Rockenbach Alexander, Sudarsan Suresh, Berens Judith, Kosubek Michael, Lazar Jaroslav, Demling Philipp, Hanke René, Mennicken Philip, Ebert Birgitta E, Blank Lars M, Schnakenberg Uwe
Institute of Materials in Electrical Engineering 1, RWTH Aachen, 52074 Aachen, Germany.
iAMB Institute of Applied Microbiology, ABBt-Aachen Biology and Biotechnology Department, RWTH Aachen, 52074 Aachen, Germany.
Metabolites. 2019 Sep 30;9(10):211. doi: 10.3390/metabo9100211.
Exploring the dynamic behavior of cellular metabolism requires a standard laboratory method that guarantees rapid sampling and extraction of the cellular content. We propose a versatile sampling technique applicable to cells with different cell wall and cell membrane properties. The technique is based on irreversible electroporation with simultaneous quenching and extraction by using a microfluidic device. By application of electric pulses in the millisecond range, permanent lethal pores are formed in the cell membrane of and , facilitating the release of the cellular contents; here demonstrated by the measurement of glucose-6-phosphate and the activity of the enzyme glucose-6-phosphate dehydrogenase. The successful application of this device was demonstrated by pulsed electric field treatment in a flow-through configuration of the microfluidic chip in combination with sampling, inactivation, and extraction of the intracellular content in a few seconds. Minimum electric field strengths of 10 kV/cm for and 7.5 kV/cm for yeast were required for successful cell lysis. The results are discussed in the context of applications in industrial biotechnology, where metabolomics analyses are important.
探索细胞代谢的动态行为需要一种标准的实验室方法,以确保能够快速采样和提取细胞内容物。我们提出了一种通用的采样技术,适用于具有不同细胞壁和细胞膜特性的细胞。该技术基于不可逆电穿孔,同时利用微流控装置进行淬灭和提取。通过施加毫秒级的电脉冲,在大肠杆菌和酵母的细胞膜上形成永久性致命孔,促进细胞内容物的释放;这里通过测量6-磷酸葡萄糖和6-磷酸葡萄糖脱氢酶的活性来证明。通过在微流控芯片的流通配置中进行脉冲电场处理,并在几秒钟内对细胞内内容物进行采样、灭活和提取,证明了该装置的成功应用。成功裂解大肠杆菌需要的最小电场强度为10 kV/cm,酵母为7.5 kV/cm。在代谢组学分析很重要的工业生物技术应用背景下对结果进行了讨论。