Advanced Biosensing towards Real-Time Imaging of Protein Secretion from Single Cells

Protein secretion of cells plays a vital role in intercellular communication. The abnormality and dysfunction of cellular protein secretion are associated with various physiological disorders, such as malignant proliferation of cells, aberrant immune function, and bone marrow failure. The heterogeneity of protein secretion exists not only between varying populations of cells, but also in the same phenotype of cells. Therefore, characterization of protein secretion from single cell contributes not only to the understanding of intercellular communication in immune effector, carcinogenesis and metastasis, but also to the development and improvement of diagnosis and therapy of relative diseases. In spite of abundant highly sensitive methods that have been developed for the detection of secreted proteins, majority of them fall short in providing sufficient spatial and temporal resolution for comprehensive profiling of protein secretion from single cells. The real-time imaging techniques allow rapid acquisition and manipulation of analyte information on a 2D plane, providing high spatiotemporal resolution. Here, we summarize recent advances in real-time imaging of secretory proteins from single cell, including label-free and labelling techniques, shedding light on the development of simple yet powerful methodology for real-time imaging of single-cell protein secretion

Clickable surfaces for the selective recognition of oligosaccharides

With the emergent role of protein glycosylation as a potential biomarker of disease, new technologies are required to selectively recognise unique glycan structures symptomatically expressed in disease states. Currently, glycan recognition is achieved through methods lacking high affinity and selectivity or through complex analytical methods and are poorly suited to clinical laboratories. In this thesis we propose an alternative glycan sensing strategy based on molecular self-assembly and imprinting of glycan ligands with phenyl boronic acid (PBA), imbuing selective oligosaccharide recognition to a surface. This strategy employs extensive use of the copper catalysed click reaction providing a simple highly biocompatible and adaptable self-assembled monolayer intended to form a foundation for future modifications. The synthesis and characterisation of various SAM monomers has been described here alongside azide functionalised-PBA CuAAC ligands and PBA terminated SAM monomers. SAMs formed with these were characterised for surface organisation and the CuAAC reaction yield established thus confirming reaction utility in functionalisation of alkyne terminated SAMs. Saccharide binding of PBA functional SAMs was then examined accounting for surface organisation between SAM monomer designs. Having determined a suitable SAM for functionalisation, we then produced a novel oligosaccharide-PBA imprinted surface demonstrating high selectivity between structurally different oligosaccharides

Recent Advances in Surface Plasmon Resonance Imaging Sensors

The surface plasmon resonance (SPR) sensor is an important tool widely used for studying binding kinetics between biomolecular species. The SPR approach offers unique advantages in light of its real-time and label-free sensing capabilities. Until now, nearly all established SPR instrumentation schemes are based on single- or several-channel configurations. With the emergence of drug screening and investigation of biomolecular interactions on a massive scale these days for finding more effective treatments of diseases, there is a growing demand for the development of high-throughput 2-D SPR sensor arrays based on imaging. The so-called SPR imaging (SPRi) approach has been explored intensively in recent years. This review aims to provide an up-to-date and concise summary of recent advances in SPRi. The specific focuses are on practical instrumentation designs and their respective biosensing applications in relation to molecular sensing, healthcare testing, and environmental screening