Quasi-continuous combined scattered light and fluorescence measurements:: A novel measurement technique for shaken microtiter plates
Quasi-continuous combined scattered light and fluorescence measurements:: A novel measurement technique for shaken microtiter plates
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DOI：
10.1002/bit.20573
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发表时间：
2005-10-05
影响因子：
3.8
通讯作者：
Büchs, J
中科院分区：
文献类型：
--
作者：
Samorski, M;Müller-Newen, G;Büchs, J
提出了一种新的振荡微量滴定板准连续在线测量技术。在振荡过程中测量光散射以及细胞内和/或蛋白质荧光（例如NADH、YFP），从而允许在微量滴定板中对96种不同的同时培养物进行过程监测。与现有的测量技术相比，不必停止振荡过程来进行测量，从而避免了培养物氧气供应的相应中断和对培养物的任何不可预测的影响。实验用E.在LB、TB和MOPS基本培养基中的大肠杆菌和在改良的LB和TB培养基中的产钠弧菌。散射光和NADH荧光的强度曲线被用来区分不同的滞后期，生长速度，或接种密度。从这种新方法的数据对应离线测量的光密度和氧转移率的文化运行在同时进行摇瓶实验在同等的氧转移能力。利用与白细胞介素-6融合的黄色荧光蛋白，确定了表达E.大肠杆菌菌株可以通过在线监测产物形成来确定。因此，这种测量技术使研究人员能够在筛选水平上评估和区分不同的培养物，并改善筛选条件，工艺开发和放大。(c)2005 Wiley Periodicals，Inc.
A novel quasi-continuous on-line measuring technique for shaken microtiter plates is presented. Light scattering as well as intracellular and/or protein fluorescence (e.g. NADH, YFP) is measured during the shaking procedure, thus allowing a process monitoring of 96 different simultaneous cultures in a microtiter plate. In contrast to existing measurement techniques, the shaking process does not have to be stopped to take the measurements, thus avoiding the corresponding interruption of the cultures' oxygen supply and any unpredictable effects on the cultures. Experiments were conducted with E. coli in LB, TB, and MOPS minimal medium and V. natriegens in modified LB and TB media. Intensity curves of scattered light and NADH fluorescence were used to distinguish different lag phases, growth velocities, or inoculation densities. Data from this new method corresponded well to the off-line measured optical densities and to the oxygen transfer rates of cultures run in simultaneously conducted shake flask experiments at equivalent oxygen transfer capacities. With the aid of yellow fluorescence protein fused to interleukin-6 the optimal induction time of an expressing E. coli strain could be determined by on-line monitoring of product formation. Thus, this measuring technique enables the researcher to evaluate and to discriminate different cultures on a screening level and to improve screening conditions, process development and scale-up. (c) 2005 Wiley Periodicals, Inc.