Automated microfluidic platform of bead-based electrochemical immunosensor integrated with bioreactor for continual monitoring of cell secreted biomarkers

Abstract

There is an increasing interest in developing microfluidic bioreactors and organs-on-a-chip platforms combined with sensing capabilities for continual monitoring of cell-secreted biomarkers. Conventional approaches such as ELISA and mass spectroscopy cannot satisfy the needs of continual monitoring as they are labor-intensive and not easily integrable with low-volume bioreactors. This paper reports on the development of an automated microfluidic bead-based electrochemical immunosensor for in-line measurement of cell-secreted biomarkers. For the operation of the multi-use immunosensor, disposable magnetic microbeads were used to immobilize biomarker-recognition molecules. Microvalves were further integrated in the microfluidic immunosensor chip to achieve programmable operations of the immunoassay including bead loading and unloading, binding, washing, and electrochemical sensing. The platform allowed convenient integration of the immunosensor with liver-on-chips to carry out continual quantification of biomarkers secreted from hepatocytes. Transferrin and albumin productions were monitored during a 5-day hepatotoxicity assessment in which human primary hepatocytes cultured in the bioreactor were treated with acetaminophen. Taken together, our unique microfluidic immunosensor provides a new platform for in-line detection of biomarkers in low volumes and long-term in vitro assessments of cellular functions in microfluidic bioreactors and organs-on-chips.

Multisensor-integrated organs-on-chips platform for automated and continual in situ monitoring of organoid behaviors

“…We developed label-free electrochemical immunobiosensors for continual in situ monitoring of soluble biomarkers secreted by the organoids (48,49). Here, we have further integrated these immunobiosensors with a set of specifically designed microfluidic chips to achieve fully automated operations of the sensors for high-sensitivity and longterm organoid monitoring in our platform.…”

“…To realize these procedures, we further fabricated an in-house microelectrode system for easy integration with microfluidic chips (48,49). The system consisted of three microelectrodes, a reference electrode (RE), a working electrode (WE), and a counter electrode (CE).…”

“…S12 A and B). Au was selected as the material for WE due to its relatively good stability, favorable electron transfer kinetics with high in-plane conductivity, biocompatibility, and its ability to readily create covalent bonding for generating stable immobilization of receptors onto its surface (48,49). The annealing process allowed us to create robust and smooth microelectrodes (SI Appendix, Fig.…”

“…A resealable microbioreactor for perfusion culture of organoids was developed by modifying our recently published protocols (48,54,55). The microbioreactor was designed to possess two hemichambers embedded within a pair of PDMS (top) and PDMS/glass (bottom) pieces, which were then sandwiched between two rigid supports made of PMMA and a thin PDMS cushion layer to ensure hydraulic tightness (Fig.…”

Label‐Free and Regenerative Electrochemical Microfluidic Biosensors for Continual Monitoring of Cell Secretomes

“…The diameters of the bubbles were found to dramatically decrease and they were found to disappear after about 35 s when using a chip with pillars having diameters of 500 µm (Figure S8, Supporting Information). In this microfluidic setup, the introduction of all the reagent solutions into the chip can be completely achieved in an automated manner, therefore eliminating the possibility of releasing toxic reagents including K 3 Fe(CN) 6 , SAM, and ethanol from the electrochemical sensing unit to the media in the bioreactor 38. Furthermore, during the impedance measurements using 50 × 10 −3 m K 3 Fe(CN) 6 solution, we block the flow to the detection chamber upon closure of the valves to obtain accurate impedance measurements and to reduce noise.…”