Ogura M, Agata Y, Watanabe K, McCormick R M, Hamaguchi Y, Aso Y, Mitsuhashi M
Hitachi Chemical Research Center, Irvine, CA 92612, USA.
Clin Chem. 1998 Nov;44(11):2249-55.
Two major components of rRNA (18S and 28S rRNA) were separated by electrophoresis in injection-molded acrylic chips with a microchannel 100 microm in width, 40 microm in depth, and with 1 cm of separation distance. Microchannels were filled with 4 g/L hydroxypropylmethylcellulose as sieving polymer and 5 mg/L ethidium bromide for RNA staining. The fluorescent signals were detected by a fluorescent microscope equipped with a photometer and 590 nm emission filter. The assay is rapid (<3 min), reproducible, RNase-free, and requires only 1-2 microL of sample. The detection limit was approximately 10 mg/L (10 ng/microL), 100-fold lower than that for conventional agarose gel electrophoresis. Because only 0.1 nL of the loaded sample was used for electrophoresis, the detectable peaks of rRNA in the separation were derived from less RNA than in a single cell. Because the quality of RNA is critical for RNA-related diagnostic tests, disposable plastic chips will be useful for quality assessment of RNA.
通过在宽度为100微米、深度为40微米且分离距离为1厘米的注塑丙烯酸芯片中进行电泳，分离出rRNA的两个主要成分（18S和28S rRNA）。微通道中填充有4 g/L羟丙基甲基纤维素作为筛分聚合物和5 mg/L溴化乙锭用于RNA染色。通过配备光度计和590 nm发射滤光片的荧光显微镜检测荧光信号。该检测方法快速（<3分钟）、可重复、无核糖核酸酶，且仅需1-2微升样品。检测限约为10 mg/L（10 ng/微升），比传统琼脂糖凝胶电泳低100倍。由于仅0.1纳升的加样样品用于电泳，分离中rRNA的可检测峰源自比单个细胞中更少的RNA。由于RNA质量对于与RNA相关的诊断测试至关重要，一次性塑料芯片将有助于RNA的质量评估。
