Hydrogel patterns in microfluidic devices by do-it-yourself UV-photolithography suitable for very large-scale integration

The interest in large-scale integrated (LSI) microfluidic systems that perform highthroughput biological and chemical laboratory investigations on a single chip is steadily growing. Such highly integrated Labs-on-a-Chip (LoC) provide fast analysis, high functionality, outstanding reproducibility at low cost per sample, and small demand of reagents. One LoC platform technology capable of LSI relies on specific intrinsically active polymers, the so-called stimuli-responsive hydrogels. Analogous to microelectronics, the active components of the chips can be realized by photolithographic micro-patterning of functional layers. The miniaturization potential and the integration degree of the microfluidic circuits depend on the capability of the photolithographic process to pattern hydrogel layers with high resolution, and they typically require expensive cleanroom equipment. Here, we propose, compare, and discuss a cost-efficient do-it-yourself (DIY) photolithographic set-up suitable to micro-pattern hydrogel-layers with a resolution as needed for very large-scale integrated (VLSI) microfluidics. The achievable structure dimensions are in the lower micrometer scale, down to a feature size of 20 µm with aspect ratios of 1:5 and maximum integration densities of 20,000 hydrogel patterns per cm. Furthermore, we demonstrate the effects of miniaturization on the efficiency of a hydrogel-based microreactor system by increasing the surface area to volume (SA:V) ratio of integrated bioactive hydrogels. We then determine and discuss a correlation between ultraviolet (UV) exposure time, cross-linking density of polymers, and the degree of immobilization of bioactive components. © 2020 by the authors

Hypoxia-on-a-chip

In this work a microfluidic cell cultivation device for perfused hypoxia assays as well as a suitable controlling unit are presented. The device features active components like pumps for fluid actuation and valves for fluid direction as well as an oxygenator element to ensure a sufficient oxygen transfer. It consists of several individually structured layers which can be tailored specifically to the intended purpose. Because of its clearness, its mechanical strength and chemical resistance as well as its well-known biocompatibility polycarbonate was chosen to form the fluidic layers by thermal diffusion bonding. Several oxygen sensing spots are integrated into the device and monitored with fluorescence lifetime detection. Furthermore an oxygen regulator module is implemented into the controlling unit which is able to mix different process gases to achieve a controlled oxygenation. First experiments show that oxygenation/deoxygenation of the system is completed within several minutes when pure nitrogen or air is applied to the oxygenator. Lastly the oxygen input by the pneumatically driven micro pump was quantified by measuring the oxygen content before and after the oxygenator

Closed-loop control system for well-defined oxygen supply in micro-physiological systems

To improve cell vitality, sufficient oxygen supply is an important factor. A deficiency in oxygen is called Hypoxia and can influence for example tumor growth or inflammatory processes. Hypoxia assays are usually performed with the help of animal or static human cell culture models. The main disadvantage of these methods is that the results are hardly transferable to the human physiology. Microfluidic 3D cell cultivation systems for perfused hypoxia assays may overcome this issue since they can mimic the in-vivo situation in the human body much better. Such a Hypoxia-on-a-Chip system was recently developed. The chip system consists of several individually laser-structured layers which are bonded using a hot press or chemical treatment. Oxygen sensing spots are integrated into the system which can be monitored continuously with an optical sensor by means of fluorescence lifetime detection

Mikrofluidisches System zur Kultivierung von oder der Analyse an lebenden Zellen oder Biomolekülen sowie ein Verfahren zu seiner Herstellung

Bei dem mikrofluidischen System zur Kultivierung von oder der Analyse an lebenden Zellen oder Biomolekülen sind mehrere übereinander angeordnete Laminate aus einem polymeren Werkstoff fluiddicht und stoffschlüssig miteinander verbunden. In mindestens einem der Laminate ist mindestens eine Öffnung oder mindestens ein Ausschnitt ausgebildet, mit der/dem ein Kanal oder eine Reservoir für die Aufnahme lebender Zellen oder Biomoleküle ausgebildet ist. In mindestens einem der Laminate ist mindestens eine weitere Öffnung ausgebildet, in die ein aus einem keramischen oder einem halbleitenden Werkstoff gebildeter Aktor/Sensor oder ein aus einem keramischen oder halbleitenden Werkstoff gebildeter Träger für einen Aktor/Sensor eingesetzt ist. Auf der Oberfläche des Laminats in das der Aktor/Sensor oder der Träger eingesetzt ist oder auf der Oberfläche eines unmittelbar an dieses Laminat angrenzenden Laminats ist mindestens eine elektrische Leiterbahn gedruckt, mit der eine elektrische Kontaktierung des Aktors und/oder Sensors erreichbar ist. Der mindestens eine Aktor/Sensor steht in berührendem Kontakt mit einer Flüssigkeit für die Kultivierung von Zellen, einer Flüssigkeit in der Biomoleküle enthalten sind oder den in einem Kanal oder Reservoir enthaltenen Zellen oder Biomolekülen

Microfluidic system for enhanced cardiac tissue formation

Hereby a microfluidic system for cell cultivation is presented in which human pluripotent stem cell-derived cardiomyocytes were cultivated under perfusion. Besides micro-perfusion this system is also capable to produce well-defined oxygen contents, apply defined forces and has excellent imaging characteristics. Cardiomyocytes attach to the surface, start spontaneous beating and stay functional for up to 14 days under perfusion. The cell motion was subsequently analysed using an adapted video analysis script to calculate beating rate, beating direction and contraction or relaxation speed

Hypoxia-on-a-chip - Generating hypoxic conditions in microfluidic cell culture systems

In this work a microfluidic cell cultivation device for perfused hypoxia assays as well as a suitable controlling unit are presented. The device features active components like pumps for fluid actuation and valves for fluid direction as well as an oxygenator element to ensure a sufficient oxygen transfer. It consists of several individually structured layers which can be tailored specifically to the intended purpose. Because of its clearness, its mechanical strength and chemical resistance as well as its well-known biocompatibility polycarbonate was chosen to form the fluidic layers by thermal diffusion bonding. Several oxygen sensing spots are integrated into the device and monitored with fluorescence lifetime detection. Furthermore an oxygen regulator module is implemented into the controlling unit which is able to mix different process gases to achieve a controlled oxygenation. First experiments show that oxygenation/deoxygenation of the system is completed within several minutes when pure nitrogen or air is applied to the oxygenator. Lastly the oxygen input by the pneumatically driven micro pump was quantified by measuring the oxygen content before and after the oxygenator

Closed-loop control system for well-defined oxygen supply in micro-physiological systems

To improve cell vitality, sufficient oxygen supply is an important factor. A deficiency in oxygen is called Hypoxia and can influence for example tumor growth or inflammatory processes. Hypoxia assays are usually performed with the help of animal or static human cell culture models. The main disadvantage of these methods is that the results are hardly transferable to the human physiology. Microfluidic 3D cell cultivation systems for perfused hypoxia assays may overcome this issue since they can mimic the in-vivo situation in the human body much better. Such a Hypoxia-on-a-Chip system was recently developed. The chip system consists of several individually laser-structured layers which are bonded using a hot press or chemical treatment. Oxygen sensing spots are integrated into the system which can be monitored continuously with an optical sensor by means of fluorescence lifetime detection. Hereby presented is the developed hard- and software requiered to control the oxygen content within this microfluidic system. This system forms a closed-loop control system which is parameterized and evaluated

Design, characterization, and modeling of microcirculation systems with integrated oxygenators

Here, we describe a microfluidic system for hypoxia assays on human cell culture models. These systems are developed to replace or reduce animal testing in biomedical basic research. The presented system uses a gas-permeable membrane as a gas–liquid interface and a micropump for media actuation to influence the oxygen content in two cell culture chambers. To apply well-defined hypoxic conditions to the cells, a good understanding of the mass transport phenomena is necessary. Therefore, a complete network model of the microfluidic system is presented. This model is validated by means of micro-particle image velocimetry (µPIV) and optical oxygen measurement with fluorescence lifetime detection. Finally, the impact of several process parameters, e.g., the gas permeability of the pump, is discussed using the developed model

Microfluidic system for enhanced cardiac tissue formation

Hereby a microfluidic system for cell cultivation is presented in which human pluripotent stem cell-derived cardiomyocytes were cultivated under perfusion. Besides micro-perfusion this system is also capable to produce well-defined oxygen contents, apply defined forces and has excellent imaging characteristics. Cardiomyocytes attach to the surface, start spontaneous beating and stay functional for up to 14 days under perfusion. The cell motion was subsequently analysed using an adapted video analysis script to calculate beating rate, beating direction and contraction or relaxation speed

3D printing – a key technology for tailored biomedical cell culture lab ware

Today’s 3D printing technologies offer great possibilities for biomedical researchers to create their own specific laboratory equipment. With respect to the generation of ex vivo vascular perfusion systems this will enable new types of products that will embed complex 3D structures possibly coupled with cell loaded scaffolds closely reflecting the in-vivo environment. Moreover this could lead to microfluidic devices that should be available in small numbers of pieces at moderate prices. Here, we will present first results of such 3D printed cell culture systems made from plastics and show their use for scaffold based applications