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Bioreactor Analyte Monitoring with NIR Spectroscopy

Introduction
  • Bioreactors are an important emergent tool for development of pharmaceuticals, cells and tissue
  • Bioreactors are transitioning from research to mainstream production environments
  • Currently, there is significant undercapacity in the pharmaceutical bioreactor market
  • Opportunity costs are in the hundreds of millions because demand for bioreactor produced drugs cannot be met
  • InLight Solutions has demonstrated technical feasibility for a Bioreactor monitor that enables optimal cell growth, higher reliability, shorter runs, and superior yields

Most bioreactions are batch processes
  • Bioreactor run can last as long as 120 days, so losing a batch is potentially very costly
  • Significant effort is put into monitoring to ensure a batch run is successful
    • Based on checks, new growth media is infused and waste removed to facilitate cell growth
    • In general, the process is labor intensive and open loop between adjustments
  • Fluid dynamics limit the capacity of a bioreactor
    • Labor requirements to monitor and adjust a bioreactor limit production scale-up

Measurement of glucose and lactate during monitoring, control
  • The commonly measured analytes (oxygen, carbon dioxide, pH and temperature) don’t give a complete picture of the cell reaction
  • Measurement of glucose, lactate during a bioreactor run usually requires removal of liquid from system
    • Potential for contamination
    • Labor intensive
    • Undersampling of reaction
    • 30+ minute time lag between acquiring sample, bench-top/lab analysis and adjustment of bioreactor
  • Monitoring gaps and lack of sensor integration make closed-loop control a challenge
    • Growth not optimized, batch runs take longer, cells stressed

NIR spectroscopy expands bioreactor monitoring capabilities
  • Additional important analytes for monitoring cell metabolism include:
    • Glucose, glutamine (food for cells)
    • Lactate (waste product)
  • Other analytes of interest include:
    • Urea, bicarbonate, ammonia
  • Potential for monitoring additional amino acids, sugars, proteins, and lipids

Advantages of NIR spectroscopy for bioreactor monitoring
  • Continuous monitoring of bioreactor
    • Reduced chance of contamination because system not compromised by periodic removal of bioreactor media for measurement/monitoring
    • More info to characterize, control and optimize the process resulting in shorter runs, higher reliability and superior yields
  • Multiple analytes measured with a single spectral measurement
    • Flash analysis of bioreactor media
    • No need for discrete sensor per analyte
  • Multivariate spectroscopy has built-in diagnostic capabilities that provide confidence in the results

InLight is a leader in the development of NIR spectroscopy applications
  • InLight has spent 10 years developing a noninvasive glucose monitor based on NIR spectroscopy
  • InLight has extensive experience and expertise in measuring numerous analytes in vitro and in vivo
    • In vitro work includes whole blood, serum and aqueous media
    • In vivo work includes glucose, alcohol, urea and arterial blood gases in tissue
  • Bioreactor application plays to InLight’s strengths:
    • Spectroscopy, chemometrics, statistics
    • Clinical chemistry, tissue optics, clinical research
    • Optical, mechanical and electrical engineering

 

NASA Bioreactor Monitoring Feasibility Project

 

Project overview
  • InLight collaborated with NASA to demonstrate proof of concept
  • InLight developed a prototype based on our proprietary Fourier transform spectrometer equipped with a NIR compatible flow cell
  • InLight demonstrated simultaneous measurement by near infrared spectroscopy of the following analytes in GTSF-2 cell culture media:
    • Glucose, glutamine
    • Lactate, urea
    • pH, HCO3-, pCO2

Bioreactor monitor concept breadboard

 

Picture of Breadboard w/o Cover

 

Bioreactor monitoring demonstration overview
  • Built custom breadboard system for demonstration
  • Developed calibrations for glucose, glutamine, lactate, urea, pH and HCO3-
    • Spectral measurement of doped GTSF-2 samples in which analytes of interest were intentionally varied in an orthogonal manner
    • Partial least squares regression technique applied to collected spectra and analyte reference values
    • pCO2 calculated from pH and HCO3-
  • Applied analyte calibrations prospectively to a set of validation samples
    • Results of validation summarized on following pages

Examples of NIR measurement performance Bioreactor monitor concept breadboard

 
 

  • As demonstrated in these plots, spectroscopic measurement error is independent of concentration. Competing sensor technologies generally have proportional errors.

NIR accurately measures analytes in GTSF-2
Analyte Concentration Range Standard Deviation Standard Error of Prediction
Glucose 19 – 350 mg/dL 98.2 4.7 mg/dL 0.998
Lactate 0 – 47 mg/dL 16.0 1.3 mg/dL 0.994
Urea 0 – 14 mg/dL 5.1 0.6 mg/dL 0.995
Glutamine 0 – 67 mg/dL 23.8 3.7 mg/dL 0.976
pH 7.0 – 7.9 0.24 0.026 0.989
Bicarbonate 8.7 – 21.2 mmol 3.2 0.7 mmol 0.932
pCO2 (calc) 5.7 – 69.5 mmHg 20.5 2.3 mmHg 0.996

NIR can measure additional analytes in GTSF-2
  • Based on InLight’s in vitro work in blood, plasma and serum, the bioreactor monitoring analyte menu can be expanded to include:
    • Ammonia
    • Albumin
    • Total Protein
    • Triglycerides
    • HDL, LDL and total cholesterol
    • Bilirubin
    • Fructose
    • Galactose

Summary
  • Demonstrated feasibility of bioreactor monitoring with NIR spectroscopy
    • Great accuracy and precision
    • Negligible protein coating of optical flow cell during 40 day bioreactor cell run
  • Good potential to measure other analytes in GTSF-2
    • Information already contained in spectral measurement
  • Integration of NIR continuous bioreactor monitoring with closed loop control will facilitate production
    • Optimal cell growth, higher reliability, shorter runs, superior yields
  • InLight is excited by this opportunity and is seeking an industry partner to move forward

 

For information on Bioreactor Analyte Monitoring Applications, please contact Info@InLightSolutions.com

 

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