A Hypoxia-Maintaining Perfusion Device for Prolonged Cell Studies
Ameziane, Karim (2024)
2024
Bioteknologian ja biolääketieteen tekniikan maisteriohjelma - Master's Programme in Biotechnology and Biomedical Engineering
Lääketieteen ja terveysteknologian tiedekunta - Faculty of Medicine and Health Technology
This publication is copyrighted. You may download, display and print it for Your own personal use. Commercial use is prohibited.
Hyväksymispäivämäärä
2024-10-16
Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:tuni-202410089152
https://urn.fi/URN:NBN:fi:tuni-202410089152
Tiivistelmä
Oxygen plays a critical role in many of the biological processes of the human body. Nevertheless, it is often overlooked in cell and tissue culture. Organ-on-chip applications aiming to control oxygen levels offer suitable solutions to overcome this issue. Namely, with adequately engineered cell culture platforms, it is possible to expose cell and tissue cultures to various oxygen conditions such as hypoxia and physioxia.
This thesis presents a one-millimetre-thick device the size of a microscope slide consisting of a perfusable medium channel surrounded by a gas channel that controls oxygen levels. The device is capable of setting and maintaining hypoxia while simultaneously implementing continuous medium perfusion. This is especially suitable for long-term monitoring outside of incubator conditions when aiming to maintain set hypoxia.
This thesis also documents the fabrication of said device. Its channels were fabricated from a single polydimethylsiloxane (PDMS) bulk using a 3D-printed mould and were sealed with a microscope slide by plasma-bonding. The topmost layer of the device consists of a laser-cut oxygen barrier that isolates the medium channel from ambient oxygen conditions.
Characterization measurements were performed as part of the main objectives of this work. This was possible with the use of an optical ratiometric oxygen-sensing layer coating the microscope slide. This characterization process involved measuring oxygen concentrations from the device’s cell culture region in both flow and no-flow conditions. Additionally, the maximum flow rate at which the device is capable of maintaining desired oxygen levels was identified. The obtained data in these various conditions was analysed and time points of interest of the dynamic response were determined using appropriate approaches.
Additionally, a finite element simulation model of the device was built to replicate its behaviour in silico. This model was successfully capable of reproducing the step responses observed in the measured area of its physical counterpart in both flow and no-flow conditions. This model was then used to estimate the dynamic response at the identified flow rate limit as well as gaining insight on oxygen behaviour in parts of the medium channel outside of the measured area.
The measurement results indicate that the fall and rise times of the device under a 4 µL/min flow rate are 52 min and 37 min respectively when oxygen concentration in the gas channel is switched from 19% to 0% and vice versa. Under no-flow conditions, these same times become 2.1 h and 1.5 h respectively. The flow rate limit at which the device can upkeep set oxygen levels was identified to be 10 µL/min. Moreover, the simulation model estimates the fall and rise times to be 27 min and 26 min respectively at said limit. Additionally, replicating the step response measurements with a 3 mm thick device show that a thicker PDMS bulk slows down the device dynamics.
Overall, the device presented in this thesis was successfully capable of combining both oxygen control and perfusion of the medium channel. This device could be used in future works involving cell cultures to better study their behaviour under both hypoxia and normoxia.
This thesis presents a one-millimetre-thick device the size of a microscope slide consisting of a perfusable medium channel surrounded by a gas channel that controls oxygen levels. The device is capable of setting and maintaining hypoxia while simultaneously implementing continuous medium perfusion. This is especially suitable for long-term monitoring outside of incubator conditions when aiming to maintain set hypoxia.
This thesis also documents the fabrication of said device. Its channels were fabricated from a single polydimethylsiloxane (PDMS) bulk using a 3D-printed mould and were sealed with a microscope slide by plasma-bonding. The topmost layer of the device consists of a laser-cut oxygen barrier that isolates the medium channel from ambient oxygen conditions.
Characterization measurements were performed as part of the main objectives of this work. This was possible with the use of an optical ratiometric oxygen-sensing layer coating the microscope slide. This characterization process involved measuring oxygen concentrations from the device’s cell culture region in both flow and no-flow conditions. Additionally, the maximum flow rate at which the device is capable of maintaining desired oxygen levels was identified. The obtained data in these various conditions was analysed and time points of interest of the dynamic response were determined using appropriate approaches.
Additionally, a finite element simulation model of the device was built to replicate its behaviour in silico. This model was successfully capable of reproducing the step responses observed in the measured area of its physical counterpart in both flow and no-flow conditions. This model was then used to estimate the dynamic response at the identified flow rate limit as well as gaining insight on oxygen behaviour in parts of the medium channel outside of the measured area.
The measurement results indicate that the fall and rise times of the device under a 4 µL/min flow rate are 52 min and 37 min respectively when oxygen concentration in the gas channel is switched from 19% to 0% and vice versa. Under no-flow conditions, these same times become 2.1 h and 1.5 h respectively. The flow rate limit at which the device can upkeep set oxygen levels was identified to be 10 µL/min. Moreover, the simulation model estimates the fall and rise times to be 27 min and 26 min respectively at said limit. Additionally, replicating the step response measurements with a 3 mm thick device show that a thicker PDMS bulk slows down the device dynamics.
Overall, the device presented in this thesis was successfully capable of combining both oxygen control and perfusion of the medium channel. This device could be used in future works involving cell cultures to better study their behaviour under both hypoxia and normoxia.