{"_buckets": {"deposit": "6eae6eb4-22f7-4a01-abe5-8a63053baef1"}, "_deposit": {"created_by": 1, "id": "250", "owners": [1], "pid": {"revision_id": 0, "type": "depid", "value": "250"}, "status": "published"}, "_oai": {"id": "oai:ir.soken.ac.jp:00000250", "sets": ["9"]}, "author_link": ["0", "0", "0"], "item_1_biblio_info_21": {"attribute_name": "書誌情報（ソート用）", "attribute_value_mlt": [{"bibliographicIssueDates": {"bibliographicIssueDate": "2008-03-19", "bibliographicIssueDateType": "Issued"}, "bibliographic_titles": [{}]}]}, "item_1_creator_2": {"attribute_name": "著者名", "attribute_type": "creator", "attribute_value_mlt": [{"creatorNames": [{"creatorName": "張, 振〓"}], "nameIdentifiers": [{"nameIdentifier": "0", "nameIdentifierScheme": "WEKO"}]}]}, "item_1_creator_3": {"attribute_name": "フリガナ", "attribute_type": "creator", "attribute_value_mlt": [{"creatorNames": [{"creatorName": "ツァン, ツェンロン"}], "nameIdentifiers": [{"nameIdentifier": "0", "nameIdentifierScheme": "WEKO"}]}]}, "item_1_date_granted_11": {"attribute_name": "学位授与年月日", "attribute_value_mlt": [{"subitem_dategranted": "2008-03-19"}]}, "item_1_degree_grantor_5": {"attribute_name": "学位授与機関", "attribute_value_mlt": [{"subitem_degreegrantor": [{"subitem_degreegrantor_name": "総合研究大学院大学"}]}]}, "item_1_degree_name_6": {"attribute_name": "学位名", "attribute_value_mlt": [{"subitem_degreename": "博士（理学）"}]}, "item_1_description_1": {"attribute_name": "ID", "attribute_value_mlt": [{"subitem_description": "2008008", "subitem_description_type": "Other"}]}, "item_1_description_12": {"attribute_name": "要旨", "attribute_value_mlt": [{"subitem_description": "　　Ion channels play key roles in many cellular processes. There are many distinct\u003cbr /\u003edysfunctions known as channelopathies caused by ion channel mutations. Therefore the\u003cbr /\u003einvestigation of ion channels is of great significance for understanding how they work\u003cbr /\u003eand for drug discovery of channelopathies. Pipette patch-clamp technique has been\u003cbr /\u003eproven to be a powerful technique for the investigation of fundamental ion channel\u003cbr /\u003ebiophysics and for drug discovery. This technique allows one to monitor the gating of\u003cbr /\u003eion channels under defined conditions and enables the coupling of functional and\u003cbr /\u003emolecular studies on ion channels at the single cell level. However, this technique has \u003cbr /\u003esome weaknesses, such as the requirements of precise micromanipulation and skillful\u003cbr /\u003eexperimenter, and electrode being an individual glass pipette, which are not suitable　for\u003cbr /\u003ethe long time measurement of the cell function and the application to high-hroughput\u003cbr /\u003escreening. Recently planar patch-clamp method has attracted great attentions. Because\u003cbr /\u003eit has some advantages compared with the pipette patch-clamp method, such as the \u003cbr /\u003eminiaturization and parallelization of the planar substrates and the availability to\u003cbr /\u003ecombine with other physical probes. Many materials have been used to make the planar \u003cbr /\u003epatch-clamp substrates, such as glass, Si, quartz and PDMS etc. It has been considered\u003cbr /\u003efor Si that the background noise current is large due to the high density of free charge\u003cbr /\u003ecarrier in the substrate [Fertig et al., Recept. Channels 9(2003)29]. However, they have\u003cbr /\u003edemonstrated that the noise current can be significantly reduced by using\u003cbr /\u003esilicon-on-insulator (SOI) wafer. There are several other advantages in using SOI wafer:\u003cbr /\u003e1) the structure of the micropore through the substrate can be precisely controlled by\u003cbr /\u003eusing the large etching rate difference between Si and SiO2 in both plasma and wet\u003cbr /\u003eetching, and 2) it is possible to produce significantly miniaturized device by　integrating\u003cbr /\u003ethe biosensor and Si electronic circuits of preamplifier into the same SOI substrate.　　\u003cbr /\u003e　　In the present study, he has fabricated the planar patch-clamp substrates using SOI\u003cbr /\u003ewafer. A device, called conventional type planar patch-clamp biosensor, was assembled\u003cbr /\u003ewith the SOI-based substrate. He used this to demonstrate that SOI is a versatile\u003cbr /\u003ematerial for planar patch-clamp substrate fabrication used in biosensor. However, the\u003cbr /\u003elifetime of the cell on the micropore is too short in the planar patch-clamp biosensor,\u003cbr /\u003ewhich is a serious problem for measuring various cell functions. To elongate the \u003cbr /\u003ecellular lifetime, the substrate in the conventional type planar patch-clamp biosensor\u003cbr /\u003ewas modified with fibronectin, an extracellular matrix protein, and cells were cultured \u003cbr /\u003eunder culture medium instead of buffer solution. By using this method, he has \u003cbr /\u003edeveloped a new generation planar patch-clamp device, called incubation type planar\u003cbr /\u003eion-channel biosensor.\u003cbr /\u003e　　In the former half of his doctoral course research, he developed several elementary\u003cbr /\u003eprocesses to fabricate the planar patch-clamp substrates using SOI wafer, which\u003cbr /\u003eproduce low access resistance and low capacitance.\u003cbr /\u003e　　Two procedures of planar patch-clamp substrate fabrication were developed. One is \u003cbr /\u003ebased on electron beam lithography (EBL) combined with reactive ion etching (RIE), \u003cbr /\u003ethe other is based on focused ion beam (FIB).\u003cbr /\u003e　　In the fabrication method with EBL combined with RIE, firstly circular patterns\u003cbr /\u003ewere made with EBL and RIE techniques. Then a SiO\u003csub\u003e2\u003c/sub\u003e layer with 1 \u0026micro;m thickness was \u003cbr /\u003egrown at 900\u0026ordm;C with thermal oxidation in which O\u003csub\u003e2\u003c/sub\u003e bubbled water vapor at 95\u0026ordm;C was \u003cbr /\u003eused as the reactive gas. After that large holes on the backside of the substrate were\u003cbr /\u003emade with 1-mm-diameter diamond drill polishing, followed by 8%(v/v) TMAH etching \u003cbr /\u003eat 90\u0026ordm;C to the buried SiO\u003csub\u003e2\u003c/sub\u003e layer. Finally the buried SiO\u003csub\u003e2\u003c/sub\u003e layer at the bottom of the\u003cbr /\u003epatterns was removed with 10% (v/v) HF solution from the topside of the substrate,\u003cbr /\u003efollowed by 1-\u0026micro;m-thick SiO\u003csub\u003e2\u003c/sub\u003e layer formation at 900\u0026ordm;C with thermal oxidation in the\u003cbr /\u003epresence of O\u003csub\u003e2\u003c/sub\u003e bubbled water vapor at 95\u0026ordm;C. The scanning electron microscope (SEM)\u003cbr /\u003eimages indicate that the initially round pore becomes faceted after the thermal\u003cbr /\u003eoxidation due to the crystallographic growth-rate dependence of single-crystal silicon, \u003cbr /\u003eand the sharp edge at the rim of the pore becomes dull after the thermal oxidation. \u003cbr /\u003eThese are possibly unsuitable for the tight contact formation between the cell \u003cbr /\u003emembrane and the substrate surface. \u003cbr /\u003e　　In the fabrication method using FIB, firstly a 0.2 \u0026micro;m thickness SiO\u003csub\u003e2\u003c/sub\u003e layer was \u003cbr /\u003eformed on the silicon surface by thermal oxidation at 900\u0026ordm;C with the water-saturated O\u003csub\u003e2\u003c/sub\u003e\u003cbr /\u003eflow. Then large holes (～1 mm diameter and ～400 \u0026micro;m depth) on the backside were made by diamond drill polishing. Subsequently the pyramid-shaped holes were formed by 8% (v/v) TMAH etching at 90\u0026ordm;C for about 40 min, which reached the buried SiO\u003csub\u003e2\u003c/sub\u003e layer. \u003cbr /\u003eFinally micropores through the Si and SiO\u003csub\u003e2\u003c/sub\u003e layers were made by FIB milling from the\u003cbr /\u003ebackside. SEM images indicate that a round pore with sharp edge can be obtained with\u003cbr /\u003ethe FIB method. Therefore the planar patch-clamp substrates made with FIB were used\u003cbr /\u003eto make the device. The substrate with 1.2 \u0026micro;m diameter pore was used to make the\u003cbr /\u003econventional type planar patch-clamp biosensor. The substrate was assembled into the\u003cbr /\u003emicrofluidic circuit. The human embryonic kidney 293 (HEK-293) cell transfected with\u003cbr /\u003etransient receptor potential vanilloid type 1 (TRPV1) was positioned on the micropore\u003cbr /\u003eand the whole-cell configuration was formed by suction. Capsaicin was added to the\u003cbr /\u003eextracellular solution as a ligand molecule, and the whole-cell current of HEK-293 cell\u003cbr /\u003eshowing desensitization unique to TRPV1 in the extracellular solution containing Ca\u003csup\u003e2+ \u003c/sup\u003e\u003cbr /\u003e[Caterina et al. Nature 389(1997)816] was measured successfully.\u003cbr /\u003e　　In the latter half of his research, to overcome the cellular short-lifetime problem, \u003cbr /\u003ethe incubation type planar ion-channel biosensor was developed. He modified the\u003cbr /\u003esubstrate of the planar patch-clamp biosensor with fibronectin, and cultured the cell \u003cbr /\u003epositioned on the micropore under the culture medium instead of the buffer solution. \u003cbr /\u003eAtomic force microscope (AFM) and cell spreading assays of the FN-coated substrates\u003cbr /\u003eindicate that they are promising biomaterials for cell adhesion and spreading. After the\u003cbr /\u003ecell attached and spread on the pore of the FN-coated substrate, the resistance of the\u003cbr /\u003eelectrolyte in the cleft between the cell membrane and the substrate surface was\u003cbr /\u003eevaluated based on a schematic model and its equivalent circuit. The obtained values of\u003cbr /\u003eseal resistance quite well agree with the experimentally measured values. Using this\u003cbr /\u003emethod, the resistance of the electrolyte in the narrow space between the cell on the \u003cbr /\u003epore and all the contacting cells was evaluated. The whole-cell configuration of the\u003cbr /\u003eHEK-293 cell spreading on the pore was obtained with nystatin perforation and the\u003cbr /\u003ewhole-cell current of TRPV1 was successfully measured. It is demonstrated that\u003cbr /\u003efibronectin modification of the substrate can make the cell live for a long time and the\u003cbr /\u003ewhole-cell current of the cell spreading on the pore can be obtained during cell culture.\u003cbr /\u003eMoreover this planar ion-channel biosensor has high potential application to\u003cbr /\u003einvestigate the various cell functions and the neuronal signal transductions, because the\u003cbr /\u003ecells can be cultured on this FN-coated planar ion-channel biosensor substrate.\u003cbr /\u003e", "subitem_description_type": "Other"}]}, "item_1_description_7": {"attribute_name": "学位記番号", "attribute_value_mlt": [{"subitem_description": "総研大甲第1115号", "subitem_description_type": "Other"}]}, "item_1_select_14": {"attribute_name": "所蔵", "attribute_value_mlt": [{"subitem_select_item": "有"}]}, "item_1_select_8": {"attribute_name": "研究科", "attribute_value_mlt": [{"subitem_select_item": "物理科学研究科"}]}, "item_1_select_9": {"attribute_name": "専攻", "attribute_value_mlt": [{"subitem_select_item": "07 構造分子科学専攻"}]}, "item_1_text_10": {"attribute_name": "学位授与年度", "attribute_value_mlt": [{"subitem_text_value": "2007"}]}, "item_creator": {"attribute_name": "著者", "attribute_type": "creator", "attribute_value_mlt": [{"creatorNames": [{"creatorName": "Zhang, Zhenlong", "creatorNameLang": "en"}], "nameIdentifiers": [{"nameIdentifier": "0", "nameIdentifierScheme": "WEKO"}]}]}, "item_files": {"attribute_name": "ファイル情報", "attribute_type": "file", "attribute_value_mlt": [{"accessrole": "open_date", "date": [{"dateType": "Available", "dateValue": "2016-02-17"}], "displaytype": "simple", "download_preview_message": "", "file_order": 0, "filename": "甲1115_要旨.pdf", "filesize": [{"value": "321.9 kB"}], "format": "application/pdf", "future_date_message": "", "is_thumbnail": false, "licensetype": "license_11", "mimetype": "application/pdf", "size": 321900.0, "url": {"label": "要旨・審査要旨", "url": "https://ir.soken.ac.jp/record/250/files/甲1115_要旨.pdf"}, "version_id": "a6ed28f1-6bf0-4170-a1a1-58b6694cc200"}]}, "item_language": {"attribute_name": "言語", "attribute_value_mlt": [{"subitem_language": "eng"}]}, "item_resource_type": {"attribute_name": "資源タイプ", "attribute_value_mlt": [{"resourcetype": "thesis", "resourceuri": "http://purl.org/coar/resource_type/c_46ec"}]}, "item_title": "Fabrication of incubation type planar ion-channel biosensor using silicon-on-insulator substrate", "item_titles": {"attribute_name": "タイトル", "attribute_value_mlt": [{"subitem_title": "Fabrication of incubation type planar ion-channel biosensor using silicon-on-insulator substrate"}, {"subitem_title": "Fabrication of incubation type planar ion-channel biosensor using silicon-on-insulator substrate", "subitem_title_language": "en"}]}, "item_type_id": "1", "owner": "1", "path": ["9"], "permalink_uri": "https://ir.soken.ac.jp/records/250", "pubdate": {"attribute_name": "公開日", "attribute_value": "2010-02-22"}, "publish_date": "2010-02-22", "publish_status": "0", "recid": "250", "relation": {}, "relation_version_is_last": true, "title": ["Fabrication of incubation type planar ion-channel biosensor using silicon-on-insulator substrate"], "weko_shared_id": 1}