JPS63500539A - Sensor with ion selective electrode - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

[Detailed description of the invention]

Sensor with ion selective electrode explanation background ion selective electrode Ion-selective electrodes respond preferentially and selectively to specific ion species in a liquid. They are used in potentiometric measurements of the activity of ions in liquid samples. . Potentiometric measurements measure the voltage between two electrodes that form an electrochemical cell in contact with a liquid. Measure the difference in position. The half-cell potential of one electrode - the reference electrode - is essentially constant; That of the other electrode - the indicator electrode - varies with the ionic activity of the liquid being analyzed. Ru. The potential across the electrode is proportional to the logarithm of the activity of the ion to which the ion-selective electrode responds. Ru. The Nernst equation describes this logarithmic relationship. The potential difference is like an electrometer Can be measured using a potentiometer. Several types of ion-selective electrodes are available, e.g. Includes conventional glass electrodes for pH measurements. Glass electrode is alkali ion silicic acid Based on salt composition. The electrode for pH measurement is lithium silicate glass or borium. It can be made from silicate glass and is permeable to hydrogen ions (H+). However, the anion or fc is not permeable to other cations. Selected for H+ When a selectively permeable glass thin film is placed between two solutions of different H+ concentrations, the H+ + ions move across the glass from a high concentration solution to a low H+ solution. Such transfer results in the addition of positive ions to solutions of low U+ concentration at °C, glass It leaves negative ions on the other side, creating an electrical potential across the glass. References independent of pH Dipping the electrode into the solution completes the circuit and allows the potential difference to be measured. Another type of ion-selective electrode uses a liquid ion exchanger, cellulose acetate. or supported by an inert polymer such as a film of polyvinyl chloride. be done. An important example of this type of electrode is the calcium salt of an oil-soluble diester of phosphoric acid. This is the basic Ca-responsive electrode. U.S. Patent 3,429.785; 3.438 ．． 886; and 3,445.365#'i, divalent cations (e.g. c!L2 +, M g 2 +) Ion exchange organic liquid that selectively responds to K An ion-selective electrode with a second layer filled with a porous inert material is also used. It is listed. Neutral carrier-based sensors for monovalent and divalent thiones are ion-exchange Similar to body-based electrodes. Both contain ion exchange sites, especially mediator Negative mobile site or carrier provided by a substance (mediator) contains negative fixation sites that result from hydrolysis of the molecule. Neutral carriers, they are cyclic chains or 7' (can be an open class, but is generally in a hydrophobic complex form with a cation) It is a compounding agent. Such compounds can be selectively extracted for K'', H&'' and Ca. , thus resulting in selective permeability, and those ions d otherwise simple As an inorganic salt, it does not dissolve in agricultural phase. An example of such a carrier is palinomycin, It can be used in electrodes that respond selectively to +. Electrodes for measuring the H+ content of liquid samples have been described by others. They are Commonly, ion-selective components (ion permeable carriers (1onophore) and their Plastics with solvent/plasticizer compounds capable of dissolving ion-selective components Contains a protective membrane. A glass frame that is selectively permeable to H ions. Oxidative phosphorylation uncoupling agents such as lipophilic derivatives and lipophilic tertiary amines Hydrogen ion permeable carriers have been used. For example, Simon et al. describes a micro pH 1f electrode containing a hydrogen ion-permeable carrier (Alamine) as a hydrogen ion permeable carrier. Man analytic Chemistry 53:2267-2270 (1981 ). The authors found that this electrode had excellent response to pH changes between 5.5 and 12. I am reporting that I will give you. However, there are at least two important drawbacks when properly It arises from the fact that in order to function, the agricultural phase must contain dissolved carbon dioxide. Cheating. First, the electrode performance (i.e., CO□ content) due to changes in humidity temperature and pressure. It affects me badly. Second, mass production requires careful control of f'Lfc conditions. It is extremely difficult to do so, and moreover, dissolved Co diffuses away during electrode storage. Other types of ionophores act as uncouplers of oxidative phosphorylation in mitochondria. Basic. An example is described by Finklestein. Biochim ica Biophysica Acta. 205:1-6 (1970). 2.4-dinitrophenol and 1-chlorophenol Weakly acidic uncoupling agents, such as phenylhydrazone mesooxalonitrile, support H+ ions. Acts as a body. those measles

[1. Membrane components that are incorporated into ion-selective electrodes are unsuitable due to their certain water solubility (ie, they do not remain bound to the membrane and leach out of the membrane). Brown et al. describe a pH sensor that is claimed to be suitable for long-term intravascular implantation. The pH-sensing element is a thin film of elastomeric polymer made permeable to ions through the addition of a hydrophobic and lipophilic specific H+ ion carrier. The carrier used was p-octadecyloxy-m-chloro7-ether hydrazo/menoxalonitrile (OCiPH, where f'L is weakly acidic conjugate theory-chlorophenylhydrazone mesooxalonitrile). Salonitrile polymer! is a homologous series). Brown and co-workers suggested that although OCPH molecules act as mobile C carriers in each elastomer, pH-responsive properties of a quality good enough for practical applications are guaranteed by a point specially devised for the purpose. It is stated that only a small amount can be obtained using a reamer matrix. US special A3, 743, 588 (1973); 0, H. Leblanc, J.F. Blau Journal of Applied Physxology, 40. : 644-647 (1976) The preparation of polymers that can be used with 60GPH molecules has been described by Vaughn. US Pat. No. 3,189,662 (1965); H.A. Vaughn and J.P. Goldberb's Poxylier Letters. 7: 569-572 (1969). Rice 11! No. 3,691.047 C1972), Ross and Martin describe an ion-sensing membrane for potentiometric measurements. The membrane is described as a gelled mixture in which the solid phase is a polymer (e.g. cellulose triacetate) and the liquid phase is an organic solvent dissolved in an organic solvent. organic ion exchange materials (e.g., carbon dissolved in dioctyl phenylphosphonate) It has been stated that sium (bis-dibromooleyl-phosphate) is l. Ru. Its ion-sensing electrode consists of an organic ionizer whose main component is dissolved in a non-volatile organic solvent. It is said to have a membrane that is an on-exchanger. In US Pat. No. 4,214,968 (1980), Battaglia et al. describe a dry operable ion-selective eleven pole for use in determining the ionic content of liquids. The electrode consists of a dry internal reference electrode in contact with a hydrophobic ion-selective membrane. It is being said. Its internal reference %: the pole is dry [primary metal/metal salt reference semi-℃ pond or the pole is dry redox couple Sanfu 1 and wet when applying the liquid sample. Let's go. The ion-selective membrane contains an ion carrier (eg, palinomycin), which is dispersed in an aqueous binder and dissolved in a carrier. 9; (6 (1980), Ball and Baba Ogle describes an apparatus for tracing and determining ionic activity in liquid samples. In that case, one with an ion-selective membrane containing vr4i, silica, and an ion-selective membrane on the support (- I haven't said enough Ru. The two electrodes of the device are solid and are preferably said to be dry. In U.S. Patent No. 4,053,381 (1977 N'c), Hamplen et al. described another device for measuring the ion activity t in a liquid. @pole is an ion selector whose internal reference electrode has several layers. Including the Selection Jugokukai. Their Nk is a metal layer, a layer of insoluble salt in the metal layer, and a preferable or dry chicken W, containing a liquid-containing layer. The claimed device includes a solid electrode. There are currently many types of ion-selective electrodes available for measuring the ionic content of liquids. Exists. However, these ion selective electrodes have limitations. These limitations require that membranes made with specially designed polymeric polymers (IJ Luxm) be pre-neutralized with base to improve membrane sensitivity and reduce response time. The need for storage under well-controlled conditions; and reduced sensitivity and reliability during storage. Additionally, currently available ion-selective electrodes at a constant pH require relatively large samples (i.e., 1.0 ml+ or more) and are expensive for accurate operation. Made of high-strength glass and suitable for automatic processing of extremely small quantities of samples. cannot be incorporated into electrodes that Ion selectors that can be used to quickly and accurately measure the ion content of samples smaller than those encountered in current applications (e.g. microliter size) There are Also. There also exists a need for ion-selective electrodes that can accurately and more rapidly measure other constituents of smaller samples than is possible with currently available electrodes. Differential Measurement Techniques Differential potentiometry techniques rely on the potential difference that occurs between two equivalent electrochemical half-cells separated by a salt bridge and immersed in solutions of different activity. These two The half-cells together constitute a concentration cell. In this case, one half Pond activity (ILl) F! It is fixed (8M), and on the other hand, the active f(a(trial) EL of the battery is defined as follows. Varying: Here, m59.1mv, (for 5m=1 F at 298°K). l, = emf of the reference half-cell E, = amf of the sample bovine cell 34o = standard reference electrode potential R2 molar gas constant; 8,314 volts e coulombs to 70 = absolute temperature Ox n = charge on the ion F = Faraday constant Number; 96,493 coulombs a1 and a, = io of reference sample In the case of a cation such as potassium (K+), the half-cell a is used as a reference. If defined as a photovoltaic cell and assigned zero potential, the emf of the concentration cell is positive if a, )a, and a! If < a t, it is negative. , t Ll are fixed, so the equation for the battery potential F contains only 11 unknowns (a, )'e, and by measuring Kcel'1, the equation can be solved for h. Differential measurement techniques detect components of biological fluids other than H+, including other ions such as sodium (Na''), potassium (K''), calcium (Ca++) and chloride (C-!-). It has been used to indirectly measure the concentration or activity of Furthermore, such techniques often utilize biosensors or enzyme electrodes, which are the products of a biologically catalytic reaction or a biological catalyst coupled to an electrode that is sensitive to a cosubstrate (e.g. immobilized enzymes, cells, tissue layers). Enzyme or substrate concentration can be determined using differential measurement techniques. For example, many enzymatic reactions result in the release of bases. The ionization of the acid or base results in the release and uptake of H+ and a change in the pH of the solution. The measured change in H+ concentration or pH releases hydrogen ions, which can be the basis for stoichiometric measurements of the concentration of uptake substrates (eg, glucose, urea, etc.). In the case of differential pH measurements, each half-cell has p)I! Including poles. The hydrogen ion activity of the a1#tli pond is fixed and varies depending on the pH of the sample to that of -1 (sample). It can be used to calculate the hydrogen ion activity of the emf order Ka, which is measured across the cell by an electrometer. For differential measurements that require an enzyme, place the enzyme in one half-cell or both half-cells and add the sample to both half-cells, or add the sample to one half-cell. and calibrator to the other half cell. As a result, when the enzyme reacts with the substrate in the sample, an increase or decrease in pH occurs; the magnitude of the change is directly proportional to the amount of substrate in the sample. Similarly, the substrate can be replaced with an enzyme and the cell can be used to measure the enzyme activity of a given sample. For example, Nirunn and co-workers have used hydrogen ion glass electrodes to create enzyme-pH electrodes that measure glucose, urea, and penicillin in solution. It describes the development of poles. The enzymes used in this measurement are glucose oxidase, urease, and and penicillinase. % Biochimia et Biophygica Acta, 320: 529-534 (1973). Moss and co-workers have also described the determination of glucose concentration by differential pH measurements. The technique is based on the measurement of the pH change produced by the hexokinase catalyzed reaction between glucose and ATP. They describe two systems that are said to be useful for measuring pH differences between 1 ml of aqueous samples. The concentration of glucose is determined by its measured pH change according to the formula derived by those authors. Calculated from . Soska, A. et al., Analytical Biochemistry, 112: 287-294 (1981). A differential measurement of pH for measuring glucose in whole blood and plasma and an automated system for doing so have since been described by this group. C11nicalC h s m18 try I 29: 80 85 (198A) of Lutsana 1M, et al. The same device and differential pH technique has been described for use in measuring lipase activity in biological fluids such as serum, plasma, and decadal fluid. C11nical Chemistry of Kariotsuta F, et al. 31; 257-260 (1985). The refinement of this differential pH measurement technique As a basis for the determination of urea, creatinine and glucose in plasma and whole blood It is said to be helpful. Enzyme, urease, creatinine iminohydrolase, and hexokinase are used for their measurement. The concentrations of these three substrates are calculated based on the observed changes in the pH of the solution after the reactions have taken place. In U.S. Pat. No. 4,353,867 (1982), Lutsana describes an apparatus and method for measuring substances such as glucose, urea, and enzymes in biological solutions (e.g., blood, serum, urine). ing. The method uses differential pH measurement, where 21 glass pH electrodes are placed in separate solutions. The pH change in the solution after the reagent is added is measured, and the concentration of the substance of interest is calculated from the observed pH change. The device also includes a sample cuvette and two glass capillary electrodes. one cell, a means for measuring the pH at these two electrodes, and a means for measuring the pH at the two electrodes. electronic means for calculating the concentration of a substance. Immunosensor Electrochemical immunosensor It may be described either as a differential measurement, or as a current measurement. electric Differential immunosensors can be used to measure either antibodies or antigens. Wear. They may be described as either direct or indirect, and can be either membranes or solid electrodes. An example of a direct potentiometric immunosensor for antigen measurement is described by Yamamoto et al. An antibody, anti-hcc, is immobilized on titanium wire. Gently connect the anti-hcG electrode and the reference electrode! Place in solution. As the antigen, hCJf, is added and the antigen binds to its immobilized antibody, the potential difference between the two electrodes changes until it reaches a flat wave. Its equilibrium is directly proportional to antigen prime. Yamamoto et al., C11nical Chemistry, 26: 1 569-1 572 (1980). Although the exact nature of the potentiometric response is not fully understood, it is generally recognized that it involves surface charge neutralization and redistribution. Another example of a direct potentiometric immunosensor is described by Keating and Rechnits. This type of immunosensor uses marker ions. On the contrary, the g-body responds to a specific antibody in a manner such that the change in potential is proportional to the amount of the antibody at one time. An immunosensor for the measurement of digoxin antibodies has been described, in which digoxin is bound to the marker ion carrier molecule benzo-15-crown-5. The resulting complex is assembled into a polyvinyl chloride membrane. J. et al., Keating, J. Y., and Rechnitz, G., Analytical Che+++igtry, 56:801-806 (1984). Antibody-selective potentiometer electrodes for antibody measurements were developed by Hinitsu and Tsursky. as described in US Pat. No. 4,402.819. A significant limitation to routine clinical use of this method is the need to maintain a constant background level of marker ions. Therefore, biological samples must be analytically dialyzed. Indirect potentiometric immunosensors are enzyme-linked and similar to enzyme immunoassays, except that they use electrochemical sensors to measure the production of enzyme-substrate reactions. Both homogeneous and heterogeneous potentiometric enzyme immunoassays have been described for the determination of estradiol, e.g., Boathio et al. describes a heterogeneous potentiometric enzyme-linked immunoassay. Boachio et al., C11nical Chemica Acta, 113, 175-182 (x981), antibodies against estradiol were prepared using porous gelatin. immobilized onto the membrane. The membrane is treated with peroxidase-labeled estradiol. Keep warm with isolated estradiol. After washing, the membrane is fixed onto an iodide tube sensing electrode. Peroxidase activity is measured in the presence of hydrogen peroxide and iodide ions. The iodide tube selective electrode potential is a function of estradiol concentration. A homogeneous potentiometric enzyme immunoassay for human IgG was developed by Huonong and Lech. stated by Nitsu. The method ij, Chlorope conjugated to XgG antibody oxidase. CO□ Raw with Beta Keto Adipine Distilled It is based on the formation and closure by IgG. Phonong and Rechnitz G., Analytical Chemistry, 56:2586-2590 (1984). When the D enzyme-xgeQ complex is incubated with a sample containing antigen (IgG) prior to reaction with the enzyme substrate, observation of CO2 release, measured with a potentiometric CO2 gas-sensing electrode, is required. The decrease in activity is proportional to the amount of IgG in the sample.11 Flow-metric enzyme immunosensors use an amperometric sensor, usually an electrode, to absorb the enzyme. Except that it is used to measure elementary activity. Similar to potentiometric immunosensors. For example, Aizawa, Mori Saiki, and Suzu An amperometric immunosensor for the measurement of the tumor antigen alpha-7 ethoprotein (AFP) is described. Aizawa et al. Analytica Chimica ActlL, 15: 6-67 (1980). The antibody FP is covalently immobilized on the porous membrane. Membranes were combined with catalase-labeled AFP and free AFP. Keep warm together. After competitive binding, the membranes are tested for catalase activity by amperometric measurement of oxygen after addition of hydrogen peroxide. In a similar manner, Boachio and co-workers have described an amperometric enzyme immunoassay for the measurement of hepatitis 8 surface antigen. Boatio et al., C11nic&l Chemistry, 25:318-321 (1979). The limitations of enzyme-linked amperometric sensors are the necessity of enzyme-linked synthesis and the use of oxygen electrodes. One reason is that currently available amperometric sensors are expensive. Disclosure of the Invention The present invention is a sophisticated sensor that potentiometrically measures the ion content or activity of a sample and the concentration of other components of the sample by using an ion-selective electrode. This sensor measures the hydrogen ion content and pH of the biological fluid; the pH of other ions of the biological fluid; and other components of the biological fluid, such as by differential pH measurement or immunoassay techniques. , glucose, urea, triglycerides, uric acid, aspartate amino Transferase (ACT), alanine aminotransferase (ALT), amylase, creatinine kinase (C), alkaline phosphatase, and enzymes are particularly useful for automated handling or processing. This sensor consists of ion-selective electrodes that are held in a frame. and between them a porous material (e.g. made by Borex Chichnorrhea). porous polyethylene), such as those manufactured by This porous matrix provides a means of concentrating ionic flow between the electrodes when the sample is applied to the electrodes. child These ion-selective electrodes Fiapt consist of a membrane that is selectively permeable to the ions or other substances to be measured and a reference electrode. The membrane requires pre-conditioning before use. do not have. A selectively permeable membrane consists of an ion-selective compound (or ion-permeable carrier) and a thermoplastic resin or plastic material, all of which are soluble in organic solvents. Additionally, the membrane can also include a plasticizer. The ion-selective component for the pH sensor is a compound with the following general formula, where R,, R, and R1 are independently selected and each has a number of carbon atoms of 4- 18 alkyl groups; 3) halogen-substituted alkyl group: 4) alkoxy group: 5) halogen-substituted alkoxy group; 6) 40. Acid groups represented by RK, where R4 is alkyl of 1-18 carbons: 7) Keto groups, represented by -COR, where R3 is defined for R4 and is selected from nine groups: or. 8) Hydrogen atom; In one embodiment of the invention, the membrane ij organic plastic matrix, polyvinyl Nil chloride (pvc), which is an ion-selective compound 2-oftadec 5-carbethoxyphenylhydrazone containing mesooxalonitrile. This ion-selective compound, pvc, and 2-nitrophenyl in this embodiment. All plasticizers, which are octyl ethers, are soluble in the solvent tetrahedral CI7 (THF). These ingredients make up the membrane formulation. In a preferred embodiment, the membrane is comprised of approximately 10-40 inches of PVC with a thickness RI greater than 1 mil. In a particularly preferred embodiment, it is made of about 20-354 PvC by weight and is about 3-15 mils thick. The membrane described can be used as a component of the ion selective electrode of the present invention. So? l is also commercially available. It can also be incorporated into a hand-held electrode body. The sensor of the present invention also has an internal reference element or substance that contains the known quality of the sample component to be determined. Figure IFi is a perspective view of the top of a sensor with an ion selection force pole and a handle; Does the hand have a magnetic stripe (5tripe) that records cheetahs? I have it. FIG. 1j is a perspective view of the underside of the sensor with an ion-selective electrode and a handle with a magnetic stripe on which data is recorded; Figure 3 shows the hydrogen ion activity f (pH) of the sample or the concentration of other ions in the sample t-measurement. 1 is a perspective view showing the individual components of a sensor that can be used to determine the FIG. 4 is a schematic representation of the ion-selective electrode arrangement. Figure 5#'i Ion of the invention inserted into a commercially available barre/L/type electrode. Figure 2 shows a selective membrane. Figure 6Vi used to measure the activity or once of a component of a sample by differential measurement technique. 2 is a perspective view showing the individual components of a sensor that can be The sensor that is the subject of the invention is used for potentiometric determination of the ionic content of a sample or the concentration of other components of the sample. Hydrogen ions in biological fluids (e.g. blood) It is useful for rapid determination of the concentration of other ions (e.g. glucose, urea, triglycerides, uric acid) and enzymes such as aspartate aminotransferase (AST) in biological fluids. It is particularly useful for measuring concentrations of alanine aminotransferase (ALT), amylase, creatine kinase (CK), alkaline phosphatase). An ion permeable carrier which is selective for sample components and which can be incorporated into the ion-selective membrane of a sensor of this type is also a subject of the present invention. This sensor will now be further described with reference to the figures. Figures 1 to 3 show an ion-selective electrode and a magnetic strip with two data points. With your hands? It shows the sensor with The sensors shown in these figures can be used to measure the hydrogen ion content of a sample or the pH or other ions in the sample. These figures are referred to herein when describing the sensor used to measure the hydrogen ion content of a sample. However, it is clear that the sensor can be used to measure other ions or other components as well. One sensor 30 consists of an ion selective electrode held within a frame or body 10 having an upper portion 12 and a lower portion 14. Furthermore, the sensor 30t/'i has a magnetic stripe 8 on the lower surface of the upper part 12F of the handle 5 having fourteen openings 4 and three grooves 6 disposed between the openings. The lower part 14 of the frame 10 has four open lowers and three grooves (not shown) located between the openings.The upper part 12 and the lower part 14a, the opening 4 and the open lowers are aligned in a line and the grooves 6 in the upper part are aligned with the grooves in the lower part.As a result, the openings 4 and 7 form four openings 11, and the grooves 6 in the upper part and the grooves in the lower part are aligned. The groove defines the space between three of the apertures 11 in the section. The upper section 12 has small holes 15 located between the apertures 4. The holes 15 allow the passage of air. The ion selective electrode is It is placed inside the opening 11 and has a cylindrical or or by a porous tubing 17 in the form of a rod, which provides a means for ionic flow between the KvL electrodes when a sample is applied to the electrodes. This cylindrical shape or Fi frame rod shape A porous material 17 is inserted into the spaces between the openings 11. The upper part] 2 and the lower part] 4 are placed on the porous material 17, and the frame or 7t is the body. A seal is formed between or within the two parts of the body 10. As shown in Figure 3, the sensor 30 has four positions in which ion selective electrodes can be placed. I have a place. Typically, one ion-selective charge is applied at each of these positions. Although there are poles, these can be put into various combinations depending on the analytical considerations desired. It can be used as a stand. Therefore, the positions of the grooves 6 in the upper part 12 and the grooves in the lower part 14 vary as necessary for the analysis to be performed. For example, *6 in the upper part 12 can be located as shown in FIG. The three electrodes are positioned so that a space is defined between them. Alternatively, the 2" grooves 6 can be placed parallel to each other in the upper part and the grooves in the lower part 14 can be located correspondingly. I can do it. Two, three, or all four electrodes can be used. The porous ridge 17 is either hydrophilic and electrically conductive or hydrophobic and non-conductive. The use of conductive materials or non-conductive materials between a pair of electrodes is determined by the analysis to be performed. The various combinations of dark are best described by examples. In the first example, as shown in Figure 4a, three membranes are active; All contain the same membrane. Are positions ta and b the known quality of the base ion being measured? too Position 1 C has a sample containing an unknown concentration of the target ion. Between a and b The smf generated between the solutions at b and C can be used to calibrate the sensor, and the slope W is then used to determine the #summer for that sample from the aunt generated between the solutions at b and C. determined. Furthermore, since the ion concentrations in a and bO are known, the pre-measured slope Using the slope value, calculate what the chamber potential difference between a and b should be, measure this value and compare it to the nine potential differences, or calculate once for #''taK and find this is the known value. mosquito It is possible to determine how the In a clinical chemistry sense, the 1 eL solution in a is then used as a reference standard. This a, b, a configuration can also be used in such a way that repeated samples can be analyzed. This is shown in Figure 4b. In this case, the same sample is placed in positions a and cKt, and the reference solution is placed in b. By using pre-measured slope values, two values can be determined simultaneously for the same sample. It is also possible to measure two analytical components simultaneously. This is shown in Figure 40. It is. For example, a and d can be membranes selective for one analyte, and b and c #: can be membranes selective for another analyte. There are no useful combinations. Thorium/potassium, pH/calcium, glucose/urea, and urea/crease It can be atinine. In these cases, predetermined calibration data for Kd is required. The enzyme/substrate sensor can be constructed as shown in FIG. 4. In this case, a, b, c and d can all be the same membrane1, e.g. All of these can be for pH measurement. However, a and d contain immobilized enzymes in addition to the pH membrane. An unknown analyte sample concentration is added to both C and d, fl, and a known concentration of the same analyte is added to a and b. The enzyme in turtle and d acts on the substrate and This produces a pH change that is proportional to the concentration of the analyte in the sample. Assuming that all membranes are equal, the 6-ring f generated between the tortoise and b (reference solution) is used to calibrate the electrode. I can be there. The calibration data obtained for a and b are then applied to the samples in C and d. It can be used to calculate the concentration of The enzyme/substrate sensor can be configured as shown in Figure 4K. In this case a, b and cFi are all the same membrane, for example they are all for pH measurements. However, in addition to the tz-sc, b and C pH membranes, it contains an immobilized substrate. The substrate concentrations in 1 and b are different and can be the same as the substrate concentrations in C Fia and b, or can be different. An unknown concentration of the analyte enzyme is added to C, and known concentrations fa and b of the enzyme to be measured are added. Turtle, b, and. The enzyme in a, b and c acts on the substrate, producing a pH change that is proportional to the enzyme concentration in the sample. All membranes are the same. Assuming that, the smt occurring between a and b (reference solution) can be used to calibrate the electrode. The calibration data obtained for a and b may then be used to calculate the enzyme sample concentration in C. can be immobilized in an enzyme/substrate sensor as represented in Figure 4k. Examples of substrates that can be used include amino acids such as L-alanine and L-aspartate, Reatine and starch. These are fl, alanine aminotransfer, respectively. Laase. Aspartate amino-transferase, creatine kinase, and aminotransferase It can be used for the measurement of analysinase in a sample. As described in Example 8 The enzyme/substrate as represented in FIG. 4k is also the sample creatine! It can be used to measure 1 degree. If a kinetic rate method is used, this methodology effectively eliminates any temperature effects on enzymatic reaction rates. In the case of endpoint or equilibrium measurements, the temperature effect in the Nernst equation is normalized. 17 can be electrically conductive or non-conductive, and the electrical conductivity between a pair of electrodes is determined by the use of non-conductive materials and analysis. Materials include nylon, polypropylene, polyethylene, and polyvinylidene fluoride. It can be a microporous plastic such as oleide, ethylene vinyl acetate, styrene-acrylonitrile, or polytetrafluoroethylene. child The porous material can also be a roll of filter paper or a length of fiber. - Can be cellulosic with bundle-like properties. in one embodiment This porous material is made from ultra-high molecular weight microporous polyethylene (Volex technology). Consisting of a cylindrical shape chamber (Logis, Inc., Fairburn, GA). This substance is hydrophobic in nature so that it is not water i'qfi. This porous material can be made hydrophilic by immersing it in a solution of a surfactant (approximately 0.1 to 0.6 inches by volume). Lfc surfactants found to be useful include nonionic surfactants such as Triton Polyoxyethylene ethers such as octylphenoxypolyethoxy ethanol family, Br1j 35, etc. polyoxyethylene Rubitan derivatives such as Tween 20, 3M Flu Fluoroaliphatic polymeric esters such as Orad FC-171, and Schelle A surfactant, such as Alosurf 66PK-12 from X Chemical Company. Anionic and cationic surfactants can also be used. Porous materials without wetting agents or surfactants are non-conductive. The absence of surfactant between the two chambers, of course, results in total non-conductivity. The ion selective electrode consists of an ion selective membrane 18 and a reference electrode 22 of silver/silver chloride material. Ion Selective 11118 is made of an ion selective compound (ion permeable carrier), a thermoplastic resin, and a plasticizing agent, all of which are soluble in organic solvents. The antinode is held in place within the ion selective electrode by retaining means such as retainer ring 24. As such, there is a mechanical seal between this retainer means and the lower part of the sensor body, such that there is no leakage from the membrane. Important features of the sensor The characteristics of the ion-selective membrane incorporated therein are preconditioned before the sensor can be used. No preparation (eg, immersion in a solution of the ions to be measured) is required. In the case of a hydrogen ion selective membrane, it is incorporated into the membrane. The ion-selective compound has the general formula: In this compound, R8, R, and R1 represent components of the compound that can be the same or different. R19R2'- and n, Fil) Halogen 2) an alkyl group having 4 to 18 carbon atoms, 3) a halogen-substituted alkyl group, 4) an alkoxy group, 5] halogen f! lt-substituted alkoxy group, represented by 6) -CO,R,? LR, or an acid group which is alkyl having 1-18 carbon atoms; or 7) a keto group represented by -COR, defined for R, :AZR; , a hydrogen atom. Particularly preferred derivatives of this general formula as components of hydrogen ion sensing membranes include: Mu: ζ 2-trifluoromethyl-4-octadecyloxyphenylhydrazone Sooxalonitrile b,2-octadecyloxy-5-trifluoromethylphenylhydrazone-mer Oxalonitrile C,2-octadecyloxy-5-carbethoxyphenylhydrazone-methoxy Xalonitrile cl, 2-octadecyloxy-4-fluorophenylhydrazone-mesoxa Ronitrile The ion-selective compounds are carpeethoxyphenylhydrazone mesooxalonitrile. It is particularly useful in that it shows a certain degree of inclination and does not require any pre-adjustment. That kind of membrane It is not clear whether they exhibit superior performance characteristics. However, it is against the anilino group. This is attributed to the presence of an electron-withdrawing ester group at the meta position and its proton. It may or may not happen. Its protons are clearly more acidic and therefore more easily exchanged. In addition to the ion-selective compounds described above, the ion-selective membrane 18 of the present invention also includes It consists of a plasticizer or a thermoplastic resin and, as an optional ingredient, a plasticizer. Used plastics are plastics that can be used to form films by solvent casting. FX trading is fine. For example, the plastic is polyvinyl chloride, It can be ribinyl acetate, silicone rubber or cellulose acetate. This membrane component serves the purpose of providing support and shaping into a membrane, and also serves as an ion selector. It acts as a matrix into which selective compounds are incorporated. Furthermore, since it is a hydrophobic substance, it acts as a barrier against water. In one embodiment, used in the membrane The thermoplastic resin used is polyvinyl chloride. Other things you can use are Contains lulose acetate, polyvinyl acetate, and silicone rubber. The plasticizer is an optional component of the membrane, but it is included in the membrane formulation. Any non-volatile substance suitable for the general purpose of facilitating and improving membrane flexibility may be used. stomach. Plasticizers also contribute to the dissolution of ionophores. It can for example be one or more of the following used alone or in combination: phthalates, azides. pates, sebacates, aliphatic and aromatic ethers, aliphatic and aromatic phosphates esters, aliphatic and aromatic esters, and nitrated fL7 to aliphatic and nitrated aromatic esters. In one embodiment of the invention, the plasticizer is 2-nitrophenyl octyl ether. The components of ion selective membrane 18 can be present in various amounts. The plastic used comprises about 10-30 inches by weight of the membrane, and in one embodiment is about 28% by weight. The ion-selective compound can be from about 1% to about 10% by weight of the membrane, and typically from about 3% to about 6% by weight. The plasticizer can range from about 50% to about 80% by weight of the membrane and typically comprises about 70% by weight. The ion-specific membrane 18 incorporated into the ion-selective electrode of the present invention is generally The thickness is preferably from about 3 mils to about 15 mils. The ion-specific membranes of the present invention can be fabricated by at least two different methods: solvent casting and dip coating. Steps include: 1. a) alkylation of a nitrated phenolic material to produce an ether, b) reduction of the nitro group to an amino group, C) precipitation of the hydrochloride of this amine, and and d) conversion of the hydrochloride to a menoxalonitrile derivative. Production of isomeric compounds; 2. Ion-specific compounds, plasticizers and plastics in volatile (organic) solvents; 3. Removal of volatile organic solvents (e.g. by evaporation). After the membrane has been formed, it is shaped (e.g., by cutting, using a punch-type tool) into the appropriate form.1. On Research, Cambridge, Mass., catalog 4950015). Figure 5 shows the present invention held in the electrode by an O-ring 63. An Orion electrode body 61 with a bright ion-selective membrane 62 is shown. 200-ri (not shown) prevents sample leakage around the membrane. Ion-selective membranes can also be formed by dip coating or continuous coating methods. Wear. In this method, a base material such as nylon mesh or polyester mesh (e.g. Pecap 355. Place the substance in a solvent bath (e.g., a ketone such as 4-methyl-2-be/thanone) to remove any air trapped within the mesh. This can be done ultrasonically, for example using a Branson ultrasonic cleaner. The processing time is approximately 10-15 seconds. While still wet with solvent, place the mesh into the membrane mixer j1. After taking it out, scrape off the excess formulation. The mesh containing the formulation is then dried, which can be done, for example, at room temperature or in warm air (eg 35-50°C). The internal reference material consists of a solution of the salt of the ion to be measured. Also 1.

[If the thermal electrode is silver/silver chloride, the counterion consists of a chloride tube. Agar or a gelling agent such as glucose can be used to fix the reference material in place. Ru. In the case of a pH sensor, the reference substance is a buffer solution. One embodiment scent The pH reference material consisted of 50 tnl of glycerin, 50 d of an aqueous solution consisting of phosphate buffered acupuncture water at pH 7.4 (Sigma Chemical Co., St. Louis, MO), and 2.0 t of agar. The mixture is heated to melt the agar and then placed into the cavity at the rear of the membrane. After some time, usually 5-10 minutes, the mixture gels. For the potassium sensor, the aqueous solution is 50 t/0.2 M K CJ, and for the IJium and chloride sensors, the aqueous bath t is 0.2 M NaQj. #-j to modify the sensor of Figures 1-3 to measure other ions. It is necessary to change at least one membrane component and modify the internal reference material to contain the ion to be measured. For example, to measure potassium, the membrane is ionophoric carriers specific for potassium, such as crown ether derivatives of benzo-15-crown-5 and some other neutral carriers. internal references The gel consists of a potassium chloride solution with or without a gelling agent. To modify the Figure 1-3 sensor for the measurement of sodium, the membrane can be modified with monensin, a crown ether derivative of benzo-12-crown-5, or some other sodium Neutral carrier [e.g., Analytical Cham of Simon et al., 51 :351-353(3979)n. Contains ionophoric carriers for sodium such as. Internal reference substance is sodium chloride Contains lium. To modify the sensor of Figures 1-3 to measure chloride, the membrane is coated with a quaternary ammonium salt. Contains ionophoric carriers for chloride such as Riquat 336. internal references The quality contains chloride ions. As previously mentioned, the sensor 30 has a handle 5'ft with a magnetic stripe 8. This stripe is similar to stripes commonly found on credit cards. They are very similar and hold the data recorded on the stripes during manufacturing. The data includes, for example, the following information: 1. Exam type: unique 2. Each Arabic numeral corresponds to a particular type of exam card (i.e. each (item) fully certified. This eliminates the need for operators to register test type information. 2. Calibration constant: A refined instrument method that calculates the analyte concentration based on the measured signal. The three Arabic constants used in the formula stored in - This constant may be used in conjunction with simultaneous calibration to detect possible failures of test cards. 3. Expiration 2. Instruct the meter to indicate the expiration date of the test card and ensure that the card is not used outside of the expiry period. The format of this constant is 1jYYWW, where YYti is the two Arabic digits after the week of the year (e.g. 85 in 1985) and WW is the week of the year (e.g. May The week from May 5th to May 11th is week 19). 4. Reserved: The four Arabic numerals are reserved for future use. In addition to the above data, the following 1. Administrative data can be written on the card: 1. Reader: A minimum of 15 l1101° for the magnetic stripe reading circuit. 2. Start Sentinel: A special signal used to indicate the start of data. characters. 3. LRCt: Horizontal redundancy check used in conjunction with the parity bit attached to each data character to detect errors when reading data from the magnetic stripe. Investigation. The magnetic stripe's independent functionality makes it a disposable tape designed for one-time use. This is to prevent the reuse of stock cards. This fl does not destroy the encoded information when the test card is removed from the instrument. This is achieved by The error checking function described above will attempt to reuse the card. reject the card if Figure 6fl shows a sensor of the invention that can be used to measure the activity or a degree of a sample component by differential measurement techniques. Components of biological fluids (e.g. blood, serum, plasma, urea, saliva, cerebrospinal fluid) that can be measured include, for example, glucose, urea, triglycerides, creatinine, lipase, uric acid, and other enzymes (e.g. For example, alanine aminotransferase (8GPτ spanning AL); alkaline Sexual phosphatase (MULP): creatinine kinase aspartate aminotra Contains spherase (8GO? or A, ST). Sensor-70F! It consists of an ion selective electrode, which is held within a frame or body 40 having an upper portion 42 and a lower portion 44. The upper portion 42 of the frame 40 has four openings. It has three grooves 36 located between the mouth 34 and the opening. The lower portion 44 of the frame 40 has four apertures 35 and three l1llft located between the apertures. The upper portion 42 and the lower portion 44 are in such a relationship that the openings 34 and 35 are aligned and the grooves 36 of the upper portion 42 and the grooves of the lower portion 44 are aligned. As a result, aperture 34 and aperture 35 form aperture 39, and groove 36 in upper portion 42 and groove in lower portion 44 form aperture 39. 903 spaces are defined. Upper portion 42#: a small portion located between the openings 34 Hole 43 with hole 43 allows air to enter space 41 . The ion selective electrode is placed in the opening 39 and is made of a porous material having a cylindrical or rod shape. are connected. Provides a means for ionic flow between the electrodes when applying the sample in a porous 46ij electrode. This cylindrical or rod-shaped porous material 46 is placed in the space 41 between the openings 390. Regarding the sensor of Figures 1-3, all ion-selective electrodes, less than the four depicted in Figure 6, or more than four, can be used. In addition, as mentioned above, the porous material between the electrodes can be hydrophilic or hydrophobic. class determines this property. For example, if an enzyme substrate is to be measured using the sensor shown in Figure 6 and a reference sample is to be used, the distribution ffi&: is as shown in Figure 46. It can be something like that. These ion-selective electrodes have enzymes or substrates immobilized thereon.T-layer 48: ion-selective membrane 50: membrane 52 placed between them: internal reference *S4; and reference electrode 56. Consists of. An additional membrane 58 may be present near layer 48. The ion-selective membrane can be selective for H+ or ammonium ions (NH: ), as in the case of urea electrodes. In the case of ammonium-selective membranes, the ionophore is an ammonium-selective membrane such as nonactina. It is a selective substance. The ion selective membrane is held in place within the ion selective electrode by retainer means, such as retainer ring 60. R148 has at least one enzyme or substrate immobilized thereon. Layer 48 is, for example, a nitrocellulose membrane (e.g., type AK100, Schleicher & Schnill, Keene) having a thickness of about 180 microns. , NJ). This layer can also be made of nylon, cellulose, cellulose acetate, It may be made of lath fibers, or any porous material that can serve as a solid support for the immobilization of enzymes or substrates. Enzyme fI:, substrates can be immobilized by physical attachment (eg, adsorption onto a solid phase) or chemical attachment (eg, covalent cross-linking of yci'). Layer 48 is separated from ion-selective membrane 50 by a membrane 52, which is typically thin (e.g., less than 20 microns thick) and, for example, a dialysis membrane (e.g., Spectrum membrane). Tora Ball 2, Spectrum Medical Industries, Inc., Los Angeles It is possible to be a crab. Membrane 52 serves the purpose of preserving the activity of the immobilized enzyme in layer 48 by, for example, preventing components of ion-selective membrane 50 from migrating into the layer, where migration may denature or inactivate the enzyme. It causes Membrane 58Fi is optional and its presence depends on which component of the sample is to be measured. S1 is determined by L7 and/or how that component is measured. The function of the ribs 58 is to help physically retain (immobilize) the enzyme or substrate in layer 48, and the layer 58 is to serve as a diffusion barrier to the substrate. Ru. For example, to measure the enzyme substrate once immobilized in layer 48, the endpoint measurement It may be desirable to perform a dynamic traversal instead of a constant. To extend the range of M properties for dynamic measurements, it is necessary that the enzymatic reaction be diffusion limited. In this case, membrane 58 serves both purposes. However, in Saiai, where endpoint measurement is desirable, the diffusion barrier must be small. Film 58Fi is not necessary. Regenerated cellulose (dialysis membrane) or other ultrafiltration or reverse permeation membrane material that can serve as a diffusion barrier to the membrane 58tj substrate It can be made with quality. Internal reference substance 541-j consists of a buffer solution. In one embodiment, the rt is gQI! at pH 7.4! i! It consists of a filling solution of salt-buffered acupuncture water. In a preferred embodiment, it contains about 50M of pH 7.4 phosphate buffered acupuncture water, about 50M of glycerin, and about 2.0M of phosphate buffered acupuncture water. O? It is a gel made of agarose. The reference electrode 56 is fixed and has nine potentials! It is polar and exhibits no fluctuation in liquid junction potential when the sample solution is changed or replaced by a calibration solution. The main requirements for satisfactory reference electrode performance are reversibility (in the electrochemical sense), reproducibility and stability. The three 凰 reference electrode systems are currently in use. It is used. Metal ama such as saturated calomel electrode (SGC), Yagam: Kinhi Redox couples such as doro/or ferroferrocyanide electrodes; silver-silver chlorideff,! metal/halogenated metal electrodes such as poles; Smell of the present invention In this case, the reference electrode is typically a silver/silver chloride button. The subject matter of the invention is illustrated by the following examples, which are illustrated by l! Ill@ means It should not be thought of as something to taste. The hydrogen ion specific compound 1-2-octadecyloxy-5-carboethoxyphenyl hydrazone menxalonitrile is prepared by the following method. First, organoleptics Organie8ynthegis, Goil, vol, 3, pp140-! 4] according to the general method described in Prepare C-3-nitrobenzoate. The following mixture is refluxed for 72 hours in a 11 J tube 3-necked flask equipped with a mechanical stirrer: 0.12 moi of ethyl-4-hydroxy-3-nitrobenzoate Lancaster 5 synthesis . Windham (winaham), N) 1 catalog +645141? (0.12 mol) of 96-thi-1-bromooctadecane Aldrich Chemical Co., Milwaukee. WX, catalog≠19,949-4 16.4 F anhydrous potassium carbonate 500 d anhydrous acetone After the reflux time, the reaction mixture is cooled to room temperature (i.e., about 20-27° C.) and filtered to remove salts. The p-liquid is evaporated using a Buchi-Brinkman rotary evaporator. The residue was dissolved in 500 ml of toluene and washed successively four times with 400 ml of 5% aqueous hydrogen carbonate solution and once with 400 ml of saturated aqueous sodium chloride solution. The toluene layer is dried over anhydrous sodium sulfate and evaporated to dryness. This allows the oil to crystallize if left unattended. The solid was recrystallized with hexane and decolorized with carbon. let The yield is ethyl J%/-4-octadecyloxy-3-nitrobenzoate (m, p, 48 56°) 36? (yield 66s). The 2)o compound is reduced to an amino compound by catalytic hydrogenation. Above; Toro The compound is added to 200 it/hexano with 15 g of 10-thipalladium O supported on 10 Pft carbon. The mixture is placed in a parr hydrogenator and heated to 40° C. under 6 pslg hydrogen pressure and shaken in the same for 18 hours. reaction mixture Filter the mixture while it is hot (for example, at about 50°C) through a filtration auxiliary medium [usually diatom ± (ManvilX!)]. The solution is treated with hydrogen chloride gas to precipitate the amine hydrochloride in near quantitative yield (m, p, 141-145°). Ethyl-4-octadecyloxy-3-aminobenzoate hydrochloride was prepared by a modification of the method of Brown et al. (U.S. Pat. No. 3,743,588). The above hydrochloride salt (8.5 P CO 0,018 mol) is dissolved in 12 dimethylformamide (DMF) & C while warm. Bring the temperature to approx. 31LJ in this cold solution? Add # of hydrochloric acid, then dissolve in 500 id of DMF and add 7'c1°3P of sodium nitrite. The reaction mixture is magnetically stirred for 1 hour. Then add 1.15 m of pre-warmed malononitrile and The mixture is magnetically stirred for about 10 minutes. Add enough triethylamine to obtain a strongly basic solution (e.g., pH greater than 9). Warm the mixture to room temperature and stir for 18 hours. Acidify the reaction mixture (to pH approximately 2.3) with good hydrochloric acid. A precipitate is formed. The precipitate is filtered, washed with water, and recrystallized with methanol. is about 7.1' (76 inches) of 2-octadecyloxy-5-carboethoxyphene. Nylhydrazone menxalonitrile (+m, p, 76-77°). Other derivatives shown in Table 1 were prepared in a similar manner. Example 2 - Solution casting of PVC membrane and incorporation into ion-selective electrodes 1.51 2-2 Trophenyl octyl ether (Fluka Chemical Co., Hauppa) Hauppauge, NY. Catalog ÷737323 and 0.10 S# hydrogen ionophore (as prepared by the method described in Example 1vc) is dissolved in 10 d of tetrahydrofuran. 0.6 in this solution? Powdered polyvinyl chloride having a very high molecular weight of 1.385 fP/cc and a density of 1.385 fP/cc [AzariOh Chemical Company, Milwaukee, 11. Force 20g + 18,261-: a)? Add-t-le. Shake the mixture (eg, swirl) until the P v025 is dissolved. The PVC bath liquid was poured onto a stress-relieved Boria 0 pyrene plate with a thickness of 4, and Conditions are maintained that allow evaporation of Dorofuran. For example, hume is removed as tetrahydrofuran evaporates. Evaporation can occur at room temperature under a hood. The resulting product is a plastic membrane containing ionic and qualitative compounds. Cut out the disk from the membrane using a +7 cork borer (inner diameter 0.51 nm) and insert the orion into the polar body. 0rion Research Inc., Cambridge. Installed on MA, catalog +9500153. The internal reference electrode is a silver/silver chloride reference electrode. Pole and internal filling solution FipH 7.4 phosphate buffered saline [Sigma (8XGM A) Chemical Co., St. Louis. MO, Catalogana 1000-3). The pH-sensing performance of the electrode was evaluated. All a+ constants are 2501 beakers A double contact Ag/Ag0-1 reference electrode [Konin (Corning, catalog +476067)]. For comparison, a combined pH/reference electrode was immersed in the test solution and the KpH change was recorded as the change in V in the membrane was related to the change in pH measured with the glass electrode. This is evidenced by the fact that the membrane of the invention did not tear or break when stretched. As shown, it showed good mechanical strength. It also showed good analytical performance, obtaining slope values of 10 units and 55-85 mv over the pH range of 6 to 8. However, performance deteriorated slightly at both ends of this range. For monovalent ions, the theoretical slope is ±59.1 m at 25°C. Furthermore, the response time, or time to reach equilibrium, is less than 1 minute and the electrodes do not require any test preparation and are easy to use. The properties of the above membrane can be modified by adding the hydrogen ion selective component to the previously mentioned 3-chloro-2-ol Same person except tadecyloxyphenylhydrazonemethoxalonitrile Compared with the properties of fusuma with +Xl+ according to the method. Compared to the previously mentioned membranes, membranes prepared from chlorinated menxalonitrile showed inferior performance. In other words, the slope value is within the same pH range. The sensitivity was less than 40 mV over the entire range, indicating a decrease in sensitivity. The performance of various levels of ionophores is summarized in Table 1. relative to the ionophore position. The three derivatives with lipophilic 18 carbons are derivatives in which the lipophilic chains are discrete. This result shows that the performance is superior to that of conductors. What can you infer from this? What is it? The lipophilic chain adjacent to the hydrogen ion exchange site probably facilitates the response to hydrogen ions. The lipophilic chain forms a lipophilic nine region around the active position. 7, which may increase the selectivity for hydrogen ions. Example 3 - Preparation of enzyme electrode for urea determination A sensor as shown in Figure 6 is used for urea measurement. A urea-selective sensor uses another FM containing the enzyme urease as a urea-selective sensor. are created by incorporating them into an ion-selective electrode. Therefore, pH membrane or ammonia It can be a nium ion (NH:) membrane. Electrode a shown in the fourth fl d on the 1/ase-containing membrane. b, and cIC are present. The two membranes are separated by a third membrane (eg, membrane 52 in FIG. 6). Add the test sample (eg, white liquor, plasma), water and calibration sample to the electrode as shown in Figure 4e. A and B are cores made of porous material that can be attached to conductive poles by adding a wetting agent. It is. a, b, and cFi ammonium ion selective electrodes. Two known levels of urea are added to the patient's excipient C. C also has urease. Reactions catalyzed by urease take place in b and c. Hello The patient's sample can be compared with the two calibration samples, and the test card calibration can therefore be considered. In the presence of urease, the following reaction occurs: NH,C0NH+HOe2NH 5+00°NH,-)-H,O□ N! (4-)-OH-co,-4-a,o-ma=publicHCO,-+H"As a result, ammonium-selective molecules can be used to detect changes in ammonium ion (NH4-) concentration. Gamata FipH selection A selective membrane can be used to detect the increase in pH caused by the formation of OH-. The urease-containing membrane is prepared as follows: 5 ay/d of urease (300 y/d) Nitz/9 New England Enxyme Center, Boston. MA) solution was added to a phosphate buffered saline solution (Sigma) at pH 7.4. Other buffers found to be effective: 10 mol/L Sodium Hepes (pH 7,4) in Mikal MNaCj and 0.11 J NaQt. 10 mmol/L NaH, PO4, 10 mmol/L) tris base (pH 7,4). 12 micron pore size nitrocellulose A 47J IJ diameter disc of Schleicher & Schuell, Inc., Keene, NH, 03421 grade Axloo:l is immersed in 700 ul of enzyme solution for 5 minutes at room temperature. Remove the membrane from the solution and scrape off excess liquid with a stir bar. Place the membrane on a hydrophobic surface (for example, a polypropylene silicone). Place the plate on top of the plate and air dry for 30 minutes at room temperature. The membrane is stored at 4°C in a sealed container. A disk is cut out from the membrane to form electrodes a, b, and. (See Figure 4r). sample solution You can put the solution (25 microliters) into C and the calibration solutions a and bK. Ru. Urease in a, b and C acts on urea in the calibration solution and sample, and ammonia Ni and carbon dioxide (GO)'fr are liberated. Measure the increase in pH or ammonia. For example, a buffer solution of pH 7,5 containing 40 mmol/L NaH, PO (40 mmol/L), and 100 maol/L Na(J) was prepared. After preparing a buffer solution containing 5 mmol/L urea, put it in A and C, and put the buffer solution without urea in B. After about 1 minute, a difference of 2511V was observed. A difference of 50 mv was observed in case of solution.Urine No difference in electromotive force was observed when there was no element. A similar behavior was observed when the apH membrane was replaced with an ammonium-selective nonactin membrane. Regarding FIG. 4e, cells a, b, and C each contain immobilized glucose oxidizers. contains membranes with enzymes and catalases. When glucose oxidase (COD) and catalase are present, the following reaction occurs. The purpose of catalase is to recirculate the oxygen consumed in the oxidation of glucose, so that glucose is measured by LK. It is to expand the 9 area. The oxidase/catalase membrane was prepared as follows: Nitrocell as described above. The loin membrane was washed with 5w/at glucose oxidizer in phosphate buffered saline at pH 7.4. The solution was soaked in 700 microliters of a solution consisting of 300 units/l of catalase (300 units/l from Boehringar Mannbsin) and 1 g/47 ml of catalase (40,000 units/l from Sigma, catalog-+C-100). The membrane was dried as in Example 3. Enzyme discs were placed in a, b, and c. It was found that the concentration of glucose in the sample is proportional to the potential difference created between b and 0. Other glucose measurement methods use hexokinase/ATP as the enzyme system. Referring to Figure 46, cells a, b, and C each contain a membrane with immobilized hexokinase, ATP, and a magnesium salt such as chloride, acetate, or sulfate. Ru. When hexokinase, ATP, and magnesium ions are present, the following reaction occurs. Gelcose + A'l'P corye; Glucose-6-phosphate + ADP + Iτ "Pekinkinase/TP membranes were prepared as follows: Nitrocellulose as previously described. 10 i/L magnesium acetate in a buffer (pH 7,8) consisting of lo% glycerol, 0.25% Triton 1. Solution consisting of 101119/d hexokinase. Immersed in Od. It was found that the degree of glucose in the sample is proportional to the potential difference created between B and C. Referring to Figure 4j, a membrane containing an enzyme (in this case lipase) is placed in cells a, b, and and 0. In the presence of lipase, the following reaction occurs: Lipase Triglyceride + H80-Taglycerol + Fatty Acid (H+) Similarly to the Sagi example, the nitrocellulose membrane was coated with 5η/d lipase buffer [Sigma Chemical Co., Ltd., The membrane was completely conditioned by immersing it in the liquid (catalog≠L4384). Trigri A decrease in pH caused by the addition of a sample containing cerides was detected at the pH 4l' pole. Example 6 Preparation of enzyme electrode for measuring uric acid As in the case of the previous example, uricase [50 units/U, Sigma Chemical Co. Catalog, 1 (U18 78 ) and catalase (1#/m40.Oo. unit/~Sigma Chemical Co., catalog 4cm100': prepare a timed electrode for uric acid by immersing a nitrocellulose membrane in a solution of , b, and 0. In the presence of uricase and catalase, the following reaction results in a decrease in p) i which is detected at the pH pole. Example 7 Preparation of an immunosensor for measuring theophylline In this case, K14.Theophylline The measurement of caffeine is also sensitive to the difference in immunochemical reactivity between theophylline and caffeine. Caffeine differs from theophylline by one methyl group. Theophylline and caffeine are coupled to the crown ether moiety, and each conjugate is incorporated into the PvC membrane. Theophylline membrane forms the bovine cell, while the cuff The main membrane forms the other half cell. The caffeine membrane serves as a reference electrode, and the 6 membranes alone respond to potassium ions. However, when six membranes are assembled into a six-flame cell and the same solution containing potassium ions is placed in each half cell, the electromotive force d produced between the two cow cells is very close to zero millivolts. theophylline and caffeine Although the two conjugates exhibit identical behavior in a gas chemical sense, there are significant differences in their immunochemical reactivity to theophylline antibodies. This difference in immunochemical reactivity and the same iontophoretic electrochemical reactivity may result in competitive binding to theophylline. This is the basis for the differential potentiometric measurement method. This method can also be used at a constant ionic strength or It also does not require a certain potassium ion activity [Rechnitz research]. As shown in the study]. The main advantage of this method lies in the fact that undiluted, undialyzed, and undialyzed serum samples can be used. The sensor arrangement can be as shown in FIG. The sensor consists of theophylline-crown ether conjugates in cells a and d and caffeine-crown ether conjugates in cells b and C, separated by porous connectors and C. Add the sample containing theophylline at 0 and aK, and Add calibration bath solutions a and bVc. Competitive binding involves a competitor to caffeine. The presence of theophylline antibodies with low cross-reactivity is required. antibody? L2 may be added to the sample and calibration solution prior to analysis on the sensor, or may be included in the sensor so that competitive binding occurs within the sensor. Competition for antibody sites is initiated between theophylline immobilized in the membrane and theophylline in the sample. -((Membrane) (Membrane) When the theophylline antibody binds to theophylline contained in membranes (a and C), a change in potential occurs, the magnitude of which is inversely proportional to the concentration of theophylline in the sample. Give an example. Therefore, as the theophylline t1 degree in the sample increases, fewer antibodies are able to bind to the membrane, and therefore the generated electromotive force becomes smaller. In a typical measurement. The change in potential difference due to antibody binding is usually on the order of several millivolts. Another advantage of using the theophylline membrane differential method versus the caffeine “reference” membrane, i.e. in combination with it, is that it is conson mode, i.e. the initial onset potential is zero millivolts. This means that the signal to be measured is on the order of a few millivolts. The lower the common mode potential, the higher the sensitivity when measuring the signal voltage, so if the background or common mode voltage is several orders of magnitude lower than the signal voltage, the This enables accurate and accurate measurements. As mentioned before, theophylline half cell and Ka The potential difference between Fein and the half cell is almost zero when treated with the same potassium ion-containing sample. However, the raw cell is SOB or c1 silver-silver chloride reference 1! ' When the electrode is substituted, the difference in 1i with the same potassium ion active solution is 50 V or more. The upper part increases. The method for synthesizing the conjugate and the method for preparing the membrane will be described below. Theophylline benzo-15-crown conjugate reaction preparation mechanism l-Methyl-7-((pivaloyloxy)methyl]xanthine 4'-amino-benzo-15-crown-(11-bromoundecanoamide) Theophylline-benzo-crown-5 The first step of conjugate synthesis is 1″11-methyl group. Including blocking Sanchin's 7th place. The method of Hu, Sink and Ullman [J, Am, Chew. Soc, 45:17iX-1713 (1980)). 11 conical frass 3.2 t (18.9 m+aol) of 98-thi-l-methylxanthine (Aldrich Chemical Co.) in 700 d of anhydrous DMF in a tank. Catalog 428,098-4] was added and the mixture was heated with magnetic stirring until dissolution occurred, then cooled to room temperature. While stirring this solution, add 2.0 ? (N 8.9 rnIaol, ) of anhydrous sodium carbonate was added, followed by 97 cyclo of Lomethylbiparate (Aldrich Chemical Co., catalog fr14.118-6) was added dropwise over 1 hour [also. The reaction mixture was left at room temperature overnight. Stir for a while. The reaction mixture was evaporated to dryness leaving a residue of 5011! 1 of methylene chloride. Wash the insoluble substances separately with 1sxxcl and add 1.24S' 1-methane Thirxanthin was obtained. The methylene chloride solution was evaporated to dryness to remove the -protected and diprotected groups. Get a mixture of cantin. -protected 1-methyl-7-[(pivaloyloxy)methy Rufuxanthin was separated from the diprotected xanthine by crystallization from ethyl acetate. 1.5 by this method? A monoprotected xanthine (In, 9.204-206°) was obtained. - For the ethyl acetate p solution from which the protected xanthine was separated. Contains diprotected xanthine, and after evaporation to dryness, 20d of 2N hydroxide) Both were refluxed for 18 hours. Cool the reaction mixture. The mixture was made acidic with concentrated hydrochloric acid and extracted with chloroform. The aqueous layer was evaporated to yield 0.5 g of 1-methylxanthine. The second step of the synthesis is the preparation of 4-amino-benzo-15-crown-5 hydrochloride. 1:1 acetic acid/chloroform 8001/%3 (1 (6,11 mol)) of benzo-15-crown-5 [PILrish Chemical Co., Ltd. A solution of 15*/concentrated nitric acid dissolved in 50 id of glacial acetic acid was added dropwise to the stirred solution over a period of 1 hour. The reaction mixture was stirred for 1 hour and then evaporated to dryness on a rotary evaporator. The residue was treated with 500 ic/ml of 5% aqueous sodium bicarbonate solution and the mixture was partitioned with 500 ic/ml of chloroform. Saturate the chloroform layer with sodium cyclization solution It was washed with liquid and dried with anhydrous sodium sulfate. Evaporate chloroform and take 30? (E16) No. 4 nitrobenzo-15-crown-5? Obtained. The product was recrystallized from ethanol and its 15fi' (48 mmol)? It was dissolved in dioxane at 250 °C and catalytically reduced over Raney nickel at 50 °C under hydrogenation. The reaction mixture was filtered, the PR was evaporated to dryness and the residue was dissolved in 200 d of ethyl acetate i9. I understand. The solution was then treated with hydrogen chloride gas to give 41-amino-benzo-15-chloride. The hydrochloride of Laun-5 was precipitated. Yield 9.05' (58 inches). The third step in the synthesis is the preparation of 4hibenzo-15-crown-5-(11-promoundecanoamide). Dissolve fcl 0? in 100i & aH,cl, in a 250d conical flask. (37,71101) 11-bromounde Camoic acid [Al-zich Chemical Co., Catalog +83,28 0.4:1 to 5.5 altD] ethyl aquine was added. The mixture was cooled to 0°C and 9.6751' of N,N-bis[2-oxo-3-oxazolidinyl] phosphodiamidic chloride [BOPCL Chemical Dynamics, catalog 4iz-1370-00] was added. Ta. The mixture was stirred for 0.5 h and then 12.06 PCs (37.7 mmol) of 4μ amino-benzo-15-chloride was added. Unhydrochloride'fr was added together with 5.5 m of triethylamine. This reaction mixture Another 50m1 of cti, cl! 5,5d triethylamine dissolved in water was added dropwise over 1 hour. The reaction mixture was kept warm in the room for 18 hours. Next, add water (50sd) and mix The material was made acidic with concentrated hydrochloric acid. CH, 01, layers sequentially. The tank was flushed 3 times with 200 m/ml of saturated sodium chloride solution and 3 times with 200 ml/l of 5% sodium bicarbonate solution. OH,Cj1 layer was dried with anhydrous sodium sulfate and then evaporated. It was dried and solidified. The residue was crystallized with hexane/ethyl acetate to give 8.814 t%) of 4μ benzo-]5-crown-5-(11-bromoundecanoamide) (g+, p. 101-103°). . Analysis results: , C,, H4°BrN0. Calculated value: C: 56.60; Fl near, 60; Br: 15.06; N: 2.62° Actual value: c: 56.44; H near, 46; Br: 14.89: N: 2 .60° 4th stage In the next step, a crown ether was coupled to a l-methylxanthine. To 50d DMF in a 125d conical flask was added 1.0 PC3,57t*aol)no1-mefk-7-[(pivaloyloxy)methylfuxanthine. The mixture was warmed (e.g., about 80° K. with stirring) until dissolved. Cool the solution to room temperature and add 0.76 P C7, 1 ml 1 lal) of anhydrous sodium carbonate. Then 1.90 t (3.57 mmol) of crown ether amide was added. The reaction mixture was stirred under nitrogen for 3 days. The DMF was removed using a rotary evaporator, the mixture was added to 100 d (7) of water, and the mixture was extracted three times with 50 d of chloroform. The combined chloroform extracts were washed once with 100 ml of saturated aqueous sodium chloride solution and dried over +7 ml of anhydrous sulfuric acid. Chrollo Evaporation of the form gave a pale brown oil. The fifth step is to remove the protecting groups on an aqueous basis. To the above oil in a 250 ml round bottom flask was added 75 d of 0.2M NaOH and 25 ml of methyl alcohol. The mixture was refluxed for 1 hour and cooled to room temperature. melt The liquid was acidified with concentrated hydrochloric acid while cooling in an ice/acetone bath. Ethyl alcohol precipitate Recrystallization from coal-water gave a yellowish-white crystalline solid (m, p, 166-170°) of 1.2PC5Fl). High performance liquid phase chromatography for immaterial substances It was confirmed that the material was homogeneous by analysis. Analysis results: -a,, u4. Calculated values for N, o, and: C: 60.47; H near, 37; N: 11.37. Measured values: C: 60.25; H near, 23; N: 1109° A benzo-12-crown-4 derivative was prepared in the same manner. The caffeine conjugate was prepared according to step 4 of the reaction mechanism, except that the protected 1-methylxanthine'fr1,7-dimethylxanthine was substituted. 125111 in 50 ml of DMF in a conical flask. o? (5,56 mmol) of 1,7-dimethylxanthine was added. The mixture was warmed with stirring until dissolved. Cool the solution to room temperature and add 0.595' anhydrous sodium carbonate. Then, the above-mentioned crown ether amide was added to λ95) (5,56 mmol 1.). Then, the reaction mixture was stirred for 3 days. Remove DMF from the rotary evaporator and use X0OJI! 7 ml of water was added and the mixture was extracted 3 times with 50 i*l of chloroform. Combine the extracts Wash once with 100% saturated sodium chloride aqueous solution. It was dried with anhydrous sulfuric acid (IJum). Evaporation of the chloroform gave a brown oil, which was crystallized from hexane-ethyl acetate. ．． 7 tons of 0P (29%) product was obtained, and the product was recrystallized from ethyl acetate/I/hexane and decolorized with soot carbon. Body chromatography result” “3! Calculated value of H47Nloa: C:61. 03; H near, 52; N: 11.12° Actual value C: 60.78; H near, 5 3; N: 10.97° A benzo-12-crown-4 derivative was prepared in a similar manner. Ta. Theophylline membrane components ho, oost theophylline-crown ether conjugate, 0.02i potassium tetra(p-chlorophenyl)borate, 0.5d bis(2-ethylhexyl) sebacate, 0.31 high molecular weight pvc and 5 It was made of THF of Go. The membrane can be formed by solution casting or dip-casting as previously described. Caffeine film The components of the caffeine film are: o, o syO force 7 Ein-Klau/Niehl M combination, o, 02f potassium tetra(p-chlorophenyl)borate, 0.5t/bis(2-ethylhexyl) ) Sebaket, 0.3? of high molecular weight PVC and 5 ml of THF. The membrane can be formed by solution casting or dip coating, respectively, as described above. One film of phosphorus and one film of caffeine were placed in an Orion 95 electrode body. The filling solution inside the mefc6 consisted of a phosphate buffered saline solution with a pH of 7.4 and was a diagnostic electrode Fi@/silver chloride wire. Attach the two electrodes to rubber stoppers and immerse them in 5.0 at pH 7.4 phosphate buffered saline solution. The solution was magnetically stirred. The electromotive force measured between the two electrodes was 0.23m. The ninth time the response to potassium ions was tested, 100 uJ of 0.2 mol KCj was placed into the stirring buffer using the top head. A transient response was observed in the output of the strip chart recorder as indicated by interlayered peaks (0.3 m). The potential difference is within 30 to 60 seconds. Returned to sline value. The transient response to potassium is probably due to potassium near the membrane surface. This is probably due to local concentration of the ions, which disappear as the solution is mixed. To examine the response to protein, 3.5)/61 albumin and 3.5)/61 albumin were tested. 0 Ouj standard protein solution [Sigma (51 g1aa) Chemical Co., St. Louis, MO, Cat. added to the liquid. Standard protein solution had no effect on the electrode. However, 100uJ of theophylline antibody [Immunotech, Alston, MA, catalog $651 or retail] Search Plus (Re5search Plug). When Bayonne, NJ, catalog +0l-9603-09) is added, the potential difference between the two electrodes shifts by 0.85 mv over a period of about 20 to 30 minutes. This type of sensor can be used for competitive binding measurements. The amount of antibody remaining unbound in the sample is measured and serves as an indirect measure of the antigen present in the sample. trial The more antibodies bound by the antigen in the sample, the smaller the electrical signal obtained. If this is not done, fewer antibodies will be available for binding to the antigen immobilized in the sensor membrane. In other words, the antigen 6r! t is creatinine, which is important for kidney function. It is a common indicator, and its measurement in serum is a routine blood test. Currently, There are two methods of measuring creatinine in the floor laboratory. more widely used The method used is the Jaffe reaction, which is based on the formation of a red complex of O-creatinine and vicrate in an alkaline solution. This method gives erroneous results in the presence of certain metabolites and drugs. The second method uses enzymes. Although enzymatic methods are highly effective, the cost and time required to perform the tests limit their routine use in clinical laboratories. electrode (recently, a creatinium ion-selective electrode was described [E.P.D. Diamondiss (K.P.) and Timbee Hadjiia Creatinine and Tetraphenylboronate A creatinium ion exchanger is prepared by mixing equimolar aqueous solutions of lithium and then adding a salt solution to form creatininium tetraphenylborochain salt. The salts were extracted with 2-nitrotoluene and the complex salt solution was placed as a liquid ion exchanger in an Orion 92 [Orion Research, Cambridge, MA) IIt pole equipped with a Teflon membrane. I used it. In order to achieve this type of ion selectivity, the ion exchange material is in liquid form contained in a receiver. The sample solution constantly leaches through the membrane and must be replenished periodically. The electrodes were conditioned by immersion in 0.01 mol/L creatininium chloride for 24 hours before use. The response of the creatinium electrode at pH 3 was linear with a slope of 57 at 20°C in the range of 10-3 to 0-1+aol/L. The slope decreased to H37mv in the range of 10°4 to 10-'' sol/L.The present invention utilizes a plastic membrane with the ion exchange substance immobilized thereon.2-Nitrotoluene is used as a plasticizer. When incorporating creatininium tetraphenylboron into the PVC membrane, a slope of 44 mol/L was observed between %10'' mol/L and 10-'! ``#'i decreased to 14mv in the mol/mu range. The onset of creatinine The critical range is 10-4 to 10" anal/L. With other nitrated plasticizers such as 2-nitro-p-cymene and 2-nitrophenyl octyl ether, 10-4 11 has good sensitivity in the range of 10 to 10-” mol/L. Nakatsu next. The significantly improved response in the range of 10-4 to 10-'' mol/L is expressed by the general formula: (7=Qj, Br, F, CF, ) In one embodiment, 0.025' creatininium tetra(p-chlorophenyl)boron, 1.5 2-nitrophenyl A membrane was prepared consisting of ctyl ether, 0.35' very high molecular weight PVC, and 5g THF. As mentioned in the previous section, removal is done by solution casting method or dip coating method. That's FMM Satl. ? A slope of 55 rms between $gu 10-31oool/L and 10-'' m0 1/L creatininium chloride, and a slope of 40zv between 10-4 and 10'' r+ol/L, Does not require any testing preparation It was. Substituted tetraphenylboron salts exhibit much higher sensitivity in the clinical range than unsubstituted tetraphenylboron salts [-9]. The performance of various t-substituted tetraphenylboron salts is summarized in Table 2. Unexpected effects of substituted tetraphenylboron salts compared to unsubstituted tetraphenylboron The increased sensitivity can be attributed to the increased lipophilicity and the difference in the nature of the ion exchange properties of the 7'iC@ forms formed with substituted tetraphenyl boron salts. In reality, The pH of a solution containing atinine can be lowered to below 3 pl by adding an acid. Nachinai. This can be done by diluting the sample with an acid solution. deer However, in the case of serum samples, significant dilution may have no negative effect on measurement sensitivity by decreasing the concentration of creatinine. In the case of the present invention, a different approach was taken. The pH of samples containing creatinine was lowered by treating the samples with a carrier containing anhydrous glycine hydrochloride. This can be achieved by using a membrane barrier such as nitrocellulose (Schleicher and Schull 12 micron grade Hx-Zoo). By immersing the support in a glycine hydrochloride aqueous solution (0.1 to 1.0 mol/L) So I went. The nitrocellulose membrane is made of creatininium tetra(p-riprofe). A sandwich was formed to cover the surface of the boron PvG film. Fourthly, regarding Figure 2, each cell a. b, and nitrocellulose containing creatininium PVC membrane and glycine hydrochloride Contains a base film. The serum sample is in G, the reference solution and calibration solution are in a and bK. Add each. Glycine hydrochloride converts creatinine to creatinium hydrochloride, which is detected by the membrane. The electromotive force generated between a and b can be used to calibrate the sensor. The slope from a and b and the electromotive force generated between b and C further increase the creatinine in the sample. It can be used to increase the degree of Creatinium tetraphenylboron salt is prepared by mixing equimolar aqueous solutions of creatininium hydrochloride [Sigma (81g+ia) Chemical Co., St. Louis, MO, catalog +C-6257) and the corresponding monosodium salt of substituted tetraphenylboron. did. The precipitated salt was separated, washed with water and dried, and the sodium salt of the substituted tetraphenylboron derivative was assorted. According to the method of Mclafferty and Moors (Anal, Chew, Acta 32:376-380 (1968)) Table 2: Creatinium Tetraphene Response of Nylboron Salt to Creatininium Hydrochloride Slope (mv) Slope (mv) H5010 F 45 26 (iJ 55 36 CiF, 57 40 All thesis readings seen in Table 2 are double contact silver-silver chloride The reference is to the pole. The creatininium solution used was prepared using 0.05 M glycine-glycine hydrochloride buffer (pH 3.0) containing 150 mM sodium chloride and 6-M potassium chloride. A membrane selective for Sodium rumborate, 2-nitrophenyl octyl ether of 2.0111, polymeric tPVC of 1.25', 12tl (Df) RahiYloa9:/CTHF) and 4-methyl-2-pentanone of 511tl . The membrane can be formed by solution casting or dip coating as described above. The membrane was placed in the electrode housing described above, and a test was conducted using an aqueous solution containing sodium ions. The sensor produced a slope of 57 to 60 mv per 10 units at 25°C. Methylated Monensin from Y4 250d 1,4-dioxane [20id iodine in a 500d round bottom flask Methane and 5.01 sodium monensin [Sigma (131gma) Chemica Le Co., St. Louis, MO, Catalog + M2513] was added. Mixture at 6pm It was gently refluxed for a while and then evaporated to dryness on a rotary evaporator. The residue was dissolved in 200 d of methylene chloride and washed successively twice with 500 d of a saturated aqueous sodium chloride solution, twice with 200 d of a saturated aqueous sodium bicarbonate solution, and 11 times with 200 d of a saturated aqueous sodium chloride solution. The methylene chloride solution was diluted with anhydrous sulfuric acid, dried over IJ um, and evaporated to dryness. The residue ft was treated with 50 wt/hexane while warming in a 50°C water bath. The hexane solution was cooled in a freezer for 2 hours and then filtered to remove precipitated salts. Evaporate 1 liter of hexane solution to dryness to obtain a light brown oil of 2.5-4.01 liter. The oil was used to prepare a sodium selective membrane (DIC). A membrane selective for the sodium ion was prepared using the following proportions of fractions: : 0.2p palinomycin, 0. ．． 04 P potassium tetraphenylborate 2.0 m of 2-nitrophenyl octyl ether, 1.21 m of high molecular weight PV C, 10at7 (D'rHF and 5 M! of 4-methyl-2-pentanone. The membrane was solution cast as described above. Or it can be formed by dip coating method. A selective membrane was prepared using a roux with the following ratio of components: 7'C:1-Omj Alifor (A11quat) 336 [Aldrich Chemical Co., Milwaukee, WX, Catalog ≠20, 561-3), 2. O d tris(2-butoxyethyl) phosphate (Aldrich Chemical Co., Milwaukee, WI, catalog +13,059-1), 1.21 very high molecular weight PVC (Aldrich It can be formed by a solution casting method or a dip coating method as described in Le Co., Milwaukee, Wis., Catalog 4xs, 26x-33, 1. The present invention has industrial utility in determining the ionic content or concentration of other components in biological fluids such as blood, disintegration, plasma, urine, saliva, and brain fluid. The present invention provides a method for determining not only the ionic activities of biological samples, but also glucose, urea, triglycerides, and creatine. It is particularly useful for rapid determination of concentrations of other sample components such as tinine, uric acid, lipase, other enzymes, and drugs. Ion selective membrane trial%! No preparation required, fast conclusion The results obtained make it suitable for use in clinical or research situations. Furthermore, the present invention can be used for similar vortexing in other samples such as beverages, canned meats and processed foods, fruit extracts, etc. Equivalents Those skilled in the art will recognize, or be able to ascertain, using no more than routine experimentation, many equivalents to the specific ingredients and materials specifically described herein. Such equivalents are intended to be covered within the scope of the following claims. Cleansing (no change in content) 9@, Odbeke cleansing i! F (no change in content) Kosho (no change in content) Kog! (No change in content) (1, b + (A) ocreananium odor 1 ζ pancreatic R'mat. Procedural documents are as follows: 1. Indication of the incident PCT/US86101370 2. Name of the invention Ion selective electrode 3. Person making the amendment Relationship to the case Applicant 6. Subject of amendment (1) A prescribed document stating the applicant's representative name (2) Power of attorney and translation (3) Typed print Translation of the written specification and claims (4) Translation of drawings Translation (φA-- Clause 7, Contents of amendments As per attached paper (Note: (3), (4) International search report address with no changes to the written content ANNEX To 'hεE ZNTER+NATiCNAL SE, !IRc HFtZ:'ORT ON＋＋Φ・Lee＋＋＋＋＋＋−−Rumor−−−・++彎−− 豐−＋＋＋Rumor−−＋＋＋＋−１１觧 groan・−ENTE Translation ATZONAf, APP tJCATrON No, PCT/US 86101370 (SA 137 52) Hibiki−++Addiction −・−１１＋−−Rumor−−−−+彎piece++＋demand・＋−・中 −+Fist ・−１１＋−quarrel−・Shizuku−++bone−＋＋＋−・++−Th@European Patent Offica is in no way Liabla for thesepartials which ar@merly given for the pulpo!l! Cited in 5earch ciat@member(s) darer ep mouth rt

Claims (1)

[Claims] (1) In a sensor for potentiometric measurement of ion activity in a sample, a. flame; b. Each electrode is disposed within the frame, 1. selected for the ion whose activity is to be measured. 2. membranes with selectively permeable columns containing selective ionophores; and 2. reference electrode at least two ion-selective electrodes comprising; c. When it is conductive, there is no charge to the electrode. a porous material between the electrodes that provides on-current; and d. Internal reference material with a known concentration of the ion whose activity is to be measured More sensors. (2) In a sensor for potentiometric measurement of ionic activity or other component concentration in a sample, a. Each section has an opening therethrough and a corresponding in-plane recess between the openings. the upper section and the lower section form an internal channel between the openings Align the holes in the two sections in a straight line so that the corresponding in-plane depressions It has an upper section and a lower section that correspond to the matching relationship. a frame; b. Each electrode is 1. Ionophore selective for the ion to be measured; thermoplastic resin or plastic an ion-selective membrane consisting of a stick; and a plasticizer; 2. reference electrode an ion-selective electrode within the aperture; c. An ion-selective membrane is fixed inside the opening. holding means for holding; d. between ion-selective electrodes within the aperture when electrically conductive Give an ion flow to. porous materials in the spaces between the openings in the sensor frame; Beauty e. Internal reference material with a known concentration of the ion whose activity is to be measured More sensors. (3) Each ion-selective electrode provides ionization that requires no test preparation before using the sensor. 3. The sensor according to claim 2, comprising a selective membrane. (4) the ion-selective membrane has a thickness of about 1 mil to about 15 mils, and about 1 to about 10 mils; % by weight ionophore; about 10 to about 30% by weight thermoplastic or plastic and about 50 to about 80% by weight of a plasticizer. Sir. (5) The ion-selective membrane further includes a mesh-like substance as set forth in claim 2. On-board sensor. (6) The mesh material is a nylon mesh material or a polyester mesh material. The ion-selective membrane of the sensor (7) according to claim 5, which is a substance like , an enzyme that catalyzes a reaction that causes a dilution of the pH of the sample, or Includes a substance that is a substrate for a reaction that causes a change in pH, and the enzyme or substance is The sensor according to claim 2, which is fixed to a membrane. (8) The activity of an enzyme in a sample that catalyzes a reaction that results in a change in the pH of the sample. the amount or concentration in a sample of a component that is a substrate for a reaction that results in a change in the pH of the sample In a sensor for measuring phase difference, a. Each section has an opening through it, and and a corresponding in-plane recess between the openings; Align the holes in the two sections to form an internal channel within the opening. The upper part is arranged in a line and corresponds to the relationship of matching the depressions in the corresponding plane. a frame having a section and a lower section; b. Each electrode is 1. at least one enzyme as a catalyst for a reaction that results in a change in pH, or a layer having immobilized thereon at least one substrate for a reaction resulting in a change in pH; 2. Ionophore selective for hydrogen ions or ammonium ions; thermoplastic an ion-selective membrane containing a plastic resin-lined plastic; and a plasticizer; and 3. reference electrode an ion-selective electrode within the aperture; c. An ion-selective membrane is fixed inside the opening. holding means for holding; d. When conductive between ion-selective electrodes within the aperture a porous material in the space between the openings in the sensor frame that provides ion flow to the Beauty e. Internal reference with known hydrogen ion concentration or known ammonium ion concentration material More sensors. (9) Each ion-selective electrode is an ion-selective electrode that requires no test preparation before using the sensor. 9. The sensor according to claim 8, comprising a selective membrane. (10) The enzymes immobilized on the layer are urease, glucose oxidase, katara The substrate fixed on the layer is selected from the group consisting of L-ase, uricase, and lipase; - from alanine, L-aspartate, starch, creatinine, and creatine. and the ion-selective membrane has a thickness of about 1 mil to about 15 mils; , about 1 to about 10% by weight ionophore, about 10 to about 20% by weight thermoplastic resin. or Blastec, and about 50 to about 80% by weight of a plasticizer. The sensor according to item 8. (11) According to claim 8, the ion-selective membrane further includes a mesh-like substance. Sensors listed. (12) The mesh material is a nylon mesh material or a polyester mesh material. 12. The sensor according to claim 11, which is a tussock-like substance.