The work packages described in this report cover the implementation of complex DNA reactions on chip, the in situ formation of hydrogels in microchannels and the use of microfluidics for performing a precise sample dilution step on chip.
ESR1 – Complex DNA reactions on chip
ESR1 designed, fabricated and characterized a programmable hydraulic resistor array for silicon microfluidic chips using electrogate arrays. Taking advantage of an electrogate array the flow rate inside microfluidic chips can be programmed easily without using external and bulky equipment. As an illustration she demonstrated the possibility to use such an array to create laminar flows and “tune” an enzymatic assay in a capillary-driven microfluidic chip. The signal of these enzymatic assays was localized on receptor-coated microbeads trapped in microfluidic structures.
ESR1 also worked on the implementation of complex nucleic acid reactions inside capillary-driven microfluidic chips taking advantage of a self-coalescence module (SCM) and spotting of reagents inside the chip. An SCM allows the reconstitution of spotted reagents inside the chip in a precise manner. In particular, two isothermal reactions, a molecular beacon and a more complex clamped hybridization chain reaction (C-HCR), have been implemented in chip. For both reactions, the concentration of an initiator DNA sequence was measured using a fluorescence measurement. The formation of fluorescent DNA polymers was observed inside the chip as a result of a C-HCR (Fig. 2). Such results demonstrated the possibility to implement and study complex reaction systems inside microfluidic chips.
Moreover, ESR1 worked on a module allowing the filtration of red blood cells. Using a 3D printed device, she filtered successfully red blood cells from a few microliters of a whole blood sample, which allowed the filling of capillary-driven microfluidic chips with plasma only.
In summary, it was elaborated that the various techniques developed by ESR 1 can be combined to pave the way for rapid, accurate, and amplified DNA-based assays in silicon microfluidic chips.
ESR2 – Sequential assays
ESR2 developed an innovative method for forming hydrogels with precise geometries in a sealed microfluidic chip. This method can be used to create in situ compartments on chip, filter samples, and localize bioassays. In particular, ERS2 identified a robust two-component chemical system for performing a fast and reliable polymerization reaction for forming PEG-based hydrogels. The polymerization reaction is based on thiol-maleimide coupling and can be performed in physiological conditions. ERS2 designed a specific capillary-driven microfluidic chip for shaping the interface of the two precursor solutions. This allows to form hydrogels with a well-defined geometry in a sealed microfluidic chip by interfacial polymerization within 2 minutes. The molecular weight cut off (MWCO) of the hydrogel was characterized by monitoring the diffusion of fluorescent molecules with different molecular weights through the hydrogel. Moreover, ERS2 was able to change the MWCO of the hydrogel easily, by tuning the concentration of the precursor solutions and by modifying the length of the PEG chains of the precursors. ERS2 demonstrated that receptors such as antibodies could be easily immobilized in the hydrogel and would retain their functionality. This allowed to perform a novel format of heterogeneous competitive assay on chip, which does not require a rinsing step.
Additionally, ERS2 also implemented a microfluidic module for the precise metering of two liquids by performing volumetric mixing. Such a module can be used for example for diluting a sample solution with a precise volume of buffer/reagents independently on their viscosity. Sample dilution is a critical step in rapid tests, which is often performed by the user and is prone to errors. Therefore, it could be useful to integrate such a dilution module in rapid tests to automate sample dilution and make tests more reliable.
