Online analysis of protein inclusion bodies produced in E. coli by monitoring alterations in scattered and reflected light
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- 作者：Christian Ude ; Nadav Ben-Dov ; André Jochums…
- 关键词：Online scattered ; light sensor ; Inclusion bodies ; Flow cytometry ; Reflective interference Fourier transform spectra ; Silicon photonic arrays ; Optical biosensor
- 刊名：Applied Microbiology and Biotechnology
- 出版年：2016
- 出版时间：May 2016
- 年：2016
- 卷：100
- 期：9
- 页码：4147-4159
- 全文大小：1,437 KB
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Nadav Ben-Dov (2)
André Jochums (1)
Zhaopeng Li (1)
Ester Segal (2)
Thomas Scheper (1)
Sascha Beutel (1)
1. Institut für Technische Chemie, Gottfried Wilhelm Leibniz Universität Hannover, Callinstraße 5, 30167, Hannover, Germany
2. Department of Biotechnology and Food Engineering, Technion—Israel Institute of Technology, 32000, Haifa, Israel - 刊物类别：Chemistry and Materials Science
- 刊物主题：Chemistry
Biotechnology
Microbiology
Microbial Genetics and Genomics - 出版者：Springer Berlin / Heidelberg
- ISSN：1432-0614
文摘
The online monitoring of recombinant protein aggregate inclusion bodies during microbial cultivation is an immense challenge. Measurement of scattered and reflected light offers a versatile and non-invasive measurement technique. Therefore, we investigated two methods to detect the formation of inclusion bodies and monitor their production: (1) online 180° scattered light measurement (λ = 625 nm) using a sensor platform during cultivation in shake flask and (2) online measurement of the light reflective interference using a porous Si-based optical biosensor (SiPA). It could be shown that 180° scattered light measurement allows monitoring of alterations in the optical properties of Escherichia coli BL21 cells, associated with the formation of inclusion bodies during cultivation. A reproducible linear correlation between the inclusion body concentration of the non-fluorescent protein human leukemia inhibitory factor (hLIF) carrying a thioredoxin tag and the shift (“Δamp”) in scattered light signal intensity was observed. This was also observed for the glutathione-S-transferase-tagged green fluorescent protein (GFP-GST). Continuous online monitoring of reflective interference spectra reveals a significant increase in the bacterium refractive index during hLIF production in comparison to a non-induced reference that coincide with the formation of inclusion bodies. These online monitoring techniques could be applied for fast and cost-effective screening of different protein expression systems.