Novel microfluidic unit operations enabling cell handling on a pressure-driven lab-on-chip system

Abstract: Lab-on-a-Chip (LoC) systems offer a way to perform point-of-care analysis of patient samples in a microfluidic environment. Nowadays, there are numerous commercialized systems based on different technologies for various applications. Within that context, it is desirable to combine as many processing and analysis workflows as possible in one system to cover a broad range of different applications. One application area becoming increasingly important is the examination of cells in the field of tumor diagnostics. Accordingly, the aim of this work is to extend an existing LoC system in terms of microfluidic unit operations and functionalities to enable cell transport and analysis. In particular, the Vivalytic system of Bosch Healthcare Solutions GmbH is studied exemplarily, which is a pressure-driven system based on elastomeric membranes.

Three topics were addressed in the present work: I) Characterization and simulative design of the micropump as well as determination of the influencing parameters on fluidic processes. Both, a contact-free, fluorescence-based measurement method for determining transient flow rates and an analytical modeling approach for the pressure-based deflection of elastomeric membranes in the fluidic network were investigated. II) Development and implementation of new pumping mechanisms for the processing of fluids in the microfluidic system. Here, options for controlled pumping and new peristaltic pumping processes were investigated based on flow rate measurements as well as model calculations. III) Investigations on the influence of microfluidic stimuli on cells transported in the LoC system. In particular, the viability of the cells and changes on the molecular level were examined. Microfluidic chips with defined structures were fabricated by rapid prototyping to investigate the influence of different parameters. Furthermore, experiments were performed in the cartridge of the Vivalytic system to investigate the influence of the previously developed pumping mechanisms on the cells. Finally, this work addressed the implementation of a unit operation in the field of liquid biopsy for the retention of tumor cells from a blood sample. The cell retention element was also designed and optimized in the microfluidic chip, followed by a transfer onto the Vivalytic cartridge.

For the first topic, the results of this work include the calculation suitable concentrations, verification of the linearity assumption, and validation of the volume determination through the fluorescence-based measurement method. This shows that the optical setup and subsequent image analysis enable contact-free determination of the volume flow rate. For the standard pumping process in the Vivalytic system, maximum flow rates between 180 and 700 µL/s are measured exemplary, depending on the fluidic path and additional elements such as the filter unit. Through the analytical model, the nonlinear pressure-based deflection of an elastomeric membrane is derived from the Young-Laplace equation, and the modeled cubic relationship between applied pressure and deflection of the membrane is verified experimentally. Transient volumetric flows in the microfluidic network are calculated via differential equations based on Hagen Poiseuille's law. A comparison between modeling and measurement method shows that system-dependent parameters such as the course of the actuation pressure have to be integrated into the model in order to enable a reliable calculation of the fluidic processes. The combination of the measurement method and analytical model provides helpful insights into dynamics and fluidic relations in a membrane-based LoC.

Based on this understanding, in the second part, new pumping mechanisms are developed and implemented in the Vivalytic system. These include pressure- and viscosity-controlled pumping, reducing maximum flow rates in the system to < 100 µL/s. Furthermore, peristaltic pumping mechanisms are implemented using chambers and valves in the system, which in particular achieves a reduction in volume quantization from 20 µL (volume of a pump chamber) to volumes of 0.1 to 20 µL. In peristaltic pumping with valves, it is also shown that a non-pressurized elastomeric membrane in the fluidic path can smooth the pulsatility of the volume flow and maximum flow rates are reduced to < 10 µL/s.

Finally, in the third part of the present work, cell transport through the previously described pumping mechanisms and the development of a functionality for retaining cells in the studied LoC system are considered. Thereby, the hypothesis that cell characteristics such as viability and surface marker expression can be influenced by the processing in the microfluidic environment is confirmed. It is shown that flow rates above 50 µL/s and the resulting fluidic shear forces of 30 dyn/cm^2 can result in a decrease in viability of over 50%. Because cell viability decreases in a time range of 30 to 60 min, analysis of target cells should either be performed in a time range of 30 min, or the shear forces on the cells should be reduced by processing them at flow rates < 50 µL/s.
To develop a unit operation for retaining tumor cells from a blood sample using a filter element, the unit is particularly designed as a microfluidic chip so that the filter area is optically accessible. This allows the process to be observed in real-time to detect cells that are forced through the filter pores during the course of filtration due to the fluidic pressure applied on the filter. This is confirmed experimentally by an increased retention rate of 73 % when counting cells in real-time compared to 57 % when using an endpoint determination. Furthermore, the integration and processing of the retention unit on the Vivalytic cartridge is carried out, providing comparable results to the microfluidic chip.

Overall, this work paves the way towards a comprehensive microfluidic understanding of membrane-based, pressure-driven LoC systems for enhanced functionalities and applications in the field of cell biology. In the Vivalytic system, these findings can now be used to implement and optimize fluidic processes for the transport and filtration of tumor cells from a blood sample

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• Standort: Deutsche Nationalbibliothek Frankfurt am Main
• Umfang: Online-Ressource
• Sprache: Englisch
• Anmerkungen: Universität Freiburg, Dissertation, 2022

Klassifikation und Themen

• Schlagwort: Pressure ; Mikrofluidik ; Mikropumpe ; Elastomer ; Membranpumpe ; Tumorzelle ; Lab on a Chip ; Diagnostik

Beteiligte, Orts- und Zeitangaben

• Ereignis: Veröffentlichung
• (wo): Freiburg
• (wer): Universität
• (wann): 2022
• Urheber: Bott, Hannah
• Beteiligte Personen und Organisationen: Zengerle, Roland ; Woias, Peter ; Albert-Ludwigs-Universität Freiburg. Institut für Mikrosystemtechnik ; Albert-Ludwigs-Universität Freiburg. Fakultät für Angewandte Wissenschaften

Weitere Informationen

• DOI: 10.6094/UNIFR/228445
• URN: urn:nbn:de:bsz:25-freidok-2284459
• Rechteinformation: Open Access; Der Zugriff auf das Objekt ist unbeschränkt möglich.