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Application of Ion-Sensitive Field Effect Transistor to Biosensor

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Application of Ion-Sensitive Field Effect Transistor to Biosensor 石 油 学 会 誌 Sekiyu Gakkaishi, 34, (5), 399-406 (1991) 399 Application of Ion-sensitive Field Effect Transistor to Biosensor Jun-ichi ANZAI, Shouryu LEE, and Tetsuo OSA* Pharmaceutical Institute, Tohoku University, Aobayama, Sendai 980 (Received January 11,1991) The principle and operation of an ion-sensitive field effect transistor (ISFET) are summarized briefly, and the application of the device to development of biosensors is described. Several kinds of techniques are discussed to immobilize ionophores and enzymes on the ISFET. for/solution interface. The surface potential at 1. Introduction the gate surface is often described reasonably, based The concept of an ion-sensitive fieldeffect tran- on the site-binding model, which explains the sistor(ISFET) was derived from the structure of the interaction between an inorganic insulator and an metal oxide semiconductor field effect transistor adjacent electrolyte solution, with the assumption (MOSFET), which is a purely electronic semicon- that the surface contains a discrete number of ductor device1),2). In Fig. 1 are schematically surface sites, which can bind or liberate H+ ion3). illustrated the similarities and dissimilarities In the case of ISFET with Si3N4 gate insulator, the between MOSFET and ISFET, together with the gate surface contains silanol groups which can electrical equivalent circuit common to both bind or liberate H+ ions, depending on the pH of devices. The variance between them is that the the measuring solution, as shown in Fig. 2. The metal gate of the MOSFET is replaced by the ISFET is often applied in a feedback circuit to reference electrode for ISFET, while the sample obtain an electrically stable operation. For this solution ensures the electrical contact with the reason, the drain-source voltage has a constant original gate insulator. The pH sensitivityof the preset value and the Id value is also constant, which ISFET is ascribed to the fact that the drain current, affords a gate-source voltage equal to the value of Id,depends on the surface potential at the insula- surface potential at the gate surface. The present paper describes the application of ISFET to the fabrication of microbiosensors. The advantages of ISFET in biosensor application are: (1) easy fabrication of miniature devices through semiconductor manufacturing process, (2) feasibility to accumulate multiple sensors on single tip, (3) applicability of non-conductive materials for sensitive layer ascribed to low output impedance, and (4) suitability for mass production. 2. ISFET Ion Sensors The standard type of pH-sensitive ISFET has been used to detect ionic species other than H+ ion by coating the gate surface with ion-sensitive membranes. A polyvinyl chloride (PVC) membrane containing valinomycin has been used Fig. 1 Schematic Representation of MOSFET (A) and ISFET (B) and the Equivalent Circuit (C) * To whom correspondence should be addressed. Fig. 2 pH Response of Si3N4 Gate Surface 石 油 学 会 誌 Sekiyu Gakkaishi, Vol. 34, No. 5, 1991 400 to prepare K+-ion sensor4). The PVC membrane- based ISEFT ion sensor shows some advantages over conventional type of membrane electrode sensors by its small size and low output impedance. The disadvantages, however, such as slow response, short life, etc. remain to be improved. The polymer materials such as polyacrylates and polysiloxanes have been used to improve the potentiometric response of ISFET ion sensors5). We have recently prepared two types of ISFET K+-ion sensors with ion-sensitive layers of mono- molecular level: chemically modified gate and LB membrane-coated gate ISFETs. The fabrication and performance characteristics of these sensors are described below. 2.1 Chemically Modified ISFET Sensors The chemical modification of electrode surface is one of the most promising techniques in devel- oping chemically functional electrodes6). There are some advantages in applying the chemical modification method to the preparation of ISFET ion sensors: owing to the thinness of the ion sensitive layer, minute devices (even in micron Fig. 3 The Proceduree for Surface Modification with order) can easily be prepared, and very rapid Crown Ether response can be expected. Additionally, the covalent bonding between ionophore and gate surface ensures a long life for the sensors without deterioration in sensitivity and selectivity. A procedure for chemical modification of ISFET is shown in Fig. 37). The probe-type Si3N4 gate ISFET coated with poly (p-xylylene) (parylene) film was used as an underlying device. The base insulator material for chemical modification should have less binding site for ions to suppress non-specific noise response. From this view- point, parylene is an excellent material8). First, the carboxylic groups were introduced at the surface of parylene by annealing. The carboxylic Fig. 4 Potentiometric Response of K+-ion Sensitive groups were treated with thionyl chloride to be ISFET Chemically Modified with Crown Ether activated to acid chloride form. The ISFET probe thus activated was immersed in toluene solution of 4'-aminobenzo-18-crown-69), at room temperature, sensor were improved as compared with those of for one day. It was ascertained that the crown PVC membrane-based sensors. The response ether residues are introduced to the parylene time was within 10 sec. After the initial decrease surface through amide linkage, by FTIR in sensitivity, the sensor maintained full response measurement: signals at 1,665cm-1 (amide I) and even after one month. There are some problems 1,569cm-1 (amide II). to be overcome for the chemically modified ISFET The potentiometric response of the K+-ion ion sensors: ca. 20mV/pH of pH response, ionic sensor is shown in Fig. 47). The sensitivity to K+ intensity response, and initial drift of potential. ion was ca. 35mV/log[K+], whereas the Since these problems stem from the chemical conventional type PVC/crown ether membrane- properties of modified parylene film, measure- coated device exhibited ca. 55mV/log[K+]. The ments in different mode with an unmodified difference in sensitivity between the two devices parylene film ISFET may solve the problems. seems to originate from the difference in the 2.2 LB Membrane-modified ISFET Sensors density of crown ether at the gate surface. On the A thin membrane with K+ ion sensitivity was other hand, the response time and the life of the deposited on the parylene gate ISFET by Lang- 石 油 学 会 誌 Sekiyu Gakkaishi, Vol. 34, No. 5, 1991 401 3. ISFET Enzyme Sensors It is envisaged that the ISFET enzyme sensors can be prepared by modification of gate surface with enzymes which catalyze the reaction con- suming or producing H+ion. Based on this idea, many kinds of ISFET enzyme sensors have been developed so far13)-17). In general, enzymes can be immobilized to the gate surface of ISFET by the use muir-Blodgett (LB) technique10). A mixed of organic thin membranes. The organic mem- monolayer composed of stearic acid and crown branes where enzyme is immobilized should be a ether 1 (9: 1 in weight) was spread with benzene on crucially important factor in determining the water subphase at ca. pH 7 in Langmuir trough. performance of the sensors, because the signal, The surface of the parylene gate ISFET was usually H+ion, is produced as a result of an enzyme covered with the Y-type mixed monolayer. The catalyzed chemical reaction of analyte on and/or in senosor showed a response to K+ ion, over the the membrane. Additionally, the enzyme activity concentration range of 0.01-100mM, with a slope and the diffusion rates of analytes and reaction of ca. 35mV/log[K+]. This sensor, however, products should depend on the chemical and suffers from a drawback in that, in spite of the low physical properties of the membrane. The pH response (5-10mV/pH) of the parylene gate usefulness of direct covalent bonding of enzyme to ISFET without LB layer, the gate potential is ISFET gate has also been discussed very recently18). sensitive to pH, presumably ascribed to the ion- Bearing these in mind, we have examined the exchange sites resulting from the carboxylic performance of ISFET enzyme sensors with two groups in the LB layer. In Table 1 are shown pH different types of organic thin membranes: cross- response of ISFET sensors with and without stearic linked albumin membrane and LB membrane. acid LB membrane11). The use of non-ionic 3.1 Albumin Membrane-modified ISFET En- amphiphiles may be useful for the removal of pH zyme Sensors response. The long-term stability of this sensor is Bovine serum albumin (BSA) has been widely not satisfactory. The improvement of perfor- used to immobilize enzyme in its cross-linked gel mance characteristics of the LB membrane- matrix, because of the inertness of BSA and modified ISFET ion sensors may be possible by the relatively good surface adhesion to solid support. use of LB membranes with higher content of crown We have prepared enzyme sensors by covering the ether, polymerized LB layers, etc. ISFET gate with the enzyme/BSA cross-linked It is possible to miniaturize body of the sensor by membranes19),20). The enzymes used were urease, the use of ISFET device described above. At the trypsin, α-chymotrypsin, penicillinase, and present stage, however, the use of ISFET sensors is glucose oxidase. In this paper, the results of premature in that no reference electrode of urease-modified sensor are described. comparable size is available. Reinhoudt et al. About 10% urease and BSA solutions were mixed have demonstrated that polyacrylate-modified reference FET shows superior pH insensitivity (within 1mV/pH), as compared with previously reported reference FETs coated with ion-blocking thin films by plasma deposition method12). Table 1 Effect of LB Membranea) Deposition on the pH Response of ISFET A: ISFET without urease/albumin membrane. B: ISFET coated with 2μm membrane.
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