Comparison of time‐gated surface‐enhanced raman spectroscopy (TG‐SERS) and classical SERS based monitoring of Escherichia coli cultivation samples

Abstract

The application of Raman spectroscopy as a monitoring technique for bioprocesses is severely limited by a large background signal originating from fluorescing compounds in the culture media. Here, we compare time-gated Raman (TG-Raman)-, continuous wave NIR-process Raman (NIR-Raman), and continuous wave micro-Raman (micro-Raman) approaches in combination with surface enhanced Raman spectroscopy (SERS) for their potential to overcome this limit. For that purpose, we monitored metabolite concentrations of Escherichia coli bioreactor cultivations in cell-free supernatant samples. We investigated concentration transients of glucose, acetate, AMP, and cAMP at alternating substrate availability, from deficiency to excess. Raman and SERS signals were compared to off-line metabolite analysis of carbohydrates, carboxylic acids, and nucleotides. Results demonstrate that SERS, in almost all cases, led to a higher number of identifiable signals and better resolved spectra. Spectra derived from the TG-Raman were comparable to those of micro-Raman resulting in well-discernable Raman peaks, which allowed for the identification of a higher number of compounds. In contrast, NIR-Raman provided a superior performance for the quantitative evaluation of analytes, both with and without SERS nanoparticles when using multivariate data analysis. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., :1-10, 2018.

Time-gated Raman spectroscopy and proteomics analyses of hypoxic and normoxic renal carcinoma extracellular vesicles

“…A maximum of 5 mW of CW excited laser emission was guided through a lens (Leica, N Plan, Wetzlar, Germany) with a magnification factor of 20× and numerical aperture (NA) of 0.4, while the spectral acquisition time to the samples was set to 10 s. In comparison, the overall measurement time for each sample with TG-RS/SERS was set to be 3 times 60 s including repetitions to achieve an appropriate signal without sample evaporation. For performing RS measurements small aluminum microwells with a cavity in µL scale were used, as previously shown 14 . This allows to first measure the Raman response of isolated EV samples while subsequently measure the SERS response.…”

“…“gating out” the sample derived autofluorescence. TG-RS has been applied earlier to biological samples 14 – 17 (for more detailed information about the TG-RS see Kögler and Heilala 18 ).…”

Assessment of recombinant protein production in E. coli with Time-Gated Surface Enhanced Raman Spectroscopy (TG-SERS)

“…The final concentration for hCNTF and HspA1 1 hour after induction was approximately 2.4 * 10 8 cells/ml, considering the volume of NC and Ag NP solutions. The aluminum has proven to not interfere with the measurements 27,28 . In addition, due to the relatively low intensity of background from complex media from the E. coli cell samples with NC and Ag NPs this contribution was not subtracted from final spectra (Fig.…”

Automated Bioprocess Feedback Operation in a High-Throughput Facility via the Integration of a Mobile Robotic Lab Assistant

“…Especially in HT scenarios, the frequency of sampling to determine the status of microbial cultures can be a major challenge (Janzen et al, 2019). Although a variety of online, inline, and atline probes for realtime process analytics exist and essential substrates or metabolites can be estimated in situ (Lee et al, 2004), methods such as Raman spectroscopy or near-infrared spectroscopy (Tamburini et al, 2014;Kögler et al, 2018) introduce additional complexity with regard to data handling, model calibration, and analysis (Mercier et al, 2014). A typical approach to overcome this issue is the implementation of enzymatic assays for atline determination of metabolites and substrates.…”