Development of On-Site Environmental Monitoring Microsystems Using Miniaturized Ultrathin Slab Gel Eletrophoresis

利用小型化超薄平板凝胶电泳开发现场环境监测微系统

• 批准号: 13124201
• 负责人: UMEMURA Tomonari
• 金额: $ 24.9万
• 依托单位: Gunma University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research on Priority Areas
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13124201/
• 关键词: Two-dimensional electrophoresis; Protein; Isoelectric focusing; Optical waveguide; Interface; Air monitoring; Diesel exhaust particular; Micro TOF-Mass spectrometer; マイクロチップ; 質量分析; インターフェイス; 環境分析; 電気泳動; プロテオーム解析; ナイロン糸ゲル; オンサイト分析; 内分泌かく乱化学

1-1. Miniaturized slab gel electrophoresis was developed and experimental conditions for the miniaturized gel were optimized. Miniaturized slab gel of 30 mm wide, 30 mm long, and 0.1 mm thick was applied to two-dimensional electrophoresis (2-DE), and feasibility of high performance separation of proteins were investigated.1-2. Multifilament string-gel (MFS-Gel) was proposed as an isoelectric focusing gel (first dimension gel of 2-DE). The MFS-Gel could provide 1 mm thin, blow concentration gel down to 2.5%, which resulted in applicability to proteins of higher molecular weight that 200 kDa.1-3. Fast 2-De using the MFS-Gel was developed. Colored proteins were used to evaluate performance of the isoelectric focusing system, and focusing field could be applied up to 430 V/cm, and the migration time could be reduced from 2 hours to 30 min. Practical samples such as E. Coli and chicken skeletal muscle extract were separated in 2 hours in the first dirnension and in 1 hour in the second dimension, which was at least three times faster than the conventional 2-DE.1-4. Optical waveguide detection system using the evanescent wave was developed for microchip analysis and interfacial substances were successfully detected.2-1. Microchip-device has been developed and optimized in order to measure for atmospheric NO_2 with high sensitivity. The detection limit was approximately 1 ppb in wind tunnel.2-2. Pre-treatment microchip was developed for eluting PAHs from diesel exhaust particle (DEP). PAHs in DEP were trapped into silica gel beads, which were packed into the microchip, and extracted into chloroform. The recovery of this pre-treatment microchip was same as bulk scale methods.2-3. MS interface has been developed. This interface has monolith structure in the capillary, and investigated the capacity to resist pressure between vacuum bath and liquid delivery system. As a result of connecting this interface and micro liquid delivery system, flow rate has reduced below 1 μL/min.

1-1.建立了微型平板凝胶电泳技术，并对微型凝胶电泳条件进行了优化。将30 mm宽、30 mm长、0.1 mm厚的微型平板凝胶应用于双向电泳（2-DE），并研究了高效分离蛋白质的可行性。提出了一种等电聚焦凝胶（二维凝胶），即多细丝串凝胶（MFS-Gel）。MFS-Gel可以提供1 mm薄的、吹塑浓度低至2.5%的凝胶，这导致适用于分子量高于200 kDa的蛋白质。开发了使用MFS-Gel的快速2-De。以有色蛋白质为例，对等电聚焦系统的性能进行了评价，聚焦场可达430 V/cm，迁移时间从2 h缩短到30 min。大肠杆菌和鸡骨骼肌提取物的分离在第一维上在2小时内，在第二维上在1小时内，这比传统的2-DE至少快3倍。开发了用于微芯片分析的使用倏逝波的光波导检测系统，并成功地检测了界面物质。为了实现高灵敏度的大气NO_2气体测量，研制了一种新型的微芯片测量装置。在风洞中的检测限约为1 ppb。2 -2.研制了用于洗脱柴油机尾气颗粒物（DEP）中多环芳烃的预处理微芯片。DEP中的多环芳烃被捕获到硅胶珠中，将其装入微芯片中，并用氯仿萃取。该预处理微芯片的回收率与批量规模方法相同。2 -3.开发了MS接口。该界面在毛细管中具有整体结构，并研究了真空浴和液体输送系统之间的耐压能力。将该接口与微量液体输送系统连接后，流量已降至1 μL/min以下。

项目成果

T.Korenaga, X.Liu, Z.Huang: "The Influence of Moisture Content on Polycyclic Aromatic Hydrocarbons Emission During Rice Straw Burning"Chemosphere-Global Change Science. 3(1). 117-122 (2001)

T.Korenaga、X.Liu、Z.Huang：“水分含量对稻草燃烧过程中多环芳烃排放的影响”化学圈-全球变化科学。

伊永隆史, 高林義昌: "ラボオンチップ環境計測素子-微細品産業の勃興をめざす-"化学工業. 52(11). 847-851 (2001)

Takashi Inaga、Yoshimasa Takabayashi：“片上环境测量装置 - 瞄准微型产品产业的崛起 -”化学工业 52(11)。

角田欣一, 梅村知也: "光ファイバー型キャピラリーセルの分析化学における可能性"ケミカル・エンジニヤリング. 10. 44-47 (2003)

Kinichi Tsunoda、Tomoya Umemura：“光纤毛细管细胞在分析化学中的可能性”《化学工程》10. 44-47 (2003)。

伊永隆史: "簡易分析法-大気分析"ぶんせき. 8. 424-429 (2002)

Takashi Inaga：“简单分析方法 - 大气分析”Bunseki 8. 424-429 (2002)。

M.Helaleh, S.Fujii, T.Korenaga: "Column Silylation Method for Determining Endocrine Disruptors from Environmental Water Samples by Solid Phase Micro-Extraction"Talanta. 54(6). 1039-1047 (2001)

M.Helaleh、S.Fujii、T.Korenaga：“通过固相微萃取测定环境水样中内分泌干扰物的柱硅烷化方法”Talanta。