Department of Chemical and Biomolecular Engineering
Korea Advanced Institute of Science and Technology

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[COVER PAGE] Cell-Based Quantification of Homocysteine Utilizing Bioluminescent Escherichia coli Auxotrophs

Min-Ah Woo†, Moon Il Kim†, Byung Jo Yu‡, Daeyeon Cho§, Nag-Jong Kim†, June Hyoung Cho‡, Byung-Ok Choi*, Ho Nam Chang†, and Hyun Gyu Park*†
† Department of Chemical and Biomolecular Engineering (BK21 Program), KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon 305-701, Republic of Korea
‡ MD Science Inc., 258-1 Munji-dong, Yuseong-gu, Daejeon 305-701, Republic of Korea

http://pubs.acs.org/doi/abs/10.1021/ac103350y

 

Abstract
A cell-based quantitative assay system for Hcy has been developed by utilizing two Escherichia coli auxotrophs that grow in the presence of methionine (Met) and either homocysteine (Hcy) or Met, respectively. A bioluminescent reporter gene, which produces luminescence as cells grow, was inserted into the auxotrophs, so that cell growth can be readily determined. When the relative luminescence unit (RLU) values from the two auxotrophs immobilized within agarose gels arrayed on a well plate were measured, the amount of Hcy was quantitatively determined on the basis of differences between two RLU values corresponding to cell growth of two auxotrophs with excellent levels of precision and reproducibility. Finally, the diagnostic utility of this assay system was verified by its employment in reliably determining different stages of hyperhomocysteinemia in human plasma samples providing CVs of within and between assays that are less than 2.9% and 7.1%, respectively, and recovery rates of within and between assays that are in the range of 99.1−103.5% and 97.5−105.5%, respectively. In contrast to existing conventional methods, the new system developed in this effort is simple, rapid, and cost-effective. As a result, it has great potential to serve as a viable alternative for Hcy quantification in the diagnosis of hyperhomocysteinemia



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등록일2011-08-25

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