Applied Microbiology and Biotechnology

, Volume 100, Issue 9, pp 4147–4159 | Cite as

Online analysis of protein inclusion bodies produced in E. coli by monitoring alterations in scattered and reflected light

  • Christian Ude
  • Nadav Ben-Dov
  • André Jochums
  • Zhaopeng Li
  • Ester Segal
  • Thomas Scheper
  • Sascha Beutel
Methods and protocols

Abstract

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 Escherichiacoli 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.

Keywords

Online scattered-light sensor Inclusion bodies Flow cytometry Reflective interference Fourier transform spectra Silicon photonic arrays Optical biosensor 

Supplementary material

253_2016_7403_MOESM1_ESM.pdf (28.4 mb)
ESM 1(PDF 29087 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Christian Ude
    • 1
  • Nadav Ben-Dov
    • 2
  • André Jochums
    • 1
  • Zhaopeng Li
    • 1
  • Ester Segal
    • 2
  • Thomas Scheper
    • 1
  • Sascha Beutel
    • 1
  1. 1.Institut für Technische ChemieGottfried Wilhelm Leibniz Universität HannoverHannoverGermany
  2. 2.Department of Biotechnology and Food EngineeringTechnion—Israel Institute of TechnologyHaifaIsrael

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