Figure 1: Complete bioreactor assembly
Cambridge, MA
September-December 2024 hello For my senior capstone class 2.013 Engineering Systems Design, I worked with a team of 10 to design and prototype an innovative 100mL general purpose bioreactor. As 1 of 4 members of the Sensors and Electronics Control System (SECS) subteam, I worked on developing completely non-invasive sensing for 4 key parameters: 1) optical density (OD), 2) dissolved oxygen (DO), 3) pH, and 4) temperature. These parameters are critical for maintaining the proper environment within the bioreactor to enable healthy bacterial and cell
growth and maximize product yield.
Most bioreactors on the market use invasive sensing, which typically requires a probe to enter the reaction vessel and directly contact the broth inside. However, SECS pursued entirely non-invasive sensing to eliminate the need to sterilize or replace sensors after every use, which can become expensive, and to minimize the risk of contamination to the broth. Moreover, by decoupling the sensors from the disposable vessel, the bioreactor is easily serviceable, as individual components can be replaced without affecting the rest of the system.
Thorough documentation of our bioreactor design can be found at the following files:
Figure 2: Reaction vessel and sensor plate assembly.
Trenches at the bottom of the vessel act as poke-yoke features to correctly place it onto the sensor plate.
Trenches at the bottom of the vessel act as poke-yoke features to correctly place it onto the sensor plate.
Figure 3: Sensor packaging for OD, DO, pH, and temperature sensing and control. LEDs and photodiodes not pictured.
Design Spec: 0-2 ± 0.1au
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The OD sensor is used to measure the growth rate of the culture. This is necessary to identify and monitor the growth phase of the culture, which influences several other reaction parameters, such as feed rate, product yield, and reaction efficiency.
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Figure 4: Culture growth phases can be identified via Normalized OD and Growth Rate plots.
helloOD is based on the linear relationship between light absorbance and particle concentration, which can be well described by the Lambert-Beer Law. To measure the OD of the reaction, our product uses an LED, a reference photodiode, and two measurement photodiodes at angles 90° and 135°. Light of 610nm is emitted from the LED and scattered by suspended particles in the culture.
Some of this scattered light is detected by a photodiode, which measures its incident intensity. As the culture grows, optical density increases and results in a greater intensity of light detected by the photodiode due to increased scatter.hello
Figure 5: OD sensing trench on vessel
Figure 6: OD sensor working principle and major components
Design Spec: 4-9 pH
helloTo measure the pH of the reaction, we use a polyaniline (PANI) sensing layer, which changes its absorbance based on the pH that it is exposed to. The PANI layer is deposited on the inside of a “trench” at the bottom of the reaction vessel. An LED, reference photodiode, and sensing photodiode are packaged around the trench. Light is emitted at around 630nm, passes through the PANI layer, and is detected by a photodiode, which measures its incident intensity.
Figure 7: Absorbance vs emitted wavelength of 8µm thick PANI layer.
Figure 8: pH sening trench on vessel
Figure 9: pH sensor working principle and major components
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Figure 10: Materials used in the PANI layer reaction.
Figure 11: Evolution of PANI layer formation over 10 minutes of reaction. The solution color starts clear and gradually becomes dark green.
Figure 12: Five samples from the 2nd round of PANI layer testing.
Figure 13: Quick test for light read-through using a white LED iPhone flash.
Design Spec: 2-9 mg/L
helloTo measure the dissolved oxygen (DO) of the reaction, our product uses a sensing layer made of a Tris (4,7-diphenyl-1, 10-phenanthroline) ruthenium(II) Complex in Sol-Gel, which changes fluorescence based on the dissolved oxygen concentration it is exposed to. As the dissolved oxygen concentration increases in contact with the Ru-complex layer, the fluorescent intensity is quenched, and so the emitted light is decreased. This relationship is described well by the Stern-Volmer equation.
Light is emitted by an LED at 620nm onto the Ru-complex layer, generating the fluorescence effect and causing emission at 655nm. This signal is detected using a sensing photodiode, which is placed behind a 640nm longpass filter.
Figure 14: Luminescence spectrum of Ru-complex sensing layer
Figure 15: DO sensor location on vessel
Figure 16: DO sensor working principle and major components
Design Spec: 25-50 ± 0.5°C
helloTo measure the temperature of the culture broth non-invasively, we use a thermistor and a thermal pad on the outside of the reaction vessel. Since the sensor is located outside, the measured temperature is actually that of the vessel, rather than the broth inside. Thus, we needed to calculate the thermal time constant and found it to be approximately 20 seconds, which is within our design specification of 30 seconds.
Figure 17: Temperature sensor design and its thermal resistance network.
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