RESUMO
Microfluidics evolved with the appearance of polydimethylsiloxane (PDMS), an elastomer with a short processing time and the possibility for replication on a micrometric scale. Despite the many advantages of PDMS, there are well-known drawbacks, such as the hydrophobic surface, the absorption of small molecules, the low stiffness, relatively high cost, and the difficulty of scaling up the fabrication process for industrial production, creating a need for alternative materials. One option is the use of stiffer thermoplastics, such as the cyclic olefin copolymer (COC), which can be mass produced, have lower cost and possess excellent properties. In this work, a method to fabricate COC microfluidic structures was developed. The work was divided into process optimization and evaluation of material properties for application in microfluidics. In the processing step, moulding, sealing, and liquid handling aspects were developed and optimized. The resulting COC devices were evaluated from the point of view of molecular diffusion, burst pressure, temperature resistance, and susceptibility to surface treatments and these results were compared to PDMS devices. Lastly, a target DNA hybridization assay was performed showing the potential of the COC-based microfluidic device to be used in biosensing and Lab-on-a-Chip applications.
RESUMO
Antibody therapy has been one of the most successful therapies for a wide range of diseases, including cancer. One way of expediting antibody therapy development is through phage display technology. Here, by screening thousands of randomly assembled peptide sequences, it is possible to identify potential therapeutic candidates. Conventional screening technologies do not accommodate perfusion through the system, as is the case of standard plate-based cultures. This leads to a poor translation of the experimental results obtained in vitro when moving to a more physiologically relevant setting, such as the case of preclinical animal models or clinical trials. Microfluidics is a technology that can improve screening efficacy by replicating more physiologically relevant conditions such as shear stress. In this work, a polydimethylsiloxane/polystyrene-based microfluidic system for a continuously perfused culture of cancer cells is reported. Human colorectal adenocarcinoma cells (HCT116) expressing CXCR4 were used as a cell target. Fluorescently labeled M13 phages anti-CXCR4 were used to study the efficiency of the microfluidic system as a tool to study the binding kinetics of the engineered bacteriophages. Using our microfluidic platform, we estimated a dissociation constant of 0.45 pM for the engineered phage. Additionally, a receptor internalization assay was developed using SDF-1α to verify phage specificity to the CXCR4 receptor. Upon receptor internalization there was a signal reduction, proving that the anti-CXCR4 fluorescently labelled M13 phages bound specifically to the CXCR4 receptor. The simplicity and ease of use of the microfluidic device design presented in this work can form the basis of a generic platform that facilitates the study and optimization of therapies based on interaction with biological entities such as mammalian cells.
