1. Field of the Invention
The present invention generally relates to a electrical urea biosensor and its manufacturing method, and more particularly relates to a technology for forming a urea biosensor by using a pH sensitive film with a tin oxide used as the separative gate ion-sensitive field effect transistor (EGFET) and cooperating the use of the urea enzyme.
2. Description of the Prior Art
Accordingly, the urea concentration in the blood responses the assimilation and the dissimilation catabolism of the protein and simultaneously has a closely relation of the kidney function, the liver function, and the secretion of the adrenalin. Hence, the urea nitrogen concentration of the blood or the urine is an important health index of the human body and is also an important data of the kidney function in the clinical diagnosis.
However, the conventional quantitative analysis of the organic matter has many disadvantages in the practical use, such as the complicated operation, the long analysis time, and expensive equipments, and it cannot be used in the detection of the continuous process. Hence, in order to overcome the prior disadvantage of the prior quantitative analysis, a biosensor is developed and combined with the biochemistry technology, the electronic circuits, the materials science, and the optical theory so as to design the biosensor to conform to the requirement in each fields.
The ion-sensitive field effect transistor was presented at 1970 and rapidly developed to the microminiaturized sensor. The sensor provides with the ion-sensitive electrode function and also has the character of the field effect transistor and it is completely different from the conventional electrode. The sensor has the advantages of the microminiaturization, the easy instrumentation ability, and suitable for the automation design. Following, in 1980, Caras and Janata further disclosed the gate provided with an ion-sensitive field effect transistor immobilized the aspirin within for using as the aspirin biosensor, which was called the enzyme field effect transistor.
Currently, there are many patents proposed. For example, the U.S. Pat. No. 5,922,183 in titled of “Metal oxide matrix biosensor” disclosed a substrate provided with a thin film matrix for biomolecules belonging to a general class of materials known as hydrous metal oxides and provided an amperometric biosensor or a potentiometric biosensor to perform the sensing test by the enzymes, cofactors, antibodies, antigens and the series of the nucleic acids. The U.S. Pat. No. 5,858,186 in titled of “Urea biosensor for hemodialysis monitoring” disclosed an electrochemical sensor capable of detecting and quantifying urea in fluids resulting from hemodialysis procedures. The sensor is based upon measurement of the pH change produced in an aqueous environment by the products of the enzyme-catalyzed hydrolysis of urea. The U.S. Pat. No. 5,833,824 in titled of “Dorsal substrate guarded ISFET sensor” disclosed an Ion-sensitive Field Effect Transistor (ISFET) sensor for sensing ion activity of a solution. The U.S. Pat. No. 4,877,582 in titled of “Chemical sensor device with field effect transistor” disclosed a chemical sensor having a field-effect transistor as an electronic transducer and used for the analysis of specific constituents in a liquid, the chemical sensor comprising means which permits an externally supplied sample solution to reach a chemical receptor of said chemical sensor.
Owing to the biological technology is quiet extensive, the present invention is to provide a urea biosensor belong to the formulation of the semiconductor process technology in accordance with the urea concentration of the blood or the urine and the biosensor is to detect the pH value so as to develop a structure of a disposable sensor.
The primary object of the present invention is to provide an electrical urea biosensor and its manufacturing method. The present invention utilizes a non-isolation solid-state ion-sensitive film to use as a sensitive electrode of an ion-sensitive gate field effect transistor and also utilizes the semiconductor process technology to manufacture a disposable urea biosensor.
Another object of the present invention is to provide an electrical urea biosensor and its manufacturing method. The present invention can be mass production and provides with the advantage of the low cost and the easy package so as can reduce the cost of the prior ion-sensitive gate field effect transistor simplify the package.
A further object of the present invention is to provide an electrical urea biosensor and its manufacturing method. The present invention provides with advantages of the simple production, the low cost, easily dry storage, the adjustable sensitive area, and the easy conveyance.
In order to achieve previous objects, one of the embodiments of the present invention is to provide a structure of an electrical urea biosensor. A sensitive film is positioned on a surface of a substrate, wherein a conductive layer is formed on the surface of the substrate. The sensitive film is used as an ion-sensitive electrode. The sensitive film provides with a sensitive region and a non-sensitive region. A conductive line is extended from the conductive layer for using as an external electrical contact point. The present invention utilizes a package encapsulant covering the non-sensitive region of the sensitive film to define a sensitive window at the sensitive region and a urea enzyme is immobilized within the sensitive window of the sensitive film. Then, the present invention can utilize the urea biosensor to detect of the urea concentration of the blood sample or the urine sample.
Another embodiment of the present invention is to provide a manufacturing method of an electrical urea biosensor. The manufacturing method comprises the following steps. First, a substrate is provided and a conductive layer is formed on a surface of the substrate. Then, a sensitive film is formed on a surface of the conductive layer of the substrate for using as an ion-sensitive electrode. Wherein, the sensitive film provides with a sensitive region and a non-sensitive region. Next, a conductive line is formed and extended from the conductive layer for using as an external electrical contact point. Following, a package step is performed by utilizing a package encapsulant to cover the non-sensitive region of the sensitive film so as to define a sensitive window at the sensitive region. Last, the present invention utilizes an enzyme-immobilized technology to immobilize a urea enzyme within the sensitive window of the sensitive film. Hence, the present invention completes a urea biosensor and utilizes the urea biosensor to perform the detection of the pH value so as to measure the urea concentration.
Other advantages of the present invention will become apparent from the following description taken in conjunction with the accompanying drawings wherein are set forth, by way of illustration and example, certain embodiments of the present invention.
The foregoing aspects and many of the accompanying advantages of this invention will become more readily appreciated as the same becomes better understood by reference to the following detailed description, when taken in conjunction with the accompanying drawings, wherein:
a and
The present invention utilizes the tin dioxide as the pH ion-sensitive film of the extended ion-sensitive gate field effect transistor (EGFET) and utilizes the separation structure of tin dioxide/indium tin oxide/substrate to form the urea biosensor. All the structure is the separative gate ion-sensitive field effect transistor and the suitable range of the structure is all biosensor based on the pH value detecting.
Such as shown in the
Wherein, the embodiment of the electrical urea biosensor mentioned above is using the glass substrate as its biosensor substrate. Besides, the substrate can be also selected from the group of an isolation substrate and a non-isolation substrate. Furthermore, the isolation substrate can be selected from the group of a silicon substrate, a glass substrate, a ceramics substrate, and a polymer substrate. Hence, the present biosensor has a better variation of the substrate and can change the substrate material depending on the different practical use and process condition.
Now, in order to illustrate the manufacturing method of the present invention in accordance with the structure of the
First, referring to the
Following, such as shown in the
Referring to the
A package encapsulant is used to cover the non-sensitive region of the ion-sensitive film 16 and to define a sensitive window 22. The present invention utilizes the package encapsulant 20 to define the sensitive area of the biosensor, wherein the sensitive area is about 22 mm2. A urea enzyme 24 is immobilized in the sensitive window 22 of the ion-sensitive film 16, wherein the urea enzyme 24 is composed of urease, which is embedded by a PVA-SbQ encapsulant.
Last, such as shown in the
The detail illustration of the enzyme-immobilized technology is referenced to the following description:
First, a urease (urease, EC 3.5.1.5, 50000˜100000 units/g); a PVA-SbQ encapsulant (PVA, D.P.=1700, D.S.=88; SbQ, 1.52 mol %; N.V.=12.69 wt %; and the viscosity is about 5750 cp at 25° C.); the urea (NH2CONH2=60.06), wherein its degree of purity is 99%; and the phosphate (KH2PO4=136.09), which is the normal ACS grade and used for preparing the buffer solution, are prepared.
Following, the diluted PVA-SbQ (100 mg PVA-SbQ/100 ml-55 millimole phosphate solution of the pH value 7.0) and the urea solution (7 mg urea/100 ml-millimole phosphate solution of the pH value 7.0) are mixed at the ratio of 1:1. Next, 1 ml mixture solution is taken to drop on the sensitive window 22 and then the biosensor 10 is put under the illumination of the ultraviolet of 4 watt and 365 nm to perform the photo-polymerization with about 20 minutes. Last, after finishing the photo-polymerization, the biosensor 10 is put in a dark box of 4° C. with about 12 hours to complete the enzyme immobilized process.
Such as shown in the
Referring to the
Referring to the
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Such as shown in the
Referring to the
Hence, the present invention utilizes the ion-sensitive field effect transistor of the separation structure of the extended tin dioxide/indium tin oxide/glass substrate to form the disposable urea biosensor. The structure of the urea biosensor has a best response curve under the work environment of the phosphate solution with 5 mmol and the pH value 6.0. So as the present invention can detect the urea concentration of 0.31 mg/100 ml˜120 mg/100 ml and the sensitivity of the linear portion is 169.1 mvol/p(urea concentration).
The present invention utilizes a non-isolation solid-state ion-sensitive film to use as a sensitive electrode of an ion-sensitive gate field effect transistor by integrating the semiconductor process technology to manufacture a disposable urea biosensor so as the present invention can be mass production and provides with the advantage of the low cost and the easy package so as to reduce the cost of the prior ion-sensitive gate field effect transistor simplify the package. Furthermore, the present invention simultaneously provides with advantages of the simple production, the low cost, easily dry storage, the adjustable sensitive area, and the easy conveyance.
The forgoing description of the embodiments of the invention has been presented for purposes of illustration and description, and is not intended to be exhaustive or to limit the invention to he precise from disclosed. The description was selected to best explain the principles of the invention and practical application of these principles to enable others skilled in the art to best utilize the invention in various embodiments and modifications as are suited to the particular use contemplated. It is intended that the scope of the invention not to be limited by the specification, but be defined by the claim set forth below.