Optimal treatment of cancer requires diagnostic methods to facilitate therapy choice and prevent ineffective treatments. Direct assessment of therapy response in viable tumor specimens could fill this diagnostic gap. Therefore, we designed a microfluidic platform for assessment of patient treatment response using tumor tissue slices under precisely controlled growth conditions. The optimized Cancer-on-Chip (CoC) platform maintained viability and sustained proliferation of breast and prostate tumor slices for 7 days. No major changes in tissue morphology or gene expression patterns were observed within this time frame, suggesting that the CoC system provides a reliable and effective way to probe intrinsic chemotherapeutic sensitivity of tumors. The customized CoC platform accurately predicted cisplatin and apalutamide treatment response in breast and prostate tumor xenograft models, respectively. The culture period for breast cancer could be extended up to 14 days without major changes in tissue morphology and viability. These culture characteristics enable assessment of treatment outcomes and open possibilities for detailed mechanistic studies. SIGNIFICANCE: The Cancer-on-Chip platform with a 6-well plate design incorporating silicon-based microfluidics can enable optimal patient-specific treatment strategies through parallel culture of multiple tumor slices and diagnostic assays using primary tumor material.

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We present an extremely compact field effect transistor (FET)-based electrochemical sensor for in situ real-time and label-free measurement of ion concentrations in the cell culture area of organs-on-chip (OoCs) devices. This sensor replaces the functionality of an external reference electrode, crucial in standard electrochemical sensing, by controlling the FET threshold voltage via a capacitive control gate. The silicon- and polymer-based charge sensor can be integrated in OoC platforms by means of a wafer-scale and CMOS-compatible microfabrication process. This fabrication approach inherently allows a superior level of accuracy, repeatability and scalability compared to common OoC manufacturing methods. The sensor combines in a single device the complementary benefits of silicon-based electronics and of flexible polymer membranes with integrated microelectrodes – congenial substrates to sustain dynamic stimuli and mimic physiological tissue microenvironments. The integration of the polymer membrane in the sensing region makes this miniature sensor a preferable option for high sensitivity biochemical measurements in OoC applications, including monitoring the pH of cell culture media and of tissue culturing microenvironments, quantification of ion displacement in cells, and complementary research on disease modeling.@en

We present the first Organ-on-Chip equipped with a low-impedance Poly(3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS) MicroElectrode Array (MEA). The novel device allows simultaneous mechanical stimulation with a stretchable PDMS membrane and electrical monitoring via the PEDOT:PSS MEA of multiple in vitro cell cultures. The surface area enhancement and the morphology of the PEDOT:PSS allows an increase of the charge injection per unit area at the electrode-electrolyte interface, resulting in significantly lower electrochemical impedance of the electrodes. In particular, at 1 kHz the fabricated PEDOT-MEA electrodes show a reduction of the overall impedance up to 99.4 and 93.3 % in comparison with benchmark TiN and Pt electrodes. The superior performance of PEDOT:PSS were also confirmed via Cyclic Voltammetry measurement, in which PEDOT:PSS showed a very large capacitive current, compared with the benchmark electrodes both in the forward and the reverse scans. The obtained results confirm the effectiveness of the proposed PEDOT:PSS coating, and introduce this material in the OOC field. Moreover, the quality and morphology of the fabricated PEDOT:PSS based electrodes were assessed via SEM imaging and Raman spectroscopy.

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Monitoring cell conditions and microenvironment in real time is crucial for Organ-on-Chip (OoC) functionality. In particular, biological cues such as ions, including metals and metabolites, play a critical role in physiology and homeostasis in the human body. • Real-time monitoring of ions without optical systems is an unmet need for OOCs [1]. • Electrochemical sensors, such as organic electrochemical [2] and thin-film transistors [3], may address this need. Most of these sensors however rely on reference electrodes.@en

Drug development is a complex, time-consuming (10 - 15 years) and expensive process. For a new medicine to reach the market, the net expenses covered by the pharmaceutical industry have been estimated to be around $2.6 billion. Nevertheless, the risk of finding adverse effects or toxicity cases once the drug is already on the market is still high. Thus, pharmaceutical companies have been keenly looking forward to means to eliminate this at an early stage of the development process. Recently, Organs-on-Chips (OOCs) emerged as a potential alternative to traditional drug screening. These devices, promise, in the middle-term, to enhance the in vitro screening and, in the long term, to reduce and eventually eliminate animal models in safety and efficacy essays. Nevertheless, the fabrication methods for most of these devices are hardly adaptable to scalable fabrication processes for in vitro screening application, as they rely strongly on manual techniques. This thesis demonstrates the successful development of diverse microstructures for Organ-on-Chip applications by using scalable IC and MEMS-based fabrication techniques. Chapter 3 demonstrates the development of microfabricated porous PDMS membranes for barrier modelling. A simple and reproducible method to fabricate and transfer porous PDMS membranes with a high control on pore size, porosity, thickness, is shown. Very thin (thickness <10 ¹m) porous membranes with small features sizes down to 2 ¹mand porosity up to 65% can thus be fabricated and successfully transferred with high reproducibility. The presented results on cell transmigration, topology and barrier formation demonstrated the biocompatibility of the porous PDMS membranes. Chapter 4 shows further efforts towards the realization of manufacturable OOCs. A monolithically microfabricated OOC device, an alternative to the available devices capable to address many more applications, was demonstrated. Preliminary biological experiments indicate its biocompatibility as cells (HUVEC, Cardyomyocites) are successfully cultured and maintained viable in the microchannels and the silicon cavity. Finally, Chapter 5 demonstrates other possibilities allowed by the use of IC and MEMS techniques. The integration of microstructures that enable transduction mechanisms to monitor the cell microenvironment, is shown. Specifically, strain gauges for stress sensing as an alternative to monitor in situ strain in microfabricated OOCs, are presented. Relative resistance changes of approximately 0.008% and 1.2% for titanium and polymeric strain gauges have been observed, respectively. The technological advances shown in this thesis form a significant contribution towards manufacturable fabrication of Organs-on-Chips and the standardization of OOCs as routinely used tools for drug development. @en

Organ-on-chip (OOC) is becoming the alternative tool to conventional in vitro screening. Heart-on-chip devices including microstructures for mechanical and electrical stimulation have been demonstrated to be advantageous to study structural organization and maturation of heart cells. This paper presents the development of metal and polymeric strain gauges for in situ monitoring of mechanical strain in the Cytostretch platform for heart-on-chip application. Specifically, the optimization of the fabrication process of metal titanium (Ti) strain gauges and the investigation on an alternative material to improve the robustness and performance of the devices are presented. The transduction behavior and functionality of the devices are successfully proven using a custom-made set-up. The devices showed resistance changes for the pressure range (0-3 kPa) used to stretch the membranes on which heart cells can be cultured. Relative resistance changes of approximately 0.008% and 1.2% for titanium and polymeric strain gauges are respectively reported for membrane deformations up to 5%. The results demonstrate that both conventional IC metals and polymeric materials can be implemented for sensing mechanical strain using robust microfabricated organ-on-chip devices.

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We present a novel method to easily and reliably transfer highly porous, large area, thin microfabricated Polydimethylsiloxane (PDMS) porous membranes on Lab-on-Chip (LOC) and Organ-on-Chip (OOC) devices. The use of silicon as carrier substrate and a water-soluble sacrificial layer allows a simple and reproducible transfer of the membranes to any PDMS-based OOC and LOC device. The use of IC and MEMS compatible techniques reduces significantly the fabrication time and the need of manual handling. Our method is suitable for automatic assembling systems, such as pick-and-place, crucial to significantly increase the throughput of OOC and LOC devices assembling. Membranes with 8 μm pore size and as thin as 4 μm are successfully transferred. The viability and biocompatibility of the transfer was assessed by culturing two different cell lines on an OOC with transferred porous PDMS membranes.

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We present a novel and highly reproducible process to fabricate transferable porous PDMS membranes for PDMS-based Organs-on-Chips (OOCs) using microelectromechanical systems (MEMS) fabrication technologies. Porous PDMS membranes with pore sizes down to 2.0 μm in diameter and a wide porosity range (2–65%) can be fabricated. To overcome issues normally faced when using replica moulding and extend the applicability to most OOCs and improve their scalability and reproducibility, the process includes a sacrificial layer to easily transfer the membranes from a silicon carrier to any PDMS-based OOC. The highly reliable fabrication and transfer method does not need of manual handling to define the pore features (size, distribution), allowing very thin (<10 μm) functional membranes to be transferred at chip level with a high success rate (85%). The viability of cell culturing on the porous membranes was assessed by culturing two different cell types on transferred membranes in two different OOCs. Human umbilical endothelial cells (HUVEC) and MDA-MB-231 (MDA) cells were successfully cultured confirming the viability of cell culturing and the biocompatibility of the membranes. The results demonstrate the potential of controlling the porous membrane features to study cell mechanisms such as transmigrations, monolayer formation, and barrier function. The high control over the membrane characteristics might consequently allow to intentionally trigger or prevent certain cellular responses or mechanisms when studying human physiology and pathology using OOCs.

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This paper presents an advanced voltammetric system to be used as electronic tongue for liquid and gas analysis. It has been designed to be more flexible and accurate with respect to other existing and similar systems. It features improved electronics and additional operative conditions. Among others these include the possibility to optically excite the solution and to treat the output signal by a differentiation process in order to better evidence the existence of small details in the response curve. Finally by the same type of tongue preliminary results are shown dealing with O2 and CO2 concentration measurements in appropriate solutions.@en

Polymeric (PEDOT:PSS) strain gauges embedded in PDMS membranes fabricated using a full wafer-scale fabrication process capable of realizing reproducible small features, are reported. The devices are characterized using a customized setup, which provides mechanical stretch while dynamically reading the electrical resistance. Measurements show relative resistance changes of approximately 11% for applied pressure up to 4 kPa. The process described is tailored to fabricate pressure sensors and microelectrodes for a flexible substrate-based Organ-on-Chip platform.

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Organ-on-Chips (OOCs) are micro-fabricated devices which are used to culture cells in order to mimic functional units of human organs. The devices are designed to simulate the physiological environment of tissues in vivo. Cells in some types of OOCs can be stimulated in situ by electrical and/or mechanical actuators. These actuations can mimic physiological conditions in real tissue and may include fluid or air flow, or cyclic stretch and strain as they occur in the lung and heart. These conditions similarly affect cultured cells and may influence their ability to respond appropriately to physiological or pathological stimuli. To date, most focus has been on devices specifically designed to culture just one functional unit of a specific organ: lung alveoli, kidney nephrons or blood vessels, for example. In contrast, the modular Cytostretch membrane platform described here allows OOCs to be customized to different OOC applications. The platform utilizes silicon-based micro-fabrication techniques that allow low-cost, high-volume manufacturing. We describe the platform concept and its modules developed to date. Membrane variants include membranes with (i) through-membrane pores that allow biological signaling molecules to pass between two different tissue compartments; (ii) a stretchable micro-electrode array for electrical monitoring and stimulation; (iii) micro-patterning to promote cell alignment; and (iv) strain gauges to measure changes in substrate stress. This paper presents the fabrication and the proof of functionality for each module of the Cytostretch membrane. The assessment of each additional module demonstrate that a wide range of OOCs can be achieved.
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Sensing and stimulating microstructures are necessary to develop more specialized and highly accurate Organ-on-Chip (OOC) platforms. In this paper, we present the integration of a conductive polymer, poly (3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT:PSS), on a stretchable membrane, core element of an Heart-on-Chip. The electrical conductivity along with its biocompatibility, high transparency (≈88%) and mechanical elasticity (≈1.2 GPa) make this material a candidate to develop novel microstructures for electrical monitoring and stimulation of cells in flexible-substrate based OOCs. Microstructures with different shapes and geometries of PEDOT:PSS embedded in a 9 μm-thick Polydimethylsiloxane (PDMS) membrane are developed following a wafer-level fabrication approach. PEDOT:PSS layers between 120 nm and 300 nm are obtained by varying the deposition conditions. The layers are successfully patterned and microstructures with lateral dimensions down to 2 μm. The obtained results indicate that this polymer is a suitable material for microfabrication of sensing and stimulating elements in OOC platforms.

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