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FIBS-enabled Noninvasive Metabolic Profiling
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FIBS-enabled noninvasive metabolic profiling.

Alireza Behjousiar1, Antony Constantinou, Karen M Polizzi

  • 1Centre for Process Systems Engineering, Department of Chemical Engineering and Chemical Technology, Imperial College London.

Journal of Visualized Experiments : Jove
|February 12, 2014
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Summary

Researchers developed novel in vivo biosensors for real-time monitoring of intracellular metabolites in Chinese hamster ovary (CHO) cells. This FRET integrated biological sensor (FIBS) technology enables noninvasive, accurate quantification of glucose and glutamine levels.

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Area of Science:

  • Computational biology
  • Systems biology
  • Biotechnology

Background:

  • High-throughput experimental systems are crucial for refining and validating computational models in biology.
  • Current methods for time-course data collection often involve destructive sampling, limiting real-time, noninvasive monitoring.
  • In vivo biological sensors offer a promising alternative for real-time, noninvasive monitoring of intracellular conditions.

Purpose of the Study:

  • To develop and validate a novel in vivo biosensor system for real-time, noninvasive monitoring of intracellular metabolite concentrations.
  • To construct Chinese hamster ovary (CHO) cell lines stably expressing FRET integrated biological sensors (FIBS) for glucose and glutamine.
  • To calibrate the FIBS in batch cell cultures for accurate in vivo quantification of intracellular metabolites.

Main Methods:

  • Construction of CHO cell lines engineered to constitutively express FRET integrated biological sensors (FIBS).
  • Development of FIBS based on Förster Resonance Energy Transfer (FRET) between fluorescent proteins linked by a ligand-binding domain.
  • In vivo calibration of FIBS in batch cell cultures and validation using fed-batch cultures and independent enzymatic assays.

Main Results:

  • Successfully generated CHO cell lines expressing functional FIBS for glucose and glutamine.
  • Demonstrated that FIBS exhibit spectral changes correlating with intracellular metabolite concentrations.
  • Validated the accuracy of FIBS-based quantification of intracellular glucose and glutamine using independent enzymatic assays.

Conclusions:

  • FRET integrated biological sensors (FIBS) provide a robust platform for real-time, noninvasive monitoring of intracellular metabolites in CHO cells.
  • This technology enables accurate quantification of key metabolites like glucose and glutamine, essential for bioprocess optimization.
  • FIBS represent a significant advancement for predictive modeling and bioprocess control in computational biology.