shift observed during the measurement. The voltage V1046 is then transmitted to the system-on-chip 103 via the ADC 1036.

[0160] At the end of the EIS measurement phase, the sinusoidal excitation signal is stopped so as not to interfere with the following amperometric measurement.

[0161] In a particular embodiment of the invention, and outside of the EIS measurement phases, the working electrodes 1331, 1332 are not polarized and can be left floating.

[0162] In another particular embodiment of the invention, and outside the EIS measurement phases, the working electrodes 1331, 1332 are polarized at the polarization potential VpolEIS.

[0163] Advantageously, in a particular embodiment of the invention, when the OCP 131 sensors are doubled, the OCP measurement phases are carried out in parallel. Furthermore, in a particular embodiment of the invention, the OCP and EIS measurement phases are carried out in parallel, provided that the sensors 131 and 133 are sufficiently spaced so as not to create interference between the measurements, i.e., that a space of several millimeters, for example 5 mm, separates said sensors. 43825

[0164] 22

[0165] Figure 9 represents the different stages of a generic example of a 900 measurement sequence using an electro-chemical measurement device 100 as described previously.

[0166] First, during a preliminary initialization phase 910, the bias potential VamperoRE is applied to the reference electrodes 1342a-b, VamperoWE to the working electrodes 1341a-b and VamperoCE to the counter electrodes 1343a-b of the amperometric sensors 134a-b.

[0167] Next, during a measurement phase, the steps described below are carried out.

[0168] During an amperometric measurement phase 920, the bias potentials previously applied at the level of the amperometric sensor 134 are maintained, i.e. with the voltage differential Vbias between the working electrode 1341 and the reference electrode 1342 without biasing the electrodes of the other sensors 133, a phase during which measurement signals from the amperometric sensor 134 are acquired.

[0169] During an OCP 930 measurement phase, the image polarization potential of VamperoCE is applied to the reference electrode 1312 of the OCP sensor 131, during a measurement phase of a chemical quantity or the concentration of a molecule in the solution 300, while maintaining the VamperoWE polarization potential on the working electrode 1341, the VamperoRE polarization potential on the reference electrode 1342, the VamperoCE polarization potential on the counter electrode 1343 of the amperometric sensor 134 being then induced by the VamperoWE and VamperoRE polarization potentials, in combination with the electrochemical reaction occurring on the working electrode 1341, of the amperometric sensor 134.

[0170] In a particular embodiment of the invention, the VamperoCE polarization potential is first measured, with a margin of error between -100 mV and +100 mV, by means of an analog or digital follower setup, then the measured VamperoCE polarization potential is applied to the reference electrode 1312 of the OCP sensor 131, during a measurement phase of a chemical quantity or of the concentration of a molecule in the solution 300.

[0171] In a particular embodiment of the invention, and outside the OCP 930 measurement phase, the reference electrode 1312 of the OCP 131 sensor is not polarized and can be left floating, while maintaining the VamperoWE polarization potential on the working electrode 1341, the VamperoRE polarization potential on the electrode 43825

[0172] 23

[0173] reference 1342, the VamperoCE polarization potential on the counter electrode 1343 of the amperometric sensor 134 is then induced by the VamperoWE and VamperoRE polarization potentials, in combination with the electrochemical reaction occurring on the working electrode 1341 of the amperometric sensor 134.

[0174] Advantageously, interrupting the polarization of the OCP sensor 131 slows down the aging of the reference electrode 1312 and the working electrode 1311 because no current flows through said electrodes.

[0175] In another particular embodiment of the invention, and outside the OCP measurement phase, the working electrode 1311 and the reference electrode 1312 of the OCP sensor 131 are polarized, while maintaining the VamperoWE polarization potential on the working electrode 1341 and the VamperoRE polarization potential on the reference electrode 1342 of the amperometric sensor 134, the VamperoCE polarization potential on the counter electrode 1343 of the amperometric sensor 134 then being induced by the VamperoWE and VamperoRE polarization potentials, in combination with the electrochemical reaction occurring on the working electrode 1341, of the amperometric sensor 134.

[0176] Advantageously, maintaining the polarization of the OCP 131 sensor reduces noise during measurements that are carried out during the OCP measurement phase.

[0177] During an EIS 940 measurement phase, the biasing potential is applied as close as possible to the VamperoCE biasing potential on the first working electrode 1331 as well as on the second working electrode 1332 of the EIS sensor 133, while the excitation signal is applied to the first working electrode 1331 of the EIS sensor 133, during an electrochemical impedance spectroscopic measurement phase in solution 300, while maintaining the VamperoWE biasing potential on the working electrode 1341, the VamperoRE biasing potential on the reference electrode 1342, the VamperoCE biasing potential on the counter electrode 1343 of the amperometric sensor 134 is then induced by the VamperoWE and VamperoRE biasing potentials, in combination with the electrochemical reaction occurring on the working electrode 1341, of the amperometric sensor 134,then to interrupt the polarization of the working electrodes 1331 and 1332 of the EIS sensor 133, the VamperoWE polarization potential on the working electrode 1341, the VamperoRE polarization potential on the reference electrode 1342, and the VamperoCE polarization potential on the counter electrode 1343 of the amperometric sensor 43825,

[0178] 24

[0179] 134 is then induced by the biasing potentials VamperoWE and VamperoRE, in combination with the electrochemical reaction occurring on the working electrode 1341, of the amperometric sensor 134.

[0180] In a particular embodiment of the invention, and outside the EIS 940 measurement phase, the working electrodes 1331, 1332 are not polarized and can be left floating.

[0181] In another particular embodiment of the invention, and outside the EIS 940 measurement phase, the working electrodes 1331, 1332 are polarized at the VpolEIS polarization potential.

[0182] The phases 920 to 940 described above are then repeated a number k times.

[0183] In a particular embodiment of the invention not shown here, the EIS 940 measurement phase is carried out following the amperometric measurement phase 920, the EIS 940 measurement phase then being followed by the OCP 930 measurement phase.

[0184] In another particular embodiment of the invention not shown here, the OCP 930 measurement phase and the EIS 940 measurement phase are carried out at the same time, and not sequentially.

[0185] An example of a specific sequencing of the type described above will now be given. Following a preliminary initialization phase, the VamperoRE polarization potential is applied to the reference electrodes 1342a and 1342b, and VamperoWE to the working electrodes 1341a and 1341b of the amperometric sensors 134a and 134b. These potentials are maintained throughout the bioproduction monitoring.

[0186] The following measurement sequence is then performed.

[0187] During an amperometric measurement phase of a duration for example of the order of 100 ms, the bias potential VamperoWE is maintained on the working electrodes 1341a and 1341b of the amperometric sensors 134a and 134b and the voltage differential Vbias is applied between the working electrodes 1341a and 1342b and the reference electrodes 1342a and 1342b without biasing the electrodes of the other sensors 131a-b, 132a-b and 133a-b, and measurement signals are acquired from the amperometric sensors 134a and 134b.

[0188] Then, a first pause of approximately 100 ms is performed to allow solution 300 to return to a so-called "resting" state. Next, during a measurement phase of the carbon dioxide dissolved in solution 300, the VamperoCE image polarization potential is applied to the reference electrodes 1312a and 1312b of the OCP sensors 131a and 131b, while maintaining the VamperoWE polarization potential on the working electrodes 1341a and 1341b, the VamperoRE polarization potential on the reference electrodes 1342a and 1342b, and the VamperoCE polarization potential on the counter electrodes 1343a and 1343b. This polarization potential is induced by the VamperoWE and VamperoRE polarization potentials, in combination with the electrochemical reaction on the working electrodes 1341a and 1342b. 1341b, amperometric sensors 134a and 134b;

[0189] Then, the polarization of the reference electrodes 1312a and 1312b of the OCP sensors 131a and 131b is interrupted or not, while maintaining the VamperoWE polarization potential on the working electrodes 1341a and 1341b, the VamperoRE polarization potential on the reference electrodes 1342a and 1342b, the VamperoCE polarization potential on the counter electrodes 1343a and 1343b being then induced by the VamperoWE and VamperoRE polarization potentials, in combination with the electrochemical reaction on the working electrodes 1341a and 1341b, of the amperometric sensors 134a and 134b.

[0190] A second pause of approximately 100 ms is then performed so that solution 300 returns to a so-called "resting" state;

[0191] Then, during a phase of measuring the hydrogen potential of solution 300, the image polarization potential of VamperoCE is applied to the reference electrodes 1322a and 1322b of the OCP sensors 132a and 132b, while maintaining the VamperoWE polarization potential on the working electrodes 1341a and 1341b, the VamperoRE polarization potential on the reference electrodes 1342a and 1342b, the VamperoCE polarization potential on the counter electrodes 1343a and 1343b being then induced by the VamperoWE and VamperoRE polarization potentials, in combination with the electrochemical reaction on the working electrodes 1341a and 1341b, of the amperometric sensors 134a and 134b;

[0192] Then, the polarization of the reference electrodes 1322a and 1322b of the OCP sensors 132a and 132b is interrupted or not, while maintaining the depolarization potential VamperoWE on the working electrodes 1341a and 1341b, the polarization potential VamperoRE on the reference electrodes 1342a and 1342b, the polarization potential VamperoCE on the counter electrodes 1343a and 1343b being then induced by the polarization potentials VamperoWE and VamperoRE, in combination with the electrochemical reaction on the working electrodes 1341a and 1341b, of the amperometric sensors 134a and 134b;

[0193] A third pause is then performed, for example for a duration of around 100 ms, so that solution 300 returns to a so-called "resting" state;Then, during a phase of electrochemical impedance spectroscopic measurement in solution 300, the image polarization potential of VamperoCE is applied to the working electrodes 1331a, 1332a, 1331b, and 1332b of the EIS sensors 133a and 133b, while superimposing the excitation signal on the first working electrode 1331a of the EIS sensor 133a and on the first working electrode 1331b of the EIS sensor 133b, while maintaining the VamperoWE polarization potential on the working electrodes 1341a and 1341b, the VamperoRE polarization potential on the reference electrodes 1342a and 1342b, and the VamperoCE polarization potential on the counter electrodes 1343a and 1343b, the latter being induced by the potentials of VamperoWE and VamperoRE polarization, in combination with the electrochemical reaction on the working electrodes 1341a and 1341b, of the amperometric sensors 134a and 134b;

[0194] Then, the polarization of the working electrodes 1331a, 1332a, 1331b and 1332b of the EIS sensors 133a and 133b is interrupted or not, while maintaining the VamperoWE polarization potential on the working electrodes 1341a and 1341b, the VamperoRE polarization potential on the reference electrodes 1342a and 1342b, the VamperoCE polarization potential on the counter electrodes 1343a and 1343b being then induced by the VamperoWE and VamperoRE polarization potentials, in combination with the electrochemical reaction on the working electrodes 1341a and 1341b, of the amperometric sensors 134a and 134b;

[0195] A fourth pause of approximately 100 ms is then performed in the EIS measurement phase so that solution 300 returns to a so-called "resting" state.27

[0196] Such steps are then repeated a number k times, with k an integer which can be, for example, on the order of 300000, as can be the case in the context of a CHO production carried out over a period of 21 days, and at a rate of one measurement cycle carried out continuously and every second.

[0197] Figure 10 serves to illustrate schematically one way of polarizing all the electrodes of the sensor 13, according to an embodiment as described below in connection with the timing diagrams 700a and 700b of Figures 11 A and Figure 1 IB.

[0198] Figure 10 shows how and what the polarization potentials are applied to the amperometric sensors 134, OCP 131 and 132 and EIS 133.

[0199] Similarly, Figure 10 shows that the electronic device 10 includes buffer amplifiers 1037, 1038 and 1039 to couple the bias voltage of the counter electrode 1343 of the amperometric sensor 134, respectively to the reference electrode 1312 of the OCP sensor 131, to the reference electrode 1322 of the OCP sensor 132 and to the working electrodes 1331 and 1332 of the EIS sensor 133.

[0200] Furthermore, the amperometric sensors 134, OCP 131 and OCP 132 are connected to a first potentiostat (not shown here), and the EIS sensor 133 is connected to a second potentiostat (not shown here).

[0201] The amperometric sensor is used to measure the current flowing through solution 300 between the counter electrode 1343 and the working electrode 1341.

[0202] OCP sensors 131 and 132 are used to measure the DC voltage between the reference electrode 1312 and 1322, and the working electrode 1311 and 1321.

[0203] The EIS 73 sensor is used to measure the alternating voltage and current between the working electrodes 1331 and 1332.

[0204] Figure 11 A and Figure 1 IB show timing diagrams 700a and 700b of the polarization and measurement phases for two embodiments of the invention. Timing diagram 700a shows how amperometric measurement phases 74a, OCP measurement phases 71a, 72a, and EIS measurement phases follow one another, and how the amperometric sensor 134, the OCP sensors 131 and 132, and the EIS sensor 133 are polarized during these measurement phases.

[0205] Prior to a sequence of measurements, a VamperoWE polarization potential is first applied to the working electrode 1341, then a potential of 43825

[0206] 28

[0207] VamperoRE polarization on the reference electrode 1342, with VamperoRE = VamperoWE - Vbias.

[0208] We then carry out a first amperometric measurement phase 74a, for example with a duration of 50 ms, without this being a limit.

[0209] At the end of the first amperometric measurement phase 74a, and after a pause (not shown here), the first OCP measurement phase 71a is performed. The duration of this phase can be equal to that of the amperometric measurement phase 74a, for example, 50 ms, without this being a limit. The biasing of the OCP sensor begins here before the first amperometric measurement phase 74a. The biasing potential VpolOCP is applied to the reference electrode 1311 of the OCP sensor 131, as well as to the reference electrode 1321 of the OCP sensor 132, before the first OCP measurement phase and maintained during and after the first OCP measurement phase throughout the entire measurement sequence.

[0210] At the end of the first OCP 72a measurement phase, and after a pause (not shown here), a second OCP 72a measurement phase is performed. This second phase can be of equal duration to the first OCP measurement phase, for example, 50 ms. After the second OCP 72a measurement phase, and a pause (not shown here), the first EIS 73a measurement phase is performed. It typically has a longer duration than the amperometric and OCP measurement phases. For example, the EIS measurement phase can have a duration of approximately 150 ms, although this is not a limit. During the EIS measurement phase, the polarization potential VpolEIS is used to polarize the working electrodes 1331 and 1332 of the EIS sensor 133, at least until the end of the first EIS 73a measurement sequence. The polarization of the EIS sensor(s) is not maintained here during the OCP and amperometric measurement phases.

[0211] Amperometric measurement phases 74a, OCP 71a and 72a, as well as EIS 73a are repeated a number k times, with k an integer, during a measurement sequence.

[0212] In the embodiment shown in Figure 11A, the polarization of the amperometric sensor 134 is maintained during the amperometric measurement phases 74a and the EIS and OCP measurement phases. Similarly, the OCP sensors 131 and 132 are polarized outside of the OCP measurement sequences 71a and 72a and during the other amperometric measurement phases 74a and EIS.43825

[0213] 29

[0214] Chronogram 700b represents another way of polarizing the electrodes, particularly those of the OCP sensor(s) and the EIS sensor(s). In this embodiment, and as in the previous example, a VamperoWE polarization potential is applied to the working electrode 1341 and a VamperoRE polarization potential to the reference electrode 1342, prior to the measurement sequence.

[0215] Next, this sequence of measurements starts with a first amperometric measurement phase 74b, while maintaining the polarization of the working electrodes 1341 and reference electrode 1342 of the amperometric sensor.

[0216] Following the end of the first amperometric measurement phase 74b, the first OCP measurement phase 71b is carried out while maintaining the polarization of the working electrodes 1341 and reference electrodes 1342 of the amperometric sensor.

[0217] The polarization potential VpolOCP is applied to the reference electrode 1311 of the OCP 131 sensor, during the OCP 71b measurement phases but not during the amperometric or EIS measurement phases.

[0218] At the end of the first measurement phase OCP 71b, and after a pause (not shown here), the second measurement phase OCP 72b is performed. Here too, the VpolOCP polarization potential is applied to the OCP 132 sensor, but only during the measurement phases carried out on this sensor.

[0219] At the end of the second OCP 72b measurement phase, and after a pause (not shown here), the first EIS 73b measurement phase is performed. A VpolEIS bias potential is applied to the working electrodes 1331 and 1332 of the EIS 133 sensor during the measurement phases on this sensor, but not during the amperometric and OCP measurement phases, in order to avoid generating parasitic currents during these amperometric and OCP measurement phases.

[0220] The amperometric measurement phases 74b, OCP 71b and 72b, as well as EIS 73b are repeated a number k times, during a measurement sequence.

[0221] In the embodiment shown in Figure 1 IB, only the amperometric sensor 134 is biased outside of the amperometric measurement phases 74b. In other words, the OCP sensors 131 and 132 are not biased outside of the OCP measurement phases 71a and 72b, and the EIS sensor 133 is not biased outside of the EIS measurement phase 73b.

[0222] 30

[0223] As a reminder, the amperometric measurement phases 74a-b consist of measuring the current flowing through the solution 300 between the counter electrode 1343 and the working electrode 1341.

[0224] The OCP measurement phases 71a-b and 72a-b consist of measuring the DC voltage between the reference electrode 1312 and 1322, and the working electrode 1311 and 1321 of the OCP sensors 131 and 132.

[0225] The EIS 73a-b measurement phases consist of measuring the alternating voltage and current between the working electrodes 1331 and 1332 of the EIS sensor 133. Advantageously, the amperometric, OCP and EIS measurement phases can be carried out in parallel and not sequentially in an embodiment (not shown here) in which the electronic device 10 includes at least two control circuits 103 for the polarization of the electrodes, each control circuit 103 independently polarizing the sensors 131a-b, 132a-b, 133a-b and 134a-b of the sensor set 13.

[0226] Advantageously, by means of the interlocking of sensors 131a-b, 132a-b, 133a-b and 134a-b, it is capable of carrying out real-time, or near-real-time, monitoring of the bioproduction of solution 300, by measuring different electro-chemical quantities as well as different concentrations of molecules, the duration of the complete measurement sequence as described being on the order of 800 ms.

[0227] Furthermore, the invention makes it possible to eliminate the influence of amperometric measurements on OCP or EIS measurements in the context of a multimodal measurement.

Claims

43825 31 DEMANDS 1. Electrochemical measuring device (100) for measuring physicochemical quantities and / or concentrations of molecules in a solution (300), the electrochemical measuring device comprising: - an assembly (13) of sensors arranged on a support (11), said assembly (13) comprising at least one amperometric sensor (134, 134a, 134b) and at least one "OCP" sensor for open-circuit potential measurements (131, 131a, 131b, 132, 132a, 132b), the amperometric sensor (134, 134a, 134b) comprising a working electrode (1341, 1341a, 1341b), a reference electrode (1342, 1342a, 1342b) and a counter electrode (1343, 1343a, 1343b) and the OCP sensor comprising a reference electrode (1312, 1312a, 1312b, 1322, 1322a, 1322b) and a working electrode (1311, 1311a, 1311b, 1321, 1321a, 1321b); - an electronic device (10) for the acquisition and processing of measurement signals from said sensors, the electronic device (10) further comprising at least one control circuit (103) for the polarization of the electrodes of said sensors, the control circuit (103) being configured to: according to a phase prior to a measurement sequence, apply to the amperometric sensor (134, 134a, 134b): a VamperoWE polarization potential on the working electrode (1341, 1341a, 1341b); a bias potential VamperoRE on the reference electrode (1342, 1342a, 1342b), with VamperoRE = VamperoWE - Vbias where Vbias is a voltage differential between the working electrode (1341, 1341a, 1341b) and the reference electrode (1342, 1342a, 1342b); then, perform k times (where k is a non-zero integer) a sequence of measurements comprising steps consisting of: during an amperometric measurement phase, acquire measurement signals from the amperometric sensor (134, 134a, 134b) while maintaining the VamperoWE polarization potential on the working electrode (1341, 1341a, 1341b) and maintaining the VamperoRE polarization potential on the reference electrode (1342, 1342a, 1342b); and during an OCP measurement phase, acquire measurement signals from the OCP sensor (131, 131a, 131b, 132, 132a, 132b) while biasing the OCP sensor (131, 131a, 131b, 132, 132a, 132b) by applying a biasing potential VpolOCP, dependent on the biasing potential VamperoCE of the counter electrode (1343, 1343a, 1343b) of the amperometric sensor (134, 134a, 134b), on the reference electrode (1312, 1312a, 1312b, 1322, 1322a, 1322b) of the OCP sensor (131, 131a, 131b, 132, 132a, 132b), the VamperoRE polarization potential on the reference electrode (1342, 1342a, 1342b) and the VamperoWE polarization potential on the working electrode (1341, 1341a, 1341b) of the amperometric sensor (134, 134a, 134b), being maintained during the measurement phase on the OCP sensor, in which the counter electrode (1343, 1343a, 1343b) of the amperometric sensor (134, 134a, 134b) is coupled to the reference electrode (1312, 1312a, 1312b, 1322, 1322a, 1322b) of the OCP sensor (131, 131a, 131b, 132, 132a, 132b) and / or in in which the bias potential VpolOCP is substantially equal to the bias potential VamperoCE of the counter electrode (1343, 1343a, 1343b) of the amperometric sensor (134, 134a, 134b), or to a measured value Vmes from the counter electrode (1343, 1343a, 1343b) and close to VamperoCE, with Vmes = VamperoCE + Vdelta, where Vdelta is a voltage differential between -100 mV and +100 mV.

2. Device according to claim 1, wherein the sensor assembly (13) further comprises at least one EIS sensor for electrochemical impedance spectroscopic measurements (133, 133a, 133b), the EIS sensor having a first working electrode (1331, 1331a, 1331b) and a second working electrode (1332, 1332a, 1332b), the control circuit (103) being configured to, during said sequence and during an EIS measurement phase during which measurement signals from the EIS sensor (133, 133a, 133b) are acquired: apply a bias potential VpolEIS to the second working electrode (1332, 1332a, 1332b) of the EIS sensor (133, 133a, 133b), and apply to the first electrode work (1331, 1331a, 1331b) of the EIS sensor (133, 133a, 133b) a biasing potential equal to the sum of a DC component equal to VpolEIS + Voffset, with Voffset a DC voltage between -1000 mV and +1000 mV, preferably zero,and an alternative component, while maintaining the VamperoRE polarization potential on the reference electrode (1342, 1342a, 1342b) and maintaining the VamperoWE polarization potential on the working electrode (1341, 1341a, 1341b) of the amperometric sensor.

3. Device according to claim 2, wherein the polarization potential VpolEIS is equal to the polarization potential VamperoCE of the counter electrode (1343, 1343a, 1343b) of the amperometric sensor (134, 134a, 134b) or equal to the polarization potential VpolOCP of the reference electrode (1312, 1312a, 1312b, 1322, 1322a, 1322b) of the OCP sensor (131, 131a, 131b, 132, 132a, 132b), or equal to the higher potential between said potential of the counter electrode (1343, 1343a, 1343b) of the amperometric sensor (134, 134a, 134b) and said potential of the reference electrode (1312, 1312a, 1312b, 1322, 1322a, 1322b) of the OCP sensor (131, 131a, 131b, 132, 132a, 132b).

4. Device according to any one of claims 2 or 3, wherein, after the EIS measurement phase and prior to another measurement phase, the polarization of the first working electrode (1331, 1331a, 1331b) of the EIS sensor (133, 133a, 133b), and the polarization of the second working electrode (1332, 1332a, 1332b) of the EIS sensor (133, 133a, 133b), are interrupted.

5. Device according to any one of the preceding claims, wherein the VamperoRE polarization potential on the reference electrode (1342, 1342a, 1342b), the VamperoWE polarization potential on the working electrode (1341, 1341a, 1341b) and the VamperoCE polarization potential on the counter electrode (1343, 1343a, 1343b) of the amperometric sensor (134, 134a, 134b) are maintained throughout the measurement sequence.

6. Device according to any one of the preceding claims, wherein the sensor assembly (13) comprises one or more amperometric sensors (134, 134a, 134b) for measuring an oxygen level, at least one pair of OCP sensors (131, 131a, 131b) for measuring a carbon dioxide level and / or for measuring a hydrogen potential, and at least two EIS sensors (133, 133a, 133b) for measuring a conductivity and impedance of the solution (300).

7. Device according to any one of the preceding claims, said at least one amperometric sensor (134, 134a, 134b) and said at least one OCP sensor (131, 131a, 131b, 132, 132a, 132b) comprising reference electrodes (1342, 1342a, 1342b, 1311, 1311a, 1311b, 1322, 1322a, 1322b) of equal respective areas.

8. Device according to any one of the preceding claims, wherein said support (11) is a printed circuit board, a block (14) of electrical insulating material being disposed on one face of the printed circuit board between the electrodes of the second sensor (131, 131a, 131b, 132, 132a, 132b) and the electrodes of the amperometric sensor (134, 134a, 134b).

9. Device according to any one of the preceding claims, wherein said support (11) is a printed circuit board having a first face (Ha) and a second face opposite the first face (Ha), the electrodes of the OCP sensor (131, 131a, 131b, 132, 132a, 132b) being distributed on the first face (11a), the electrodes of the amperometric sensor (134, 134a, 134b) being arranged on the second face and when the device is equipped with an EIS sensor (133, 133a, 133b), the electrodes of said EIS sensor being arranged on the second face (11b).

10. Device according to any one of the preceding claims, wherein said support (11) is in particular a printed circuit board and wherein said electronic device (10) is arranged on a second support distinct from said support, in particular a second printed circuit board distinct from the first board.

11. Electrochemical measuring probe (200) for measuring physicochemical quantities and concentrations of molecules in a solution (300) comprising a device according to any one of the preceding claims, said probe comprising: - a probe housing head (20) intended to be placed outside the solution (300); - a probe housing body (21) of oblong shape intended to be placed in the solution (300); the electronic device (10) being integrated into the probe housing head (20), and the probe housing body (21) being intended to accommodate at least a portion of a support for said sensor assembly (13), the probe housing head (20) and / or the probe housing body (21) being provided with at least one mechanical coupling element to allow a removable assembly of the housing head (20) (21) and the housing body (21) into a probe housing.

12. Probe (200) according to claim 11, wherein the probe body comprises a fixing structure (212), in particular a thread, to allow the assembly of the probe body (21) onto a receiving port, in particular provided with an additional thread.

13. Electrochemical measurement system (400) for measuring physicochemical quantities and concentrations of molecules in a solution (300), said system comprising: - an electro-chemical measurement probe (200) according to claim 12, - a bioreactor (500) equipped with a housing (50) intended to accommodate said solution and a receiving port (51) for a probe housing body (21) arranged between an external volume (53) and an internal volume (52) of the housing (50), in particular a port according to standard PG 13.5, the probe housing body (21) extending into said housing when the probe housing is assembled with the bioreactor (500).

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