Lee Jin Hyung, Mitchell Robert J, Kim Byoung Chan, Cullen David C, Gu Man Bock
National Research Laboratory on Environmental Biotechnology, Gwangju Institute of Science and Technology (GIST), 1 Oryong-dong, Buk-gu, Gwangju 500-712, Republic of Korea.
Biosens Bioelectron. 2005 Sep 15;21(3):500-7. doi: 10.1016/j.bios.2004.12.015. Epub 2005 Jan 22.
In this study, a cell-based array technology that uses recombinant bioluminescent bacteria to detect and classify environmental toxicity has been implemented to develop two biosensor arrays, i.e., a chip and a plate array. Twenty recombinant bioluminescent bacteria, having different promoters fused with the bacterial lux genes, were immobilized within LB-agar. About 2 microl of the cell-agar mixture was deposited into the wells of either a cell chip or a 384-well plate. The bioluminescence (BL) from the cell arrays was measured with the use of highly sensitive cooled CCD camera that measured the bioluminescent signal from the immobilized cells and then quantified the pixel density using image analysis software. The responses from the cell arrays were characterized using three chemicals that cause either superoxide damage (paraquat), DNA damage (mitomycin C) or protein/membrane damage (salicylic acid). The responses were found to be dependent upon the promoter fused upstream of the lux operon within each strain. Therefore, a sample's toxicity can be analyzed and classified through the changes in the BL expression from each well. Moreover, a time of only 2 h was needed for analysis, making either of these arrays a fast, portable and economical high-throughput biosensor system for detecting environmental toxicities.
在本研究中,一种基于细胞的阵列技术得以实施,该技术利用重组生物发光细菌来检测和分类环境毒性,从而开发出两种生物传感器阵列,即芯片阵列和平板阵列。二十种不同启动子与细菌lux基因融合的重组生物发光细菌被固定在LB琼脂中。将约2微升细胞 - 琼脂混合物滴入细胞芯片或384孔板的孔中。使用高灵敏度的冷却电荷耦合器件相机测量细胞阵列发出的生物发光(BL),该相机测量固定细胞发出的生物发光信号,然后使用图像分析软件对像素密度进行量化。使用三种分别导致超氧化物损伤(百草枯)、DNA损伤(丝裂霉素C)或蛋白质/膜损伤(水杨酸)的化学物质对细胞阵列的响应进行表征。发现这些响应取决于每个菌株中lux操纵子上游融合的启动子。因此,可以通过每个孔中BL表达的变化来分析和分类样品的毒性。此外,分析仅需2小时,这使得这些阵列中的任何一种都成为用于检测环境毒性的快速、便携且经济的高通量生物传感器系统。