Construction of biochemical analytical systems using the microchip technology

利用微芯片技术开发生化分析系统。在环境分析领域，建立了海洋腹足类类固醇激素和河流亚硝酸盐、亚硫酸盐的分析系统。生物活性化合物如细胞因子的分析系统也被开发出来。构建了小鼠巨噬细胞TNF-α免疫测定芯片和大鼠腹膜肥大细胞组胺生物测定芯片。首先，我们开发了一种简单的方法，通过光刻技术在玻璃板上进行微加工，使用高粘度光刻胶作为蚀刻掩膜，然后制造的玻璃用于聚二甲基硅氧烷（PDMS）微芯片的模板。我们还开发了一种用于微芯片分析的新型荧光检测单元。与传统的酶联免疫吸附测定法相比，该免疫测定芯片对海洋腹足类动物中类固醇激素的分析灵敏度提高了一个数量级，分析时间缩短了三分之一。在用于检测肥大细胞释放组胺的过敏药物筛选系统中，与传统的批量系统相比，检测所需的细胞数量减少到1%，并且通过细胞培养基的连续流动实现对组胺释放的实时监测。配备微流控梯度混合器的抗癌剂微生物测定系统。通过在微通道中简单地流动两种流体，可以很容易地进行同时梯度测定。该系统表明，胃癌细胞系的细胞活力随抗癌剂浓度的变化而变化。

Biochemical analytical systems using microchip technology were developed. In the field of environmental analyses, analytical systems for steroid hormones in marine gastropods and for nitrite and sulfite in river were constructed. Analytical systems for bioactive compounds such as cytokines were also developed. An immunoassay microchip for TNF-α from mouse macrophages and a bioassay microchip for histamine from rat peritoneal mast cells were constructed.First, we developed a simple method for micro-fabrication on a glass plate by photolithography using a high viscosity photoresist as an etching mask and then the fabricated glass was used for the template of a polydimethylsiloxane (PDMS) microchip. We also developed a novel fluorescence detection unit for microchip analyses.The analytical sensitivity for steroid hormones in marine gastropods, using the immunoassay microchip, was improved by one order of magnitude and the analysis time was reduced to one third, compared to the conventional ELISA method.In the allergic drug screening system for the detection of histamine released from mast cells, the number of the cells, required for the assay, was reduced to 1%, compared with conventional bulk systems and the real-time monitoring of the histamine release was achieved by the continuous flow of cell culture medium.Microbioassay system for an anti-cancer agent equipped with a microfluidic gradient mixer. The simultaneous gradient assay was conducted easily by simply flowing two fluids in microchannels. This system demonstrated that cell viability of a stomach cancer cell line changed according to the concentrations of an anti-cancer agent.