KR20210064064A - Cortisol Sensor Manufacturing Method to measure concentration of Cortisol, and Smart Contact Lens Manufacturing Method having the Cortisol Sensor - Google Patents

Abstract

The present invention relates to a method for manufacturing a smart contact lens having a cortisol sensor, which comprises: a step B1 of manufacturing a film structure including a sacrificial layer, a passivation layer, an electrode, and a rigid island with a cortisol sensor on a substrate; a step B2 of peeling the film structure manufactured in the step B1 to be attached to a separate handling layer; a step B3 of forming a graphene channel in the film structure attached to the handling layer; a step B4 of having a wireless communication chip, a capacitor and a resistor provided as an integrated body electrically connected to each other at one side of the film structure; a step B5 of protecting the rigid island having the cortisol sensor by a stamp; a step B6 of manufacturing a lens by inputting a structure after the step B5 and a lens material into a lens mold; and a step B7 of removing the stamp, coupling a cortisol monoclonal antibody (C-Mab) to the graphene surface, and activating the cortisol sensor. Accordingly, the cortisol concentration can be measured.

Description

A method for manufacturing a cortisol sensor capable of measuring cortisol concentration, a method for manufacturing a cortisol sensor manufactured by the method, a smart contact lens equipped with a cortisol sensor, and a smart contact lens manufactured by the method {Cortisol Sensor Manufacturing Method to measure concentration of Cortisol, and Smart Contact Lens Manufacturing Method having the Cortisol Sensor}

The present invention relates to a field of a cortisol immune sensor integrated into a soft smart contact lens capable of on-site detection of stress hormones and a manufacturing method thereof. Specifically, the present invention relates to a method for manufacturing a cortisol sensor capable of measuring cortisol concentration, a method for manufacturing a cortisol sensor manufactured by the method, and a smart contact lens equipped with a cortisol sensor, and a smart contact lens manufactured by the method.

Cortisol, which is known to occur when physically and mentally stressed, is a steroid hormone secreted by the adrenal glands to maintain homeostasis. Cortisol is secreted when the adrenal glands are stimulated by adrenocorticotropic hormone (ACTH), and ACTH is secreted when the pituitary gland is stimulated by corticotropin-releasing hormone (CRH) secreted by the hypothalamus.

This series of cortisol secretion system is called the HPA axis, and chronic stress affects this cortisol secretion system, causing abnormal cortisol secretion. When cortisol levels build up unnecessarily, they increase fat and amino acid levels, leading to serious diseases (eg, Cushing's disease, autoimmune diseases, cardiovascular complications, type 2 diabetes) as well as neurological diseases such as depression and anxiety disorders. can cause

Cortisol levels can cause Addison's disease with symptoms of hyperpigmentation, weight loss and chronic fatigue, not only when it accumulates unnecessarily, but also when it is abnormally low.

This cortisol concentration changes continuously, and since there is a difference in the degree of secretion for each individual, the change in cortisol concentration should be monitored in real time and continuously. In order to measure the concentration of cortisol, a sensor to which an immunoassay method using an antigen-antibody binding reaction is applied is being studied.

If the immunoassay method is applied to the sensor, a cortisol antibody is attached to the electrode to produce an immunosensor of the electrical measurement method, which is a relatively simple measurement method. It was mainly applied in trials.

Among many body fluids, tears contain many biomarkers and also cortisol. Among wearable electronics, contact lenses are the only platform that can measure biomarkers from the eye because they are in constant contact with tears. The use of a contact lens-type sensor enables non-invasive real-time monitoring of cortisol in tears.

In addition, the amount of tears pooled in the eyes is 6.5±0.3 μL, and since it is almost constant, it is possible to measure a certain amount of tears without a separate extraction process.

Most of the cortisol sensors available to date have measured the concentration of cortisol contained in blood, saliva, sweat, hair, urine, or interstitial fluid, but biologically active cortisol in body fluids is unstable at room temperature. There are limitations in that it is difficult to extract, store, and transport body fluids, and it is difficult to accurately measure the concentration of cortisol.

In addition, the existing method for measuring cortisol requires extraction or pre-treatment, and analysis using a bulky measuring device is required. Therefore, an on-site detection sensor is needed for disease diagnosis and prevention by measuring an accurate cortisol concentration in real time.

A cortisol monoclonal antibody (C-Mab) is attached to a gold (Au) electrode, and the cortisol concentration in saliva and interstitial fluid is measured by electrochemical impedance spectroscopy (EIS) technology. A cortisol immunosensor has been developed, but an impedance analyzer, which is a device for analyzing a nyquist plot, is required.

In addition, a cortisol sensor based on cyclic voltammetry (CV) measurement has been developed.

However, this measurement method also requires a separate electrochemical instrument to interpret the CV curve.

Recently, in an attempt to develop a cortisol sensor on a wearable platform that can detect on-site detection, a metal thin-film electrode-based sweat cortisol sensor has been developed, but this device also requires a separate measuring device for electrochemical analysis. Do.

A cortisol sensor with a field effect transistor (FET) structure has been developed, which suggests the potential to be used as a field-detectable sensor, but it is not a wearable form, and the cortisol concentration can be measured using a measuring device. measured.

(Document 1) Korean Patent Publication No. 10-2042628 (2019.11.04)

A cortisol sensor capable of measuring cortisol concentration according to the present invention and a method for manufacturing a smart contact lens having the same have the following problems.

First, it is intended to manufacture a cortisol sensor capable of quantitatively measuring cortisol concentration.

Second, we intend to non-invasively monitor the concentration of cortisol that can be obtained from the eye and tears through a smart contact lens device that can quantitatively measure the concentration of cortisol.

Third, it is intended to diagnose diseases by wirelessly transmitting the collected data to the user.

Fourth, monitoring is possible without user inconvenience.

The problems to be solved of the present invention are not limited to those mentioned above, and other problems not mentioned will be clearly understood by those skilled in the art from the following description.

The present invention provides a step A1 of moving the synthesized graphene layer to a substrate and forming a graphene channel through patterning; A2 step of exposing the graphene channel to ultraviolet ozone (UVO) to activate the graphene surface; and A3 step of binding a cortisol monoclonal antibody (C-Mab) to the surface of the activated graphene.

The synthesis of the graphene layer in step A1 according to the present invention may be performed by chemical vapor deposition (CVD).

The patterning in step A1 according to the present invention may be performed by a photolithography method.

In step A2 according to the present invention, the graphene surface exposed to ultraviolet ozone may be activated with carboxylate groups.

In step A2 according to the present invention, the hydrophobicity of graphene may be reduced by activation of the graphene surface.

In step A3 according to the present invention, a cortisol monoclonal antibody (C-Mab) may be immobilized on the graphene surface through a two-step coupling reaction between EDC and NHS.

The present invention is a cortisol sensor capable of measuring cortisol concentration, and is manufactured by the manufacturing method according to the present invention.

The present invention provides a method for manufacturing a smart contact lens equipped with a cortisol sensor, comprising: a step B1 in which a film structure including a rigid island provided with a sacrificial layer, a passivation layer, an electrode and a cortisol sensor on a substrate is manufactured; Step B2 in which the film structure prepared in step B1 is peeled off and attached to a separate handling layer; Step B3 in which a graphene channel is formed in the film structure attached to the handling layer; A step B4 in which a chip for wireless communication, a capacitor and a resistor are provided as an integrated body electrically interconnected from one side of the film structure; Step B5 in which the rigid island provided with the cortisol sensor is protected with a stamp; B6 step of manufacturing a lens by putting the structure and lens material after step B5 into a lens mold; and a B7 step of removing the stamp, binding a cortisol monoclonal antibody (C-Mab) to the graphene surface, and activating the cortisol sensor.

Step B1 according to the present invention includes a step B11 in which a sacrificial layer is provided on a substrate and a passivation layer is deposited on the sacrificial layer; B12 step of depositing and patterning an electrode material to form a source electrode connected to a cortisol sensor, a drain electrode, and an antenna pad connected to an antenna coil; Step B13 in which a hybrid antenna coil patterned in a spiral shape is formed by depositing a metal nanomaterial layer; Step B14 in which the rigid island region on which the cortisol sensor is seated is patterned with a rigid material; and a step B15 in which the lens material, which is a soft material, is coated over the entire area.

In step B11 according to the present invention, the sacrificial layer may be made of Ni/Cu, and the passivation layer may be made of parylene.

In step B11 according to the present invention, the Ni/Cu sacrificial layer may be deposited on the substrate by electron beam evaporation.

In step B12 according to the present invention, the electrode material is provided with Cr/Au, and the Cr/Au electrode is thermally evaporated and patterned to form a source electrode and a drain electrode by a photolithography method.

In step B13 according to the present invention, the hybrid antenna coil may be a flexible transparent antenna.

In step B13 according to the present invention, the hybrid antenna coil may be patterned into an AgNF-AgNF hybrid network coated with finer AgNW than AgNF on the AgNF network.

Step B2 according to the present invention includes step B21 in which the sacrificial layer is removed; and a step B22 in which the film structure formed on the substrate is peeled off from the substrate and then attached to a preset handling layer in an inverted state.

Step B3 according to the present invention may include a step B31 in which the antenna pad and a portion in which a graphene channel is to be formed between the source electrode and the drain electrode is opened while the passivation layer is patterned through etching; B32 step in which graphene is provided in the open portion; and a step B33 in which graphene channels are formed while the graphene is patterned through etching.

The step B4 according to the present invention comprises a step B41 in which a chip for wireless communication, a capacitor and a resistor are disposed and adhered to a preset position of the structure; and a step B42 in which the chip, capacitor and resistor for wireless communication are integrated by 3D printing.

In step B42 according to the present invention, the 3D printing can be printed so that the chip for wireless communication, the capacitor and the resistor are electrically connected in a stretchable manner by spraying liquid metal.

In step B5 according to the present invention, it is possible that the stamp is a PDMS stamp.

According to the present invention, in step B7, after the stamp is removed, the step B71 of exposing the graphene channel to ultraviolet ozone (UVO) to activate the graphene surface is further included.

In step B71 according to the present invention, the graphene surface exposed to ultraviolet ozone may be activated with carboxylate groups.

In step B71 according to the present invention, the hydrophobicity of graphene may be reduced by activation of the graphene surface.

In step B7 according to the present invention, the step B72 further comprises binding a cortisol monoclonal antibody (C-Mab) to the activated graphene surface.

In step B72 according to the present invention, a cortisol monoclonal antibody (C-Mab) can be immobilized on the graphene surface through a two-step coupling reaction between EDC and NHS.

The present invention is a smart contact lens equipped with a cortisol sensor, which is manufactured by the manufacturing method according to the present invention.

The cortisol sensor capable of measuring cortisol concentration according to the present invention and a method for manufacturing a smart contact lens having the same have the following effects.

First, a cortisol sensor capable of quantitatively measuring cortisol concentration can be manufactured.

Second, there is an effect of non-invasively monitoring the concentration of cortisol that can be obtained from the eye and tears through a smart contact lens device that can quantitatively measure the cortisol concentration.

Third, there is an effect of diagnosing diseases by wirelessly transmitting the collected data to the user.

Fourth, there is an effect that monitoring is possible without user inconvenience.

The effects of the present invention are not limited to those mentioned above, and other effects not mentioned will be clearly understood by those skilled in the art from the following description.

Fig. 1A shows the C-Mab immobilization process, Fig. 1B shows a schematic image of the graphene FET structure, Fig. 1C shows an optical microscope image of the fabricated graphene FET device (scale bar, 200 μm), and Fig. 1D shows The transfer curve of the drain current according to the gate voltage is shown.
Figure 2 relates to the in vitro test, Figure 2A shows the real-time relative current change at the gate voltage VG = 0 V and the drain voltage VD = 0.1 V, Figure 2B shows the correction curve of the relative current change according to the cortisol concentration, Figure 2C shows the relative change in resistance with cortisol concentration of buffer and artificial tear solvent, and Figure 2D shows the relative change of sensor resistance with cortisol concentration at 22 °C and 36.5 °C, each data point representing the mean of 10 samples Error bars represent SD.
Fig. 3 relates to NFC and a stretchable and transparent AgNF-AgNW hybrid antenna. Fig. 3A is a circuit diagram connected with an NFC chip, a cortisol sensor and components including an antenna and a resistor, and Fig. 3B is a schematic diagram of a stretchable and transparent antenna for wireless communication. , Fig. 3C is a scanning electron microscope image (scale bar, 20 μm) of a random network of AgNF-AgNW hybrid structures, Fig. 3D is the transparency of AgNF-AgNW hybrid patterned antenna coil, and Fig. 3E is a negligible strain of up to 30%. shows the stretching-relaxation cycle test (inset: AgNF-AgNW hybrid electrode strain test) performed up to 300 times that of AgNF-AgNW hybrids, and Fig. 3F shows the resonant frequency of the patterned antenna resulting in 13.56 MHz.
Figure 4 relates to smart contact lens packaging, Figure 4A is a schematic diagram of a packaged smart contact lens integrated with a three-dimensional (3D) printed stretchable interconnect and cortisol sensor located on a rigid island, with capacitors for resonant frequency and reference resistance. Resistors are respectively integrated, Figure 4B shows a photograph of the fabricated smart contact lens (inset: close-up external image of the smart contact lens) (scale bar, 1 cm), and Figure 4C shows the optical transmittance and Haze is shown, and Fig. 4D shows the radiation characteristics before and after the return of the stretchable antenna, and Fig. 4E shows the relative resonance frequency change immersed in PBS and artificial tears up to 192 h (insertion: 12 h and 192 h in artificial tears, respectively) The radiation characteristics of the antenna after immersion test during and after).
Fig. 5 relates to an in vivo test, Fig. 5A is a photograph of an adult woman wearing a smart contact lens in the left eye (inset: a close-up image of a smart contact lens in the eye), and Fig. 5B is a cortisol level using the smart contact lens Schematic images of the measurements, Fig. 5C and Fig. 5D show the simulation result SAR, Fig. 5E shows the slit lamp images (scale bar, 1 cm) of a human eye with fluorescein staining before and after smart contact lens wear, and Fig. 5F is the cortisol concentration measured as a function of cortisol solution concentration dropped into the eye using a contact lens sensor, where each data point represents the mean of 10 samples and the error bars represent the SD, and Figure 5G shows the cytotoxicity test of a smart contact lens. indicates
6 is a characteristic change of the graphene surface, showing the contact angle between a 1 μl drop of deionized water and clean UVO-exposed graphene, which was decreased by increasing the exposure time of UVO.
Figure 7 relates to C-Mab immobilization, Figure 7A shows the process of C-Mab immobilization through EDC/NHS coupling reaction, pure graphene is carboxylated by UVO exposure, and Figure 7B is a cortisol sensor (red), clean deionized water without immersion (black), and FT-IR spectrum of deionized water after immersion in 1 μg/ml cortisol monoclonal antibody (blue) solution.
Figure 8 relates to a reproducibility test, showing the ID-VG (transfer) curves of all graphene transistor samples, Figure 8A is a cortisol concentration of 1 ng / ml, Figure 8B is a cortisol concentration of 10 ng / ml, Figure 8C is 20 ng / cortisol concentration in ml, FIG. 8D is a cortisol concentration of 30 ng/ml, FIG. 8E is a cortisol concentration of 40 ng/ml, and FIG. 8F shows the absolute drain current values as a function of cortisol concentration, each data point is the mean of 10 samples. and error bars represent standard deviations.
Fig. 9 relates to a cortisol sensor characteristic test, where Fig. 9A is a schematic diagram of real-time cortisol sensing using a microfluidic channel, Fig. 9B is a calibration curve of the cortisol sensor characteristic calculated as a change in resistance, and Fig. 9C is a buffer solution (black color). ) and ALU solution (A: 50 μM ascorbic acid, L: 10 mM lactic acid, U: 10 mM urea) (red) relative change in resistance of the cortisol sensor with cortisol concentration, each data point representing the mean of 10 samples and error bars represents the standard deviation.
FIG. 10 relates to an antenna characteristic test, FIG. 10A is a photograph (scale bar, 1 cm) of an AgNF-AgNW antenna with 71% transparency, and FIG. 10B shows the resonance characteristics of the antenna for 16 days at 70°C air condition, FIG. 10C shows the relative change in the resonant frequency (blue) of the antenna and the relative change in the antenna reflection coefficient (red) in FIG. 10B.
11 shows the manufacturing process and materials of a smart contact lens.
Figure 12 is a rigid soft hybrid substrate property test, Figure 12A is a photograph of a cortisol sensor on a rigid island (scale bar is 200 μm), Figure 12B is a relative change in cortisol sensor resistance as a function of tensile strain, and Figure 12C is a photograph of a lens with a built-in rigid soft hybrid substrate, the optical transparency and haze were measured to be 93% and 1.2%, respectively (at 550 nm), the scale bar is 1 cm, and Fig. 12D is the optical characteristic of the integrated contact lens, the smart Transmittance (black) and haze (red) spectra of a contact lens are shown.
Figure 13 is an in vivo test on a human, Figure 13A shows the change in cortisol concentration measured in both eyes of a human subject using smart contact lenses, Figure 13B is a slit lamp test on a human eye, wherein the slit lamp image is a smart Photographs were taken before, during, and after wearing contact lenses (scale bar, 1 cm).
Fig. 14 is an in vivo test in rabbits, and Fig. 14A is a smart contact lens in the eyes of live rabbits. (Inset: Close-up image of rabbit eye.) Scale bar, 1 cm. Figures 14B and 14C show the thermal analysis of antenna radiation versus transmitted power (scale bar, 2 cm).
15 shows a method of manufacturing a cortisol sensor capable of measuring cortisol concentration according to the present invention.
16 shows a method of manufacturing a smart contact lens equipped with a cortisol sensor according to the present invention.

Hereinafter, with reference to the accompanying drawings, an embodiment of the present invention will be described so that those of ordinary skill in the art can easily carry out the present invention. As can be easily understood by those skilled in the art to which the present invention pertains, the embodiments described below may be modified in various forms without departing from the concept and scope of the present invention. As far as possible, the same or similar parts are indicated using the same reference numerals in the drawings.

The terminology used herein is for the purpose of referring to specific embodiments only, and is not intended to limit the invention. Singular forms as used herein also include plural forms unless the phrases clearly indicate the opposite.

The meaning of "comprising," as used herein, specifies a particular characteristic, region, integer, step, operation, element and/or component, and other specific characteristic, region, integer, step, operation, element, component and/or It does not exclude the presence or addition of groups.

All terms including technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the present invention belongs. Terms defined in the dictionary are further interpreted as having a meaning consistent with the related art literature and the presently disclosed content, and unless defined, are not interpreted in an ideal or very formal meaning.

The present invention relates to a field of a cortisol immune sensor integrated into a soft smart contact lens capable of on-site detection of stress hormones and a manufacturing method thereof.

The present invention implements a smart contact lens that measures cortisol contained in tears by fusion of a cortisol measuring sensor, a stretchable transparent element for wireless communication, and mobile interlocking communication technology. It can be used as a device.

In addition, the present invention provides a flexible transparent wireless device manufacturing technology for manufacturing a smart contact lens for diagnosing mobile-linked diseases as an element technology for manufacturing a wearable electronic device that can be attached to human skin, objects and plants. It can be used as a source technology that can be applied to fields such as , electronic skin, electronic skin, and healthcare.

The present invention can be implemented with a method for manufacturing a cortisol sensor and a cortisol sensor manufactured by the method for manufacturing the same. This is an invention about the cortisol sensor itself.

In addition, the present invention may be implemented as a method for manufacturing a smart contact lens equipped with such a cortisol sensor and a smart contact lens manufactured by the manufacturing method. A contact lens could be used for the real user's eye.

Hereinafter, the present invention will be described with reference to the drawings. For reference, the drawings may be partially exaggerated in order to explain the features of the present invention. In this case, it is preferable to be interpreted in light of the whole meaning of this specification.

First, a method of manufacturing a cortisol sensor will be described, and then a method of manufacturing a smart contact lens will be described.

Hereinafter, a method for manufacturing a cortisol sensor capable of measuring cortisol concentration according to the present invention will be described. 15 shows a method of manufacturing a cortisol sensor capable of measuring cortisol concentration according to the present invention.

The cortisol sensor according to the present invention is manufactured by binding a cortisol monoclonal antibody (C-Mab) to the graphene channel surface of a graphene FET for immune sensing of cortisol. Here, graphene acts as a transducer that converts the interaction between cortisol and C-Mab into an electrical signal.

The method for manufacturing a cortisol sensor capable of measuring cortisol concentration according to the present invention comprises: A1 step of moving the synthesized graphene layer to a substrate, and forming a graphene channel through patterning; A2 step of exposing the graphene channel to ultraviolet ozone (UVO) to activate the graphene surface; and A3 step of binding a cortisol monoclonal antibody (C-Mab) to the activated graphene surface.

Fig. 1A shows the C-Mab immobilization process, Fig. 1B shows a schematic image of the graphene FET structure, Fig. 1C shows an optical microscope image of the fabricated graphene FET device (scale bar, 200 μm), and Fig. 1D shows The transfer curve of the drain current according to the gate voltage is shown.

The synthesis of the graphene layer in step A1 according to the present invention may be performed by chemical vapor deposition (CVD). The patterning of step A1 may be performed by a photolithography method.

1A-1, the graphene layer synthesized by CVD can be transferred to a desired substrate and exposed to ultraviolet ozone (UVO) to activate the graphene surface with carboxylate groups.

1A-2, the graphene surface can be activated with carboxylate groups by exposing the patterned CVD graphene channels to ultraviolet ozone (UVO).

Figure 6 shows the contact angle between the graphene surface and the deionized (DI) water droplet. A longer exposure time to UVO can reduce the hydrophobicity of graphene while reducing the contact angle. That is, the hydrophobicity of graphene may be reduced by the activation of the graphene surface in step A2 according to the present invention.

Table 1 below is a table showing the contact angle and resistance according to UVO exposure time.

Table 1 shows the increase in the electrical resistance of graphene due to this UVO treatment. In the experiment, exposure time of 2 min to UVO decreased the contact angle from 70° to 38° without significantly increasing the resistivity of graphene.

UVO exposure time longer than this critical time excessively lowered the resistivity of graphene, so it is possible that the time of exposing the sample to UVO is limited to 2 min.

In step A3 according to the present invention, a cortisol monoclonal antibody (C-Mab) may be immobilized on the graphene surface through a two-step coupling reaction between EDC and NHS.

1A-3, the immobilization was performed on the carboxyl group of the graphene surface through the EDC [1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride]/NHS (N-hydroxysulfosuccinimide) coupling reaction. It can proceed through the amide bond of

Figure 7A shows the process of immobilizing C-Mab via an EDC/NHS coupling reaction. This two-step coupling reaction of EDC and NHS can mediate the amide bond between the carboxylate group of graphene exposed to UVO and the amine group of the protein.

Here, EDC forms a reactive O- acylisourea ester to destabilize the surface (see Fig. 7A-2).

This O-acylisourea ester reacts with NHS to form an amine-reactive NHS ester and the surface remains semi-stable (see Figure 7A-3).

The amine group of the C-Mab then reacts with the amine-reactive NHS ester to form a stable amide bond that immobilizes the C-Mab on the graphene surface (see Fig. 7A-4).

7B shows the Fourier transform infrared (FTIR) spectroscopy spectrum of DI water after the cortisol sensor was immersed for 24 h.

The spectrum of DI water immersed in the cortisol sensor was not significantly different from that of pure DI water. However, the C-Mab solution with a concentration of 1 μg/ml ^{showed significant peak intensities in the range of 3000 to 2800 cm −1} , indicating NH bonding in the C-Mab.

These results indicate that C-Mab formed a stable bond to carboxylated graphene and dissociated to a negligible degree upon exposure to water.

Therefore, graphene FETs in which graphene channels are selectively functionalized with C-Mab can be used as cortisol sensors.

1B shows a schematic diagram of a cortisol sensor. The source/drain electrodes and interconnects were passivated with a 500 nm thick epoxy layer (SU-8, MicroChem Inc.) except for the rectangular area used to expose the graphene channels.

1C shows an optical micrograph of a cortisol sensor.

After preparing an artificial tear solution containing a specific concentration of cortisol, for the characterization of this cortisol sensor, the solution was dropped into a graphene channel to measure the drain current by applying a gate voltage using an Ag/AgCl probe.

Figure 1D shows the transfer curves of this graphene FET with bipolar properties for various concentrations of cortisol.

Cortisol has an isoelectric point from pH 5.2 to pH 5.4. Therefore, it has a negative charge under physiological conditions (pH 7.35 ~ pH 7.45). Accordingly, electrons can be injected into the graphene channel in the adsorbate. And since the graphene channel (p-type) reduces the main carrier concentration, the drain current is reduced at a high concentration of cortisol.

In addition, data obtained from 10 samples are provided in Fig. 8 for further information on the reproducibility of the sensor. As shown in FIG. 8 , the cortisol sensor according to the present invention exhibited reproducible characteristics in conductivity versus gate potential at various concentrations.

On the other hand, the present invention can be implemented as a cortisol sensor capable of measuring the cortisol concentration prepared by the above-described manufacturing method.

Next, a method for manufacturing a smart contact lens equipped with a cortisol sensor according to the present invention will be described. The manufacturing method of the above-described cortisol sensor can be applied to the manufacturing method of the present invention. 16 shows a method of manufacturing a smart contact lens equipped with a cortisol sensor according to the present invention.

The manufacturing method of a smart contact lens equipped with a cortisol sensor according to the present invention consists of steps B1 to B7.

Step B1 according to the present invention is a step in which a film structure including a rigid island provided with a sacrificial layer, a passivation layer, an electrode, and a cortisol sensor on a substrate is manufactured.

Step B1 according to the present invention includes a step B11 in which a sacrificial layer is provided on a substrate and a passivation layer is deposited on the sacrificial layer; B12 step of depositing and patterning an electrode material to form a source electrode connected to a cortisol sensor, a drain electrode, and an antenna pad connected to an antenna coil; Step B13 in which a hybrid antenna coil patterned in a spiral shape is formed by depositing a metal nanomaterial layer; Step B14 in which the rigid island region on which the cortisol sensor is seated is patterned with a rigid material; and a step B15 in which the lens material, which is a soft material, is coated over the entire area.

Hereinafter, step B1 according to the present invention will be described in more detail.

In step B11, the sacrificial layer may be made of Ni/Cu, and the passivation layer may be made of parylene.

In step B11, the Ni/Cu sacrificial layer may be deposited on the substrate by electron beam evaporation.

In step B12, the electrode material is provided with Cr/Au, and the Cr/Au electrode is thermally evaporated and patterned to form a source electrode and a drain electrode by a photolithography method.

In step B13, the hybrid antenna coil may be a flexible transparent antenna.

In step B13, the hybrid antenna coil can be patterned with AgNF-AgNF hybrid network coated with finer AgNW than AgNF on AgNF network.

Step B2 according to the present invention includes step B21 in which the sacrificial layer is removed; and a step B22 in which the film structure formed on the substrate is peeled off from the substrate and then attached to a preset handling layer in an inverted state.

Step B3 according to the present invention is a step in which a graphene channel is formed in the film structure attached to the handling layer.

Step B3 according to the present invention may include a step B31 in which the antenna pad and a portion in which a graphene channel is to be formed between the source electrode and the drain electrode is opened while the passivation layer is patterned through etching; B32 step in which graphene is provided in the open portion; and a step B33 in which graphene channels are formed while the graphene is patterned through etching.

Step B4 according to the present invention is a step in which a chip for wireless communication, a capacitor and a resistor are provided as an integrated body that is electrically interconnected from one side of the film structure.

The step B4 according to the present invention comprises a step B41 in which a chip for wireless communication, a capacitor and a resistor are disposed and adhered to a preset position of the structure; and a step B42 in which the chip, capacitor and resistor for wireless communication are integrated by 3D printing.

In step B42, the 3D printing can be printed so that the chip for wireless communication, the capacitor and the resistor are electrically connected in a stretchable manner by spraying liquid metal.

Step B5 according to the present invention is a step in which the rigid island provided with the cortisol sensor is protected with a stamp. In step B5, it is possible that the stamp is a PDMS stamp.

Step B6 according to the present invention is a step of manufacturing a lens by putting the structure and lens material after step B5 into a lens mold.

Step B7 according to the present invention is a step of removing the stamp, binding a cortisol monoclonal antibody (C-Mab) to the graphene surface, and activating the cortisol sensor.

Step B7 may further include step B71 of exposing the graphene channel to ultraviolet ozone (UVO) to activate the graphene surface after the stamp is removed.

In step B71, the graphene surface exposed to ultraviolet ozone can be activated with carboxylate groups.

The hydrophobicity of graphene may be reduced by activation of the graphene surface in step B71.

Step B7 further includes a step B72 of binding a cortisol monoclonal antibody (C-Mab) to the activated graphene surface.

In step B72, a cortisol monoclonal antibody (C-Mab) can be immobilized on the graphene surface through a two-step coupling reaction between EDC and NHS.

On the other hand, the present invention can be implemented as a smart contact lens equipped with a cortisol sensor manufactured by the above-described manufacturing method.

Hereinafter, the present invention will be described more clearly while rearranging the main contents of the present invention once again based on the experiments actually performed.

Graphene FET structure sensor fabrication

Cr / Au electrodes were thermally evaporated to a thickness of 5/100 nm and patterned to form source and drain electrodes by photolithography.

The distance between them is the length of the channel, i.e. 560 μm. After the source and drain were formed, graphene was transferred between the source and the drain (water-transfer).

Chemical vapor deposition (CVD) graphene was purchased from Graphene Square.

The large-area graphene deposited on the Cu foil was cut to a desired size and spin-coated with poly (methyl methacrylate) (MicroChem Corp., 950 PMMA C2).

The PMMA unintentionally coated on the bottom of the graphene was rinsed with acetone.

Then, graphene was floated on Cu etchant (FeCl3 : HCl : H2 = 1 : 1 : 20 vol %) to etch the Cu foil on the bottom of the graphene.

When the etching was completed, the etching solution remained and the PMMA was removed by rinsing with DI water.

After rinsing, the graphene was moved to a desired position between the source and drain and patterned by photolithography and reactive ion etching (RIE) (50W, 40sccm, 120 sec).

The manufacturing process of smart contact lenses

Parylene was deposited as a passivation layer on a Ni/Cu (10/800 nm) sacrificial layer.

Cr/Au (5/100 nm) electrodes were deposited via e-beam evaporation and patterned with the source and drain of the FET type sensor and the antenna pad.

AgNF-AgNW hybrid electrodes were deposited by electrospinning and patterned on the antenna coil.

A rigid material (photopolymer) was patterned on the rigid region of the sensor.

A soft material (Elastofilcone A) was coated over the entire sample area.

The sacrificial layer was removed via wet etching. The sample was lifted from the substrate and adhered to the handling layer in the opposite position.

Parylene was patterned via RIE etching (100 W, 180 s) to drill holes in the pads of the antenna and sensor.

The graphene was transferred to the sensor to create graphene channels.

The graphene was patterned to make the channel of the sensor via RIE etching (50 W, 30 s).

An NFC chip, capacitor and resistor were coupled to the sample with lens material.

All components were integrated via 3D printing.

The sensor was protected with a PDMS stamp before molding the lens.

The lens material and the fabricated device were cast together in a lens mold at a pressure of 313 kPa and cured at 70 °C for 4 hours.

The smart contact lens was completed after removing the PDMS stamp and functionalizing the cortisol sensor by dripping EDC, NHS and C-Mab solutions.

Hard and soft hybrid film

A Ni/Cu (10/1 μm) sacrificial layer (300 nm thick, MicroChem Corporation) was deposited on a SiO2 wafer via electron beam evaporation.

Then, parylene (1 μm) was deposited on the sacrificial layer and Au source/drain portions of the sensor were deposited and patterned.

After that, an optical polymer (SPC-414, EFiRON) was rotated to a thickness of 50 μm (1500 rpm for 30 sec) and a photolithography pattern was applied as a rigid island structure.

Then, an elastomer (Elastofilcon A, CooperVision) mixed with a base and a curing agent in a weight ratio of 10:1 is rotated (1000 rpm 30 s, thickness 100 μm) and heat-cured at 100 °C.

1 h (film thickness, 5 μm). As a final step, the sacrificial layer was removed by wet etching using an etchant of FeCl3 / HCl / H2O [1 : 1 : 20 (v / v)] to peel the film _{from the SiO 2 wafer.}

Elasticity integration of soft contact lenses

To form a soft, smart contact lens, all components of the cortisol sensor, antenna, NFC chip, capacitor and resistor must be electrically integrated with a stretchable interconnect.

The softness of contact lenses can cause fractures in the existing brittle and hard materials of integrated electronic systems, damaging the cornea or eyelids.

In the present invention, a soft contact lens with a highly transparent and stress-tunable hybrid shape composed of each component (eg, cortisol sensor, NFC chip, capacitor and resistor) and an elastic joint to confirm position was formed.

For stretchable and transparent antennas and interconnecting electrodes, in one embodiment, the hard part was patterned using a thin, photo-patternable optical polymer (SPC-414, EFiRON), and the elastic part was silicone, a conventional material for soft contact lenses. Elastomer (Elastofilcon A, CooperVision) was applied.

Using a locating device, after forming a planar film of hard-soft hybrid material, all components of the cortisol sensor device, i.e., NFC chip, capacitor, resistor and antenna, were placed in the eutectic liquid metal [75 weight % (wt %) Ga and 25 wt % In; Changsha Santech Materials Co., Ltd. Ltd] were printed with a narrow line width (<10 μm) and electrically connected.

A stretchable pattern of these interconnects was formed by direct high-resolution printing of liquid metal through a nozzle at ambient conditions.

After full integration, a silicone elastomer precursor was injected to mold a flat sample (including the device) generated into the curved shape of the lens, completing the fabrication of a soft and smart contact lens.

During this shaping step the graphene channels are opened and locally exposed by the lens material (silicon elastomer) so that the channels and tears can make physical contact.

In this way, any device can be embedded inside a soft contact lens, leaving this opening for the sensor.

4 relates to smart contact lens packaging.

Figure 4A is a schematic diagram of a packaged smart contact lens integrated with a three-dimensional (3D) printed stretchable interconnect and a cortisol sensor located on a rigid island, with capacitors and resistors integrated for resonant frequency and reference resistance, respectively; A photograph of the fabricated smart contact lens (inset: close-up external image of the smart contact lens) (scale bar, 1 cm) is shown, Figure 4C shows the optical transmittance and haze of the rigid-soft hybrid material, and Figure 4D shows the stretchable The radiation characteristics before and after the return of the antenna are shown, and FIG. 4E shows the relative resonance frequency change immersed in PBS and artificial tears up to 192 hours (insertion: radiation characteristics of the antenna after immersion tests in artificial tears for 12 hours and 192 hours, respectively) .

Hereinafter, FIG. 4 will be described in more detail.

Figure 4A shows a schematic diagram of a smart contact lens for monitoring tear cortisol.

4B shows a photograph of a real lens incorporating a cortisol radio measurement system.

A contact lens based on a rigid-soft hybrid structure in which a rigid polymer island (island) is embedded inside an elastic layer can effectively dissipate mechanical strain and protect electronic components from mechanical strain of the lens.

The difference in modulus of elasticity (E) between the hard and soft parts is large enough to concentrate the tensile stress on the soft part of the lens. For example, the E of the elastomer (Elastofilcon A) is ~0.09 MPa and the E of the optical polymer is ~360 Mpa.

The sensor was placed on a hard island embedded within the soft contact lens layer (see FIG. 12A ).

This layout allowed the sensor to be mechanically separated from the deformation of the soft contact lens, and the change in resistance was negligible (see Fig. 12B).

Referring to Fig. 4C, for the transparency of the lens, the optical refractive indices (n) of the two dissimilar materials are similar (n, 1.41 of Elastofilcon A; n, 1.407 of optical polymer).

Figure 4C shows the transparency and haze curves of a flat film of a rigid-soft hybrid (Elastofilcon A thickness, 100 μm, optical polymer thickness, 50 μm).

These hybrid substrates exhibited good transmittance (~93% at 550 nm) with low haze (~1.2% at 550 nm).

For example, FIG. 12C provides a photograph of this contact lens (without the components of the device included) where the boundary between these two materials (optical polymer and Elastofilcon A) is barely visible.

In addition, no gaps were created at the interface between these disparate regions even during the stretched state (tensile strain of 30%).

12D shows that the optical clarity and haze of the integrated contact lens were measured to be 67% and 10.0% (at 550 nm), respectively.

In addition, interference to the wearer's field of vision is minimized because all components of the device are positioned outside the wearer's pupil.

Thus, contact lenses can reduce the adverse effect on the wearer's vision.

Figure 4D shows that the resonance characteristics of the antenna (embedded inside this lens) remained stable before and after flipping the soft lens.

In addition, the AgNF-AgNW antenna of this contact lens was encapsulated with a parylene elastomer passivation layer (thickness, 1 μm) to prevent oxidation of silver.

In Fig. 4E, although this smart contact lens was immersed in PBS solution (pH 7.4) or artificial tear solution for 192 hours, the resonance characteristics of the antenna were not significantly deteriorated.

Fabrication of AgNF-AgNW Hybrid Antenna

AgNF-AgNW electrodes were formed by electrospinning and subsequent electrospray.

AgNFs were formed by electrospinning for 10 s under the following conditions using a suspension of Ag nanoparticles (NPK, Korea, average diameter: 40 ± 5 nm, solvent: ethylene glycol, concentration: 50 wt. %).

The inner and outer diameters of the nozzles were 0.34 mm and 0.64 mm, respectively.

The distance between the nozzle and the substrate and the DC bias were set to 21 cm and 8 kV, respectively.

The environmental conditions were a temperature of 14°C and a relative humidity of 3.6%. AgNF was formed by coalescing Ag nanoparticles by heating at 150 °C for 30 min.

After that, AgNWs (Flexio Co., Ltd.) had an average diameter of 30 ± 5 nm and an average length of 25 ± 5 μm, and electrosprayed AgNFs on the electrospun substrates for 1 min.

The distance between the nozzle and the substrate and the DC bias were set to 13 cm and 9.5 kV, respectively.

The diameter of the nozzle was 0.33 mm. AgNF-AgNW electrodes were photolithographically patterned through a wet etching process.

in vivo tests

Figure 5A shows a photograph of a 24-year-old female subject, each wearing a fully integrated smart contact lens capable of real-time wireless detection of cortisol using a mobile phone.

For reference, FIG. 13A shows the change in cortisol concentration as a function of time for a human subject.

Figure 5B shows a schematic diagram of such a lens by wirelessly transmitting data to a reader (eg, a smartphone) via a standard NFC interface.

A smartphone's wireless power supply can provide an AC bias that operates a logic chip to evaluate the sensor's response, which includes an ADC and all computing functions for high-speed data recording and real-time wireless transmission.

In this design of the contact lens, even when all components of the device are arranged, the wearer's pupil is not blocked, thereby minimizing interference with the wearer's field of view.

Figures 5C and 5D show the simulation results for the absorption rate (SAR) in humans, indicating that the maximum SAR value of this contact lens is only 0.102 W/kg.

This value was comparable to the electromagnetic wave level of a smartphone (0.14 to 0.33 W/kg), and was about 20 times lower than the prescribed value (2 W/kg).

In addition, after wearing this lens for 12 hours, a slit lamp to examine the patient's eyes was performed.

Figure 5E shows a slit lamp image to which fluorescein staining has been applied, indicating that there is no apparent response of the human cornea to this smart lens.

In addition, the conjunctiva of the eye was examined before and after wearing the lenses for 12 hours.

Of note, Figure 13B shows that no substantial conjunctival injection was observed during this time.

In another in vivo test, an artificial tear solution with a cortisol concentration of 20 ng/ml was placed in the eyes of live rabbits and then dropped into contact lenses.

The cortisol solution (C106; Sigma-Aldrich) prepared for this experiment and a commercially available artificial tear solution were mixed at the desired concentration for 1 min at room temperature prior to administration of this eye drop.

This is because cortisol concentrations in rabbit plasma are much lower than in humans, and the cortisol sensor is calibrated to measure cortisol concentrations in humans.

After wearing these contact lenses during the antigen-antibody reaction threshold period, the cortisol concentration was detected wirelessly through automatic calibration of the smartphone software and then quantitatively displayed on the mobile phone screen.

Figure 5F shows that the cortisol concentration measured using the contact lens sensor has a good correlation with the concentration of the prepared cortisol solution.

The rabbits showed no abnormal behavior while wearing the lenses, and these contact lenses were stable even with repeated blinks.

In addition, an IR camera was used to monitor the heat generated while the rabbit wore the lenses ( FIG. 14A ).

In this case, an external metal coil was used instead of a cell phone to wirelessly transmit power to the antenna of this contact lens through an air gap of ~5 mm, as shown in Figs. 14B and 14C.

While the power was delivered for the operation of this lens circuit, the thermal fluctuations due to magnetic coupling in the low frequency band (13.56 MHz) were negligible (ΔT ~ 0.5 °C).

Although the temperature of the transmitting coil rose to ~0.5 °C to deliver power to the lens, thanks to the radio, this coil did not touch the rabbit's eyes or eyelids with an air gap of ~5 mm.

The cytotoxicity of these contact lenses was tested by measuring the viability of human dermal fibroblasts.

The cytotoxicity test used the WST-8 assay to quantify viable cells by culturing cells in extracts of test samples and controls and then detecting biological responses.

Extracts were prepared by incubating samples in minimal essential medium (1x MEM; Gibco) with 10% serum (equine serum; Gibco) at 37 ± 1 °C and 5 ± 1% CO2 for 48 h.

The extraction rate was set at 0.2 g/ml, which corresponds to an irregular model of solid medical devices. Experimental methods are described in detail in Materials and Methods.

Figure 5G shows cell viability of test samples, blanks, negative and positive controls.

The cell viability of the test sample (including contact lenses) was 83.3%, comparable to that of commercial soft contact lenses without electronics (cytotoxicity standardized according to ISO 109943-5:2009).

Therefore, it was concluded that smart contact lenses used to monitor cortisol levels are practically non-cytotoxic to people.

Calculation of cortisol concentration

The concentration of cortisol was calculated from the change in the resistance of the sensor. Therefore, the measurement procedure is to measure the resistance after adjustment, i.e. at zero concentration. The change in resistance was calculated using the following formula.

[Equation 1]

where V _ADC and V _{DAC are the} measured voltage and applied voltage, and R _Ref is the resistance of the resistor integrated with the NFC chip. Cortisol concentration can be calculated using the following equation: Here, the unit of concentration is ng/ml.

[Equation 2]

Cell culture and cytotoxicity assays

Normal human dermal fibroblasts (PromoCell) were maintained in Cascade Biologics Medium 106 (Gibco), with medium change every 3 days in a humidified atmosphere at 37 °C, 5% CO2 supplemented with low serum growth supplement (Gibco) (complete medium). do. .

When confluence was closed at 80%, cells were passaged with 0.025% trypsin/EDTA (Gibco) and cultured in complete medium conditions.

At the fourth passage, these cells were harvested and plated at a density of 5000 cells per well in each well of a 96-well plate.

Cells were cultured in complete medium for 24 hours.

The process was performed according to ISO 10993-5 for cytotoxicity test.

Human dermal fibroblasts were selected because of their high sensitivity to chemicals in cytotoxicity tests and the abundance of data related to cytotoxicity tests.

In this experiment, our smart contact lenses were compared on polyurethane films containing 0.1% zinc diethyl dithiocarbamate (Hatano Laboratories) and high-density polyurethane films (Hatano Laboratories) as positive and negative controls.

Before the experiment, all contact lenses were sterilized in 70% ethanol for 30 min, and then the samples were dried in a hood.

Each sample (n = 4) was immersed and incubated in complete medium at 37 °C for 48 h. The conditions of the extract were prepared with 0.2 g of contact lenses in 1 ml of complete medium.

The pretreated medium of the cultured cells was changed to the extraction medium immersed in contact lenses.

Cells treated with the extracted medium were incubated for 24 h. Cytotoxicity was assessed by Cell Counting Kit-8 assay (Dojindo). The procedures in the instructions were followed.

Absorbance was read at 450 nm using a multi-mode plate reader (PerkinElmer).

Absorbance values were converted to percentage values for blanks obtained only in cell growth medium.

rabbit experiment

In accordance with the guidelines of the National Institutes of Health for the Care and Use of Laboratory Animals, in vivo testing was performed with the approval of the Ulsan Institute of Science and Technology (UNIST) Animal Care and Use Committee.

Institute of Animal Care, Yonsei University (UNISTIACUC-18-02 and IACUC-A-201910-963-01). The ethics review committee consisted of the UNIST Animal Care Center and the Yonsei University Animal Care Center Use Committee.

Fourier Transform Infrared Spectroscopy (FT-IR)

FT-IR spectra of the samples were obtained using an FT-IR spectrometer (Vertex 70, Bruker) at 25 °C with an attenuated total reflectance (ATR) step.

All spectra from 256 scans were recorded between 4000 and 1000 cm-1 with a resolution of 4 cm-1.

Microfluidic Channels for Real-Time Cortisol Detection

Negative photoresist (PR) was patterned into the structure inside the microfluidic channel.

PDMS (Sylgard 184 silicone elastomer:curing agent = 10:1 wt%) was then cured on the patterned negative PR.

The PDMS was separated from the negative PR and a hole was drilled for solution injection to complete the microfluidic channel.

A PDMS microfluidic channel was placed on the sensor and diluted cortisol solution (Sigma-Aldrich., #C-106, varying concentrations) and buffer (Samcheon Pure Pharma, pH7.00 ± 0.02) were injected.

Microfluidic channel with syringe pump (New Era Pump Systems, Inc., NE-300). The amount of biofluid can be calculated using the flow rate (1 ml/h) and the response time of the sensor (3 sec).

The response time, i.e., the time required for the current level to reach 90% of the saturation level at a cortisol concentration of 1 ng/ml, was calculated in Fig. 2A.

Therefore, the minimum biofluid volume required for cortisol detection was 0.83 μl, i.e. 1 ml/hour × 3 s × 1 hour/3600 s × 1000 μl/ml, which was less than the amount of tears.

selectivity test

For the selectivity test, the sensor is tested with 1 - 40 ng/ml cortisol solution containing 50 μM ascorbic acid (product number PHR1008, Sigma-Aldrich), 10 mM lactic acid (product number PHR1113, Sigma-Aldrich) and 10 mM lactic acid.

mM urea (cat number PHR1406, Sigma-Aldrich).

Accelerated aging test for antenna

In the accelerated aging test, the accelerated aging time may be determined by the following formula.

[Equation 3]

Here, Q10, TAA and TAMB are aging factors (~2), accelerated aging temperature (70 °C) and ambient temperature (2), respectively.

Accelerated aging time was calculated to be 16 days because TAMB was assumed to be room temperature (25 °C) for a storage period of 1 year.

According to this calculation, our contact lens device was stored in air for up to 16 days.

Electromagnetic Absorption Rate (SAR) Simulation

Finite element analysis was performed to simulate a person's maximum SAR.

Simulations were performed using commercial software, namely Ansoft HFSS.

In the simulation, the antenna was placed under the human head model, and the transmit power of the transmitter or smart phone was set to 10W for extreme environment testing.

Integrated wireless circuitry built into soft contact lenses

A fully integrated device including a cortisol sensor, antenna coil, NFC chip, reference resistor and capacitor is embedded in the lens material.

The fabricated device and lens material were put together in a lens mold and cured at 100 °C for 1 hour at a pressure of 313 kPa.

After that, the soft lens with the built-in device was removed from the mold.

Slit lamp examination in human studies

The protocol of this study was approved by the Institutional Review Board of UNIST (UNIISTIRB-18-17-A) and the participants gave informed consent.

Smart contact lenses were rinsed using a commercially available contact lens cleaning solution (Frenz-pro B5 solution, JK Pharmaceutical Inc., Korea), and rinsed with PBS solution for 1 minute before wearing.

This test was performed after wearing the lenses for 12 hours.

After the human experiment, the ocular surface of the volunteers was evaluated using the slit lamp test (SL-15, Kowa Optimed Inc., Japan).

PDMS stamp for sensor protection

Negative photoresist (PR) was patterned into the internal structure of the stamp.

Then, PDMS (Sylgard 184 silicone elastomer:curing agent = 10:1 wt%) was cured on the patterned negative PR.

A PDMS stamp was formed by separating the PDMS from the negative PR.

The embodiments described in the present specification and the accompanying drawings are merely illustrative of some of the technical ideas included in the present invention. Accordingly, since the embodiments disclosed in the present specification are for explanation rather than limitation of the technical spirit of the present invention, it is obvious that the scope of the technical spirit of the present invention is not limited by these embodiments. Modifications and specific embodiments that can be easily inferred by those skilled in the art within the scope of the technical idea included in the specification and drawings of the present invention should be interpreted as being included in the scope of the present invention.

Claims (25)

A1 step of moving the synthesized graphene layer to a substrate and forming a graphene channel through patterning;
A2 step of exposing the graphene channel to ultraviolet ozone (UVO) to activate the graphene surface; and
A method of manufacturing a cortisol sensor capable of measuring cortisol concentration, comprising: a step A3 of binding a cortisol monoclonal antibody (C-Mab) to the activated graphene surface. The method according to claim 1,
A method of manufacturing a cortisol sensor capable of measuring cortisol concentration, characterized in that the synthesis of the graphene layer in step A1 is performed by chemical vapor deposition (CVD). The method according to claim 1,
A method of manufacturing a cortisol sensor capable of measuring cortisol concentration, characterized in that the patterning of step A1 is performed by a photolithography method. The method according to claim 1,
In step A2, the graphene surface exposed to ultraviolet ozone is a method of manufacturing a cortisol sensor capable of measuring cortisol concentration, characterized in that it is activated with a carboxylate group. The method of claim 4,
A method of manufacturing a cortisol sensor capable of measuring cortisol concentration, characterized in that the hydrophobicity of graphene is reduced by activation of the graphene surface in step A2. The method of claim 4,
In step A3, a method of manufacturing a cortisol sensor capable of measuring cortisol concentration, characterized in that the cortisol monoclonal antibody (C-Mab) is fixed to the graphene surface through a two-step coupling reaction between EDC and NHS. A cortisol sensor capable of measuring a cortisol concentration manufactured by the manufacturing method according to any one of claims 1 to 6. Step B1 in which a film structure including a rigid island provided with a sacrificial layer, a passivation layer, an electrode, and a cortisol sensor is prepared on a substrate;
Step B2 in which the film structure prepared in step B1 is peeled off and attached to a separate handling layer;
Step B3 in which a graphene channel is formed in the film structure attached to the handling layer;
A step B4 in which a chip for wireless communication, a capacitor and a resistor are provided as an integrated body electrically interconnected from one side of the film structure;
Step B5 in which the rigid island provided with the cortisol sensor is protected with a stamp;
B6 step of manufacturing a lens by putting the structure and lens material after step B5 into a lens mold; and
B7 step of removing the stamp, binding cortisol monoclonal antibody (C-Mab) to the graphene surface, and activating the cortisol sensor
A method of manufacturing a smart contact lens equipped with a cortisol sensor. The method according to claim 8, B1 step
a step B11 in which a sacrificial layer is provided on a substrate and a passivation layer is deposited on the sacrificial layer;
B12 step of depositing and patterning an electrode material to form a source electrode connected to a cortisol sensor, a drain electrode, and an antenna pad connected to an antenna coil;
Step B13 in which a hybrid antenna coil patterned in a spiral shape is formed by depositing a metal nanomaterial layer;
Step B14 in which the rigid island region on which the cortisol sensor is seated is patterned with a rigid material; and
A method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that it comprises a step B15 in which the entire area is coated with a lens material, which is a soft material. The method according to claim 9, in step B11
The sacrificial layer is made of Ni/Cu, and the passivation layer is made of Parylene. The method according to claim 10, in step B11
The Ni/Cu sacrificial layer is a method of manufacturing a smart contact lens provided with a cortisol sensor, characterized in that deposited on the substrate by electron beam evaporation. The method according to claim 9, in step B12
The electrode material is provided with Cr/Au, and the Cr/Au electrode is thermally evaporated and patterned, and a source electrode and a drain electrode are formed by a photolithography method. Method of manufacturing a smart contact lens equipped with a cortisol sensor . The method according to claim 9, in step B13
The hybrid antenna coil is a method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that the elastic transparent antenna. The method according to claim 9, in step B13
A method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that the hybrid antenna coil is patterned with an AgNF-AgNF hybrid network coated with AgNW finer than AgNF on the AgNF network. The method according to claim 8, B2 step
step B21 in which the sacrificial layer is removed; and
After the film structure formed on the substrate is peeled off from the substrate, the method for manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that it comprises a step B22 that is attached to a preset handling layer in an inverted state. The method according to claim 8, B3 step
a step B31 of opening the antenna pad and a portion in which a graphene channel is to be formed between the source electrode and the drain electrode while the passivation layer is patterned through etching;
B32 step in which graphene is provided in the open portion; and
Method of manufacturing a smart contact lens provided with a cortisol sensor, characterized in that it comprises a step B33 in which graphene channels are formed while the graphene is patterned through etching. The method according to claim 8, B4 step
A step B41 in which a chip for wireless communication, a capacitor and a resistor are disposed and adhered to a preset position of the structure; and
The method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that the chip, capacitor and resistor for wireless communication include a step B42 that is integrated by 3D printing. The method according to claim 17, in step B42,
The 3D printing is a method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that by spraying liquid metal, the wireless communication chip, capacitor and resistor are printed so as to be stretchable and electrically connected. The method according to claim 8, in step B5,
The stamp is a method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that the PDMS stamp. The method according to claim 8, in step B7,
After the stamp is removed, the method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that it further comprises the step B71 of exposing the graphene channel to ultraviolet ozone (UVO) to activate the graphene surface. The method of claim 20, wherein in step B71,
A method for manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that the graphene surface exposed to ultraviolet ozone is activated with a carboxylate group. The method of claim 21,
Method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that the hydrophobicity of graphene is reduced by the activation of the graphene surface in step B71. The method according to claim 20, in step B7,
Method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that it further comprises the step B72 of binding a cortisol monoclonal antibody (C-Mab) to the activated graphene surface. The method of claim 23,
In step B72, a method of manufacturing a smart contact lens equipped with a cortisol sensor, characterized in that the cortisol monoclonal antibody (C-Mab) is fixed to the graphene surface through a two-step coupling reaction between EDC and NHS. A smart contact lens equipped with a cortisol sensor manufactured by the manufacturing method according to any one of claims 8 to 24.

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