Genetic and cellular analysis on microfluidic chips

Abstract

In the past two decades, microfluidics has gone through a fast development, offering researchers better solutions for problems in various fields, such as chemical synthesis, chemical and biological analysis, and biomedical studies. There are a lot of advantages for this technique such as low consumptions of sample and reagents, less analytical time, faster heat transfer rate, and high throughputs for conducted parallel experiments. However, as the booming of genomic and proteomic era, the unprecedented demands for new microfluidic devices are being raised. In this thesis, I focused on fabricating and applying new microchips for biological studies, such as genetic analysis (Part I), single cell analysis (Part II), drug screening and cellular microenvironment or mechanics (Part III). In the first part, we employed PLL-g-PMOXA and PLL-g-PEG copolymers as the antifouling materials for chip surface passivation which greatly improved on-chip PCR efficiency. We fabricated a multi-well PCR chip for simultaneous multiple diseases verification and genotype identification. HPV genotypes of real clinical samples were identified as demonstration on this chip. In the second part, we explored single cell RT-PCR on a multi-well SiO₂ chip. GADPH transcripts in single cell were successfully amplified. In addition, we designed a PDMS microfluidic chip for high-throughput single cell proteomic study. As a preliminary study, we successfully demonstrated successive labelling of three different proteins (exogenously expressed GFP, TRPV4-V5 and β-tubulin) inside a single cell. This chip gave a glimpse for on-chip high throughput single cell proteomics study. In the third part, we fabricated a chip to generate a polydopamine gradient on a hydrophobic surface. The polydopamine modified the substrate properties and facilitated further coating of ECM proteins onto the substrate for cellular analysis. Furthermore we developed a stretchable and micropatterned parafilm membrane for study stem cell osteogenic differentiation. With this membrane, both mechanical and spatial cues were combined together for adipose-derived mesenchymal stem cells (ADMSCs) osteogenesis differentiation study. Finally, we fabricated a PDMS-glass hybrid chip which could generate a broad stiffness gradient to simulate in vivo mechanical microenvironment. With this chip we studied etoposide-induced HeLa cell apoptosis on PDMS substrates with different Young’s modulus. Our results demonstrated that both drug concentration and substrate stiffness can influence the anti-tumor drug efficiency.

Date of Award	2014
Original language	English
Awarding Institution	The Hong Kong University of Science and Technology