WO2017022923A1 - Capteur de type papier pour diagnostic du cancer de la prostate, son procédé de fabrication, et procédé de diagnostic du cancer de la prostate l'utilisant - Google Patents

Capteur de type papier pour diagnostic du cancer de la prostate, son procédé de fabrication, et procédé de diagnostic du cancer de la prostate l'utilisant Download PDF

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WO2017022923A1
WO2017022923A1 PCT/KR2016/002808 KR2016002808W WO2017022923A1 WO 2017022923 A1 WO2017022923 A1 WO 2017022923A1 KR 2016002808 W KR2016002808 W KR 2016002808W WO 2017022923 A1 WO2017022923 A1 WO 2017022923A1
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paper
prostate cancer
antibody
carbon
psa
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Korean (ko)
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김돈
지성경
이명순
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부경대학교 산학협력단
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/28—Electrolytic cell components
    • G01N27/30—Electrodes, e.g. test electrodes; Half-cells
    • G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer

Definitions

  • the present invention relates to a prostate cancer diagnostic sensor comprising a paper-type sensing unit, a manufacturing method thereof, and a prostate cancer diagnostic method using the same.
  • Prostate cancer is a form of cancer that manifests in the gland within the prostate, the male reproductive organs.
  • the cancer cells can metastasize from the prostate to other body parts, in particular bones and lymph nodes.
  • Prostate cancer is one of the most common types of cancer in men, and its incidence increases rapidly after age 50.
  • Early prostate cancer is usually asymptomatic and is most commonly found on prostate specific antigen tests or rectal balance tests, which delay the treatment and lead to death due to cancer metastasis to surrounding tissues.
  • Prostate specific antigen is a serine protease that is expressed at high levels in the prostate epidermis with a molecular weight of 34,000 Daltons, and there are many kits that measure the amount of PSA (Prostate specific antigen) in the blood. In healthy men's blood, less than 2 ng / mL of PSA is present, and the amount of prostate cancer increases.
  • the increase in the amount of PSA in the blood indicates the development of prostate diseases such as prostate cancer and enlarged prostate, and the cut-off of PSA, which is mainly used for diagnosing prostate cancer, is generally 3 to 4 ng / mL. If the amount of PSA is more than 10 ng / mL, the probability of prostate cancer is 67%.
  • the ELISA type diagnostic kit using the conventional blood requires special equipment such as an enzyme-linked immunosorbent assay (ELISA) reader and automation equipment, and is applied in areas with poor medical conditions. There is a problem in that it can not be used, and it takes a long time to obtain a test result.
  • ELISA enzyme-linked immunosorbent assay
  • An object of the present invention is to provide a prostate cancer diagnostic sensor that enables rapid diagnosis at a high reliability and low cost and can be diagnosed at an early stage for easy treatment.
  • Still another object of the present invention is to provide a method for manufacturing the prostate cancer diagnostic sensor described above.
  • Still another object of the present invention is to provide a method for effectively and reliably diagnosing prostate cancer using the prostate cancer diagnostic sensor obtained at an early stage.
  • the present invention provides a paper-type sensing unit comprising an antibody-carbon-based nanostructure, and filter paper deposited by the filtering of the antibody-carbon-based nanostructure; And a detection unit capable of measuring a change in electrical resistance of the sensing unit, wherein the paper-type sensing unit is obtained by radially cutting the edge from the circle center of the filter paper after the syringe filtering deposition.
  • a detection unit capable of measuring a change in electrical resistance of the sensing unit, wherein the paper-type sensing unit is obtained by radially cutting the edge from the circle center of the filter paper after the syringe filtering deposition.
  • the antibody may be a prostate specific antigen (PSA), but is not limited thereto.
  • PSA prostate specific antigen
  • the carbon-based nanostructures may be selected from single-walled carbon nanotubes, double-walled carbon nanotubes, multi-walled carbon nanotubes, graphite, and graphene, but is not limited thereto.
  • the filter paper may be, for example, cellulose ester filter paper having micropores of 0.3 to 0.5 um in size, but is not limited thereto.
  • the paper-type sensing unit obtained by radially cutting the filter paper may be a rectangle of 5 x 2 mm 2 , but is not limited thereto.
  • the PSA concentration may be measured at a sensitivity of 1 ng / mL in a range of 0 ng / mL to 500 ng / mL, but is not limited thereto.
  • the diagnostic sensor may include a substrate on which a paper-type sensing unit is disposed; And electrodes formed at both ends of the sensing unit.
  • Another aspect of the invention provides a method of manufacturing a paper type prostate cancer diagnostic sensor comprising the following steps.
  • the carboxylation may be performed by refluxing the carbon-based nanostructures in 20 mL of 3.0 M nitric acid for 3 hours at 105 °C, but is not limited thereto.
  • the step of obtaining the antibody-bound carbon-based nanostructures the antibody receiving step of adding the antibody to the carbon-based nanostructures mixture and reacted by stirring at room temperature for 8 hours overnight;
  • the concentration of MWCNT is 6.5 ug / mL and the purified volume of Ab-MWCNT solution per filter paper is 200 mL.
  • Another aspect of the present invention relates to a method for diagnosing prostate cancer using the paper type prostate cancer diagnostic sensor.
  • the paper-type sensing unit in the wet state at 30 ⁇ 40 °C culture for 1 to 3 hours with a biological sample to perform an antigen-antibody reaction;
  • prostate cancer may be diagnosed at an early stage if the concentration of PSA antigen is greater than 4 ng / mL based on the relative electrical resistance (R / R 0 ).
  • the present invention provides a paper-type biosensor based on carbon-based nanostructures and filter paper.
  • the PSA level changed the electrical resistance of the sensor by more than 150%.
  • the maximum detection range is 500 ng / mL and the detection limit is 1.18 ng / mL.
  • Paper-based biosensors are 23 times cheaper and 12 times faster than ELISA-based detection methods. Moreover, the maximum detection value is about 50 times higher.
  • the detection range and sensitivity are sufficient to diagnose early stage prostate cancer. Biomarkers that specifically react with various target biomolecules as well as PSA can be applied to CNTs, which is expected to be a very useful diagnostic tool.
  • FIG. 1 is Scheme 1 illustrating a process of obtaining MWCNT-activated PSA antibody to obtain antibody-MWCNT to which an antibody is attached according to an embodiment of the present invention.
  • FIG. 2 is a schematic diagram illustrating a manufacturing process of a paper-type sensing unit and a PSA detection process according to an embodiment of the present invention.
  • FIG. 3 is a scheme 3 showing a process of diagnosing prostate cancer using a paper type prostate cancer diagnostic sensor according to one embodiment of the present invention.
  • FIG. 5 is an SEM and AFM image of MWCNT based sensor units; a and b are SEM and AFM images of carboxylated MWCNTs, c and d are SEM and AFM images of Ab-MWCNTs, and e and d are SEM and AFM images of PSA-Ab-MWCNTs.
  • FIG. 8 is a graph showing the relative resistance of biosensors at PSA concentrations in (a) wide range (0-1000 ng / mL) and (b) narrow and low range (0-10ng / mL).
  • FIG. 9 is a graph of resistance uniformity measurement results according to a cutting method of a paper-type sensor (a) resistance uniformity of the sensor cut in the longitudinal radial of the present invention and (b) resistance uniformity measurement result of the sensor cut in the horizontal radial .
  • the term "about” means 30, 25, 20, 25, 10, 9, 8, 7, 6, 5, by reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight, or length. By amount, level, value, number, frequency, percentage, dimension, size, amount, weight or length, varying by 4, 3, 2 or 1%.
  • Prostate cancer in the present invention is described, for example, in Prostate. In: Edge SB, Byrd DR, Compton CC, et al., Eds .: AJCC Cancer Staging Manual. 7th ed. New York, NY: Springer, 2010, pp 457-68, which is incorporated herein by reference, can be further subdivided into stages and grades of prostate cancer.
  • abnormal prostate conditions may be classified as one or more of benign prostatic hyperplasia (BPH), androgen-sensitive prostate cancer, androgen-insensitive or resistant prostate cancer, invasive prostate cancer, non-invasive prostate cancer, metastatic prostate cancer, and non-metastatic prostate cancer. have.
  • diagnosis means to identify the presence or characteristic of a pathological condition.
  • diagnosis is to confirm the development of prostate cancer.
  • antibody is used in its broadest sense and includes monoclonal antibodies, polyclonal antibodies, multispecific antibodies (eg bispecific antibodies), and antibody fragments, as long as they exhibit the desired antigen-binding activity. But not limited to them include various antibody structures. Diagnostic antibodies may be used as raw materials such as purified monoclonal anti-PSA, rabbit anti-PSA, goat anti-rat IgG, which are raw materials of conventional commercially available antibodies that can be used for the production of strips. In a preferred embodiment, the antibody may be a prostate specific antigen (PSA).
  • PSA prostate specific antigen
  • Another prostate cancer diagnostic sensor includes a paper-type sensing unit and a detection unit.
  • the paper-type sensing unit includes an antibody-carbon based nanostructure to which an antibody is attached, and filter paper deposited by syringe filtering of the antibody-carbon based nanostructure.
  • the present invention provides a paper-based sensor using carbon-based nanostructures and filter paper.
  • carbon-based nanostructures When the carbon-based nanostructures are formed into a mat, these mutual contact barriers determine electrical conductivity.
  • the antibody is attached to the outer wall of the structure, According to the content of the antigen to be detected, changes in contact barriers between carbon-based nanostructures occur, and electrical conductivity changes accordingly, thereby determining the concentration of biomaterials such as antigens.
  • Syringe filtering method is used to form the carbon-based nanostructures in the form of a layered mat.
  • Syringe filtering is performed by putting a solution containing carbon nanotubes into a syringe and then using a pressing pressure to filter the solvent. It passes through and the nanotubes are filtered out to form a mat.
  • the thickness of the deposited antibody-carbon-based nanostructure layer may be about 3 ⁇ 5um.
  • the cut portion radially cut the filter paper from the center to the edge is used as the sensing unit. Since the uniformity of the antibody-carbon-based nanostructure layer in the radial cut is the best, it is very effective to keep the basic resistance constant throughout the sensing unit.
  • the radial cut is a rectangular shape having a long length from the center of the circle to the edge direction, in one embodiment of the present invention used a rectangular shape of 5 x 2 mm 2 , but is not limited thereto.
  • the carbon-based nanostructure refers to a nano-sized structure made of a carbon component, and may include some of the reducing acid component included in the manufacturing process.
  • multi-walled carbon nanotubes are used, but not limited thereto.
  • single-walled carbon nanotubes, double-walled carbon nanotubes, multi-walled carbon nanotubes, graphite, graphene, graphene nano Plates and the like can be applied.
  • the carbon-based nanostructure is sometimes abbreviated as a carbon structure.
  • the filter paper is preferably used to form a uniform and fine pores (micro-pore) so that the antibody-carbon structure layer can be easily formed, in one example, micro-pores of 0.3 ⁇ 0.5 um size
  • micro-pores of 0.3 ⁇ 0.5 um size
  • the maximum detection range of the paper type prostate cancer diagnostic sensor according to the present invention is 500 ng / mL, and the detection limit is 1.18 ng / mL.
  • the contact potential between the carbon structures increases according to the antibody-antigen-binding reaction, indicating that the relative resistance change value is almost linear, depending on the concentration of the antibody.
  • concentrations were measurable with a sensitivity of 1 ng / mL in the range of 0 ng / mL to 500 ng / mL.
  • the diagnostic sensor of the present invention may be used to diagnose early stage of prostate cancer.
  • the detection unit in the diagnostic sensor substrate on which the paper-type sensing unit is disposed Electrodes formed at both ends of the sensing unit; and wires are connected to the electrodes to allow current to flow.
  • Such a detector may detect a change in resistance due to a change in contact potential between the carbon structures together with a charge transfer model between the biomolecule and the antibody-carbon structure that affects conductivity.
  • the substrate may be made of glass, silicon, or a mixture thereof.
  • the electrode may be selected from the group consisting of gold, platinum, chromium, copper, aluminum, nickel, palladium, and titanium.
  • Paper type prostate cancer diagnostic sensor can be used for diagnosing the development of prostate cancer in a subject with increased risk of prostate cancer expression.
  • Subjects with an increased risk of developing prostate cancer should be monitored for additional signs or symptoms of prostate cancer.
  • the diagnostic methods provided herein for identifying subjects with an increased risk of developing prostate cancer include, but are not limited to, other known risk factors or signals of prostate cancer, such as, but not limited to, reduced urinary stream, emergency, hesitation, nocturia, nouria (incomplete). bladder emptying) and age assessment.
  • a method of manufacturing a paper type prostate cancer diagnostic sensor according to an embodiment of the present invention will be described with reference to Schemes 1 and 2 of FIGS. 1 and 2.
  • Carboxylation step of attaching a carboxyl group to the carbon-based nanostructures 110
  • Binding an antibody to the carboxylated carbon-based nanostructure to obtain an antibody-bound carbon-based nanostructure 120
  • depositing 130
  • the antibody-bound carbon-based nanostructure on filter paper by syringe filtering.
  • radially cutting the syringe-filtered filter paper from the center of the circle toward the edge to obtain a paper-type sensing unit (140).
  • the carboxylation 110 may be carried out by refluxing the carbon-based nanostructure in nitric acid solution for 3 hours to 3 days at 105 °C.
  • Obtaining the antibody-bound carbon-based nanostructures 120 includes, in one example, adding an antibody to the carbon-based nanostructure mixture and reacting by stirring at room temperature for 6 to 12 hours; And adding a blocking agent to the antibody-receiving carbon-based nanostructure and incubating for 1 to 3 hours at room temperature to 40 ° C. to obtain an antibody-bound carbon-based nanostructure (122).
  • the non-specific antigen binding can be prevented in advance by first reacting the antibody with the carbon structure and then adding a non-reactive blocking agent such as BSA to attach to the site where the antibody is not bound in the carbon structure layer.
  • a non-reactive blocking agent such as BSA
  • the antibody-bound carbon-based nanostructure deposition step 130 then forms an antibody-carbon structure layer in which the non-specific sites obtained in the previous step are blocked on the circular filter paper by syringe filtering.
  • Syringe filtering methods are well known in the art and the known methods can be equally applied to the present invention.
  • the step of cutting the filter paper 140 is radially cut in a rectangular shape with a long length from the center side to the edge of the circular filter paper, the cutting method is not particularly limited.
  • a sample is put in a paper-type sensing part in which an antibody-carbon structure layer is formed to induce an antigen-antibody reaction (210).
  • the sensing unit should not be dried, but preferably, the reaction proceeds while incubating for 1 to 3 hours at 30 to 40 ° C. in a wet state.
  • the paper-type sensing unit is dried at room temperature for about 30 minutes to 2 hours (221).
  • the dried sensing unit is mounted on a substrate (slide glass) and connected to electrodes to assemble with a detector to measure electrical resistance (222).
  • the measured electrical resistance is compared with the basic resistance of the antibody-carbon structure layer to which the antigen is not bound to obtain a relative electrical resistance (R / R 0 ) value.
  • prostate cancer can be diagnosed at an early stage if the concentration of PSA antigen is greater than 4 ng / mL based on relative electrical resistance (R / R 0 ) values.
  • Nitric acid (64.0-65.0%) was purchased from DUKSAN science.
  • N- (3-dimethylaminopropyl) -N-ethylcarbodiimide hydrochloride (EDC), N-hydroxysulfosuccinimide sodium salt (NHSS), monoclonal anti-KLK3 antibody produced in mice (species reactive This human PSA antibody) and PSA (prostate specific antigen from human semen) were purchased from Sigma-Aldrich.
  • Microporous filter paper cellulose cellulose mixture, diameter 25mm, pore size 0.45um
  • MWCNTs multi-wall carbon nanotubes, diameter 20 nm, length 5 um
  • MWCNTs were produced by the CVD method using iron and molybdenum catalysts.
  • Scheme 1 shown in Figure 1 shows the process of activating the MWCNT and PSA antibody.
  • the detection system is performed by incubating at 37 ° C. for 1.5 hours.
  • the first step is the carboxylation step 110 of the carbon-based nanostructures.
  • MWCNT 300 mg was first refluxed in 20 mL of 3.0 M nitric acid at 105 ° C. for 3 hours to produce a carboxyl group on MWCNT.
  • the functionalized MWCNT solution is centrifuged washed 10 times or more with distilled water so that the pH approaches 7.
  • Carboxylated MWCNTs were dried for 24 hours in a 100 °C oven.
  • the carboxyl groups on the MWCNT surface were identified by Fourier transform infrared spectrometer (FT-IR, JASCO FT / IR-4100).
  • the carboxylated MWCNTs may receive PSA antibodies in the next step 120.
  • the carboxylated MWCNT (1.3 mg) is dispersed in 0.4 mL of 0.1 M MES buffer with 0.4 mmol EDC and 0.1 mmol NHSS.
  • the dispersion was sonicated for 1 minute with an ultrasonic shock absorber (Jeio Tech, UC-02) and then mixed for 15 minutes at room temperature with a vortex mixer (Scientific industies, Inc).
  • the obtained liquid mixture is centrifuged at 3000 rpm for 10 minutes (Hanil Science Industrial Co. Ltd., HA-1000-3), and the supernatant is discarded. Wash with PBS at least 5 times to remove excess EDC and NHSS from the precipitate.
  • 0.5 mL (0.01 mg / mL) PSA antibody is added to the mixture and stirred overnight at room temperature to attach to MWCNT. This process is PSA antibody acceptance 121 in the second step of Scheme 1.
  • the next step is to prepare an MWCNT-based biosensor to inhibit unwanted reaction between MWCNT to which the PSA antibody is attached and the target PSA (122).
  • a mixture of 0.05% Tween20 in 1% BSA (100 uL) is added to the mixture (2 mL) to which the PSA antibody is added to prevent adhesion of PSA on the MWCNT surface.
  • BSA completely wraps the MWCNT surface to prevent indiscriminate (nonspecific) PSA attachment.
  • Tween20 cleans the binding site of PSA antibodies. The mixture was incubated at room temperature for 1.5 hours, centrifuged at 3000 rpm for 5 minutes, and the supernatant was removed.
  • Additives are washed and centrifuged at least five times with PBS buffer to remove any FREE PSA antibody and BSA.
  • the blocking agent (BSA) is reacted with the MWCNT to which the PSA antibody is attached, thereby obtaining MWCNT to which the antibody is bound (122).
  • the obtained sample is called Ab-MWCNT, which means MWCNT to which PSA antibody is attached.
  • the Ab-MWCNT is deposited on the microporous filter paper by syringe filtering. This step is the first step of Scheme 2 (see Figure 2). Ab-MWCNT deposited on the microporous filter paper should not be dried until sensing is complete for the activity of the antibody.
  • Ab-MWCNT deposited on the filter paper is cut to a size 5 ⁇ 2 mm 2 in the circumferential direction from the center of the filter paper to obtain a paper-type sensing unit.
  • This radial cutting method is very important for maintaining the resistance uniformity of all sensing parts.
  • the paper-type sensing unit (PSA-Ab-MWCNT) prepared in Example 1-3 is a BSA blocked PSA-Ab-MWCNT deposited on the microporous filter paper.
  • paper-type sensors should not be dry. That is, it should be wet until the site location specific reaction between PSA antibody and PSA is completed. The site specific reaction was completed by incubating the sensing unit at 37 °C for 1.5 hours.
  • the paper-type sensing unit is continuously dried in air for at least 30 minutes at room temperature.
  • the electrical resistance (see Scheme 3) is measured.
  • the resistance of the dried paper-type sensing unit can be measured using a digital multimeter (Hewlett-Packard Co. multimeter 3478A or Agilant 34401) under atmospheric conditions.
  • a silver paste electrode is used to maintain electrical contact between the sensing unit and the copper wire.
  • Basic morphological information of the material is observed by atomic force microscope (AFM) and scanning electron microscope (SEM, HITACHI S-2400).
  • the sensor resistance measured for a specific concentration of PSA was incubated for 1.5 hours at 37 ° C in a paper-type sensor in contact with a PBS buffer solution (100 uL) containing no PSA (PSA concentration 0 ng / mL).
  • the basic background resistance which is the resistance measured after drying.
  • the measured sensor resistance is normalized taking into account the physical shape of the sensor.
  • the thickness of the sensor is not taken into account. This is because the background resistance is compared with all resistance values, and all the sensors are made of one filter paper. In order to keep the basic resistance value of the sensor constant, the most important factor is the uniformity of the MWCNT layer deposited on the microporous filter paper.
  • Uniformity of the MWCNT layer can be obtained by adjusting the concentration of MWCNT to 6.5 ug / mL.
  • the volume of Ab-MWCNT solution used per filter paper is 200 mL.
  • the thickness of the deposited MWCNTs was ⁇ 4.5 um.
  • the radial cutting method of the sensing unit cut into a circle from the center as shown in Scheme 2 of Figure 2 is very important to maintain a constant resistance.
  • This method of measuring the concentration of PSAs has the advantages of portability, time savings and cost savings over the widely used ELISA.
  • Figure 4 shows the IR spectra of MWCNTs, immersed in nitric acid for 3 days, carboxylated MWCT, and Ab-MWCNTs.
  • the peak position corresponding to the carbon-carbon double bond of MWCNT is found at 1630 cm ⁇ 1 .
  • the carboxyl group may be located on the surface of the MWCNT and react with the amine group of the L-lysine unit in the antibody. This is a simple substitution reaction to form an amide group.
  • the carboxylated MWCNTs are treated with EDC / NHSS to induce the hydroxyl group of the carboxyl group to react with the antibody.
  • the IR spectrum of Ab-MWCNT is shown as a blue line in FIG. 1, with complex peaks corresponding to amide bonds around 1550 cm ⁇ 1 .
  • 2 a and b are SEM and AFM images of carboxylated MWCNTs, respectively.
  • the diameter of the MWCNTs in both images was about 20 nm. This corresponds to the diameter of the receptor MWCNTs. No damage was observed on the surface of the carboxylated MWCNTs. All MWCNTs were distinguishable.
  • 5 c and d are SEM and AFM images of Ab-MWCNT. Observation of the diameter of the antibody-attached MWCNTs showed similar results to the carboxylated MWCNTs. Any irregularly attached object was observed on the tube surface. This means that the attachment of the antibody may not be coated all over the MWCNT surface. Clumps such as cotton balls on SEM images are presumed to be attached antibodies.
  • 5e and f are SEM and AFM images of PSA-Ab-MWCNT.
  • Most SEM images of MWCNTs are covered with proteins (PSA antibodies and PSA).
  • PSA antibodies and PSA proteins
  • the cottony areas spread over the entire surface of the deposited film.
  • the shape of the cottony part is not clearly observed in the AFM image due to the softness of the protein.
  • the densities of the selectively observed deposition films were found in the order of carboxylated MWCNT Ab.MWCNT> PSA-Ab-MWCNT. From the AFM image of the film, RMS (effective value) roughness appeared in the same order as visual observation.
  • MWCNT, Ab-MWCNT, and PSA-Ab-MWCNT were 38, 32 and 61 nm, respectively.
  • Table 1 also shows the relative resistance values of MWCNT, carboxylated MWCNT, Ab-MWCNT, and PSA-Ab-MWCNT calculated in FIG. 6.
  • the electrical resistance of each MWCNT is listed in Table 1 and can be estimated from the straight line slope shown in FIG. Electrical resistance was increased by 500% by carboxylating MWCNTs. A decrease in the carbon-sp 2 nature that is the source of conductivity during the carboxylation process in MWCNTs can lead to this increase in resistance.
  • PSA antibody was attached to the carboxyl group
  • the electrical resistance of Ab-MWCNT was increased by about 160% higher than that of carboxylated MWCNT.
  • the increased resistance is thought to be due to the increase in the contact potential barrier between the MWCNTs by further attachment of the protein (PSA-antibody).
  • PSA-Ab-MWCNT PSA, 1000 ng / mL
  • This stepwise increase in CNTs is clearly shown in FIG. 4.
  • FIG. 7 shows the change in electrical resistance due to MWCNT surface modification.
  • Data points show MWCNT, carboxylated MWCNT, Ab-MWCNT and PSA-Ab-MWCNT (PSA; 1000 ng / mL) from left.
  • the conductivity will increase (off current increase), and the threshold voltage (V TH ) of the field transistor (FET) As the p-character increases, it moves to a more negative voltage. If biomolecules provide electrons to the CNTs, the opposite will be observed.
  • the increase in the contact energy barrier between conductive CNT strands by adsorption of biomolecules on the CNT surface leads to reduced conductivity without changing the V TH of the FET.
  • the number of target biomolecules that can be attached to the CNT can be determined by the change in resistance of the biomolecule-CNT and the V TH shift of the FET.
  • the shift of V TH due to the addition of biomolecules could not be observed, only a decrease in the off-current of the FET by exposure of the biomolecule It could be observed.
  • FETs assembled using metal-attached CNTs have excellent hydrocarbon detection capability for this reason.
  • Relative electrical resistance (R / R 0 ) caused by reaction of sensing unit (Ab-MWCNT) with PSA (where R is the resistance value of sensor that detects PSA at a certain concentration, and R 0 is the buffer solution that does not contain PSA) 8 is shown in FIG. 8.
  • FIG. 8 (a) shows that the change in the relative resistance is almost linear, depending on the concentration of PSA, in the range of 0 to 500 ng / mL. This means that if the PSA concentration is within 500 ng / mL, the concentration can be estimated by measuring the electrical resistance of the sensor. However, if the concentration of PSA is> 500 ng / mL, the resistance of the sensing section appears to be saturated.
  • the detection limit was 1.18 ng / mL.
  • the standard deviation was found to be 0.00332 from 11 sensors made by the longitudinal radial cut, and the R / R 0 value ranged from 0.99 to 1.00.
  • the resistance uniformity was shown to be constant. This results in no problem in detecting 1.18 ng / mL, which is the detection limit of the present invention sensor.
  • the paper-type sensor of the present invention uses two sensors, Background Detection Indicator and Indicator Detection Sensor, to eliminate errors that may occur due to humidity, temperature, and surrounding environment, respectively. Calculate PSA concentrations as R / R 0 values. Therefore, in order to compare the resistance of the background and indicator, the two detectors must have the same initial resistance.
  • the standard deviation of the electrical resistance obtained from 11 sensors made by the transverse radial cut was 0.0438, which is 4 ng / mL. Since PSA is larger than the detection signal, it is not suitable for sensor manufacturing. In addition, the R / R 0 value was measured as 0.91 as the lowest value and 1.06 as the highest value, thus confirming that the value was not constant and thus not suitable for the PSA detection method of the paper type sensor of the present invention. Failure to ensure resistance uniformity greatly reduces the reproducibility of the experiment and the accuracy of PSA concentration measurements.

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Abstract

La présente invention concerne un capteur de type papier pour le diagnostic du cancer de la prostate à l'aide d'une nanostructure à base de carbone et d'un papier filtre, un procédé de fabrication de ce dernier, et un procédé de diagnostic du cancer de la prostate l'utilisant, et ainsi, la présente invention permet, un diagnostic aisé, à faible coût et rapide du cancer de la prostate par comparaison avec un procédé existant, ELISA. En outre, la présente invention peut permettre le diagnostic de maladies par l'application à divers biomarqueurs, et peut ainsi être largement utilisée dans un diagnostic médical large.
PCT/KR2016/002808 2015-08-06 2016-03-21 Capteur de type papier pour diagnostic du cancer de la prostate, son procédé de fabrication, et procédé de diagnostic du cancer de la prostate l'utilisant WO2017022923A1 (fr)

Applications Claiming Priority (2)

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