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Development of Microfluidic Glucose Sensors BME 273: Kristen Jevsevar, Jason McGill, Sean Mercado, Rebecca Tarrant Advisors: Jennifer Merritt, Dr. John Wikswo, Dr. David Cliffel Department of Biomedical Engineering, Vanderbilt University, Nashville TN USA. 1.5”. 3/8” Diameter. 1/4” Diameter.

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slide1
Development of Microfluidic Glucose Sensors

BME 273: Kristen Jevsevar, Jason McGill, Sean Mercado, Rebecca Tarrant

Advisors: Jennifer Merritt, Dr. John Wikswo, Dr. David Cliffel

Department of Biomedical Engineering, Vanderbilt University, Nashville TN USA

1.5”

3/8” Diameter

1/4” Diameter

Nuts

PDMS

1/4” Ports

2.0”

7/32”

Electrode

Channel

Acrylic Plate

Bolts

2/10”

2/10”

4mm

Milled Holes

INTRODUCTION

  • Glucose is a vital component of cellular metabolism
  • Changes in metabolite consumption/product provide important information regarding cellular response
  • Metabolites are currently studied on the mL scale
  • Using microfluidics, studies can be done on μL and nL scales
  • Specially designed electrodes already exist that are able to measure glucose concentrations
    • This technology can be extended to multiple metabolites
    • They can be made more affordable while maintaining accuracy
  • Sensitivity of electrode: 172 nA/mM
  • Concentration range tested: 0 – 5 mM
  • Time delay: 80 seconds
  • Maximum flow tolerance: 300 μL/min
  • Tested durability: 6 hours

Figure 2. (left) A schematic of the Pine Instruments screen-printed electrode showing the working, reference, and counter electrodes. The microfluidic channel (red) is placed over these electrodes.

DESIGN COMPONENTS

DESIGN PERFORMANCE & RESULTS

Figure 3. (right) An enzyme film of glucose oxidase and nafion is spotted onto the electrode. A chemical reaction takes place on the electrode wherein glucose reacts with glucose oxidase to form hydrogen peroxide (H2O2). H2O2 is oxidized, releasing an electron. This generates a current, allowing the measurement of glucose concentrations.

Figure 6. Initial calibration experiments were done to determine if there was a linear response of the enzyme coating to glucose concentration. During these electrode precalibrations, when the electrode is placed in a beaker containing various concentrations of glucose, there is a linear relationship between glucose concentration and current.

channel

  • Electrode
  • Electrode Housing
  • Channel System
  • Pumping System
  • Pump Driver
  • Electrochemical Workstation
  • Bioreactor
  • Computer Software

Figure 4. A schematic of the electrode housing shown from above (left) and from the side (above). A CNC milling machine is used to make an acrylic negative. A PDMS mold is created using the negative. Acrylic plates with drilled holes are placed on either side of the electrode and PDMS mold. Nuts and bolts allow the plates to be tightened, sealing the system. Holes drilled through the upper plate allow tubing to connect to input and output tubing ports.

APPLICATIONS

Figure 7. In this experiment, the microfluidic device switches between continuous flow calibration fluids and stop/flow cell media from the multianalyte microphysiometer (MAMP) bioreactor. The continuous flow data is a smooth curve (circled in red) and the stop/flow data appears as repeating peaks (circled in green).

  • Pharmaceutical researchers are faced with the rapidly expanding challenge of designing and developing new drugs
  • Techniques to study cellular environment and metabolism are valuable for such development
  • Using a readily produced electrode and a microfluidic channel system, it is possible to measure glucose levels on a small scale

Figure 8. To determine if the microfluidic glucose sensor would work downstream of a cell culture, it was attached to the end of a MAMP. Initial experiments showed that it did measure glucose concentration downstream of cells without a loss of signal or disruption due to cellular byproducts. While running experiments downstream of the MAMP, glucose concentration was also being measured by a known device to act as a comparison. The peaks, indicating glucose consumption durinig the stop period of the stop/flow cycle, are the same for the MAMP data (blue) and our microfluidic data (red).

calibration solution

DESIGN CRITERIA

A valve is used to alternate flow between bioreactor media and calibration solutions.

  • Utilize commercially produced Pine Instruments electrode
  • Interface electrode with a microfluidic channel system, allowing microscale volumes to be studied in near real-time
  • Implement pumping system

FUTURE DIRECTION

  • Obtain results for other metabolites
  • Configure on-chip peristaltic pump
  • Interface with nanobioreactor

A bioreactor cultures a small amount of cells. An internal pump perfuses the cells with media, which is then sent to the electrode.

A LabView pump driver is used to control the Harvard Apparatus Pico Pump. The Harvard Apparatus pumps calibration solutions to the electrode.

Figure 1. Commercially produced Pine Instruments electrode.

  • Measure glucose concentrations within a biologically relevant range, between 0 mM and 6 mM
  • Affordable – less expensive than current research technology, more like disposable strips for diabetics
  • Function for at least 24 hours
  • Recalibrate automatically to account for electrochemical drift

Figure 5. This experimental diagram depicts the setup used to obtain glucose concentration measurements.

Glucose concentrations are measured using a CH Instruments electrochemical workstation. A potentiostat controls the electrical potential between the working and reference electrode, maintaining a constant voltage. The sensor detects current from the enzyme reaction.

waste products

ACKNOWLEDGEMENTS

Data is collected using CH Instruments computer software that plots current versus time.

  • Vanderbilt University Department of Biomedical Engineering
  • Dr. John Wikswo for advising and affording the team the resources needed to complete the project
  • Jennifer Merritt for acting as our graduate advisor, providing guidance, and assisting in experimental procedures
  • David Schaffer and Ron Reiserer for help in design and fabrication
  • The VIIBRE staff for advice and resources
  • Dr. Paul King and Alex Makowski for their constructive criticism and guidance through the design and development process
  • Design cost:
    • Electrode: $30
    • Tubing and electrode housing: ~$15
    • Chemicals (i.e. glucose oxidase): $60
  • Additional equipment cost:
    • Harvard Apparatus: $2,000
    • Bioreactor: $20
    • Electrochemical workstation: $2,000
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