Kim Byoung Chan, Park Kyeong Seo, Kim Sang Don, Gu Man Bock
National Research Laboratory on Environmental Biotechnology and Department of Environmental Science and Engineering, Kwangju Institute of Science and Technology, 1, Oryong-dong, Puk-gu, 500-712, Kwangju, South Korea.
Biosens Bioelectron. 2003 May;18(5-6):821-6. doi: 10.1016/s0956-5663(03)00027-7.
A high throughput toxicity biosensor has been designed and constructed using recombinant Escherichia coli cells, containing stress specific promoters (recA, fabA, or katG) or constitutive promoters (lac) fused to luciferase genes originating from Vibrio fisheri. These genetically engineered cells were immobilized in 96 well plates. By optimizing cell immobilization conditions and the strains' response specificity to toxic chemicals, bioluminescent outputs decreased or increased dose-dependently upon adding test chemicals. However, to date the toxicity data obtained using this biosensor have not been compared with the results of other toxicity tests. Phenolics were chosen to evaluate the correlation between the LD50 and the EC50 (GC2) or EC120 (DPD2540) of Daphnia magna and E. coli, respectively. Toxicity data obtained from constitutive strains by bioluminescent level decrements were compared with the results from D. magna as a standard. LD50 values were used as parameters of D. magna toxicity and EC50 of EC120 values were used for the immobilized biosensor. In the DPD2540 test, phenolics, membrane damaging toxic chemicals, for testing immobilized stress specific bacterial strains trigger dose-dependant bioluminescence increase within specific concentration. Although the stress specific responsiveness from the strains could not be compared with D. magna's LD50 values, these responses offer additional information, such as upon the mode of toxic action in the sample, in addition to the cellular toxicity results as indicated by the EC50. This novel high throughput toxicity biosensor can be implemented to investigate the toxicity of any other soluble materials, and can be used as a standardization tool for the evaluation of toxicity.
一种高通量毒性生物传感器已被设计并构建出来，它使用了重组大肠杆菌细胞，这些细胞含有与源自费氏弧菌的荧光素酶基因融合的应激特异性启动子（recA、fabA或katG）或组成型启动子（lac）。这些基因工程细胞被固定在96孔板中。通过优化细胞固定条件以及菌株对有毒化学物质的反应特异性，加入测试化学物质后，生物发光输出呈剂量依赖性降低或增加。然而，迄今为止，使用这种生物传感器获得的毒性数据尚未与其他毒性测试结果进行比较。选择酚类物质分别评估大型溞和大肠杆菌的半数致死剂量（LD50）与半数有效浓度（EC50，GC2）或120%有效浓度（EC120，DPD2540）之间的相关性。将通过生物发光水平降低从组成型菌株获得的毒性数据与作为标准的大型溞的结果进行比较。LD50值用作大型溞毒性的参数，而EC50或EC120值用于固定化生物传感器。在DPD2540测试中，用于测试固定化应激特异性细菌菌株的酚类物质（膜损伤有毒化学物质）在特定浓度范围内触发剂量依赖性生物发光增加。尽管菌株的应激特异性反应无法与大型溞的LD50值进行比较，但这些反应除了提供EC50所示的细胞毒性结果外，还提供了其他信息，例如样品中毒性作用的模式。这种新型高通量毒性生物传感器可用于研究任何其他可溶性材料的毒性，并可作为毒性评估的标准化工具。
