Study of hydrogel properties and immobilization of a bioluminescent bioreporter

by 1982- Balasundaram, Arun

The past decade has witnessed the development of a novel class of sensors; Bioluminescent Bioreporter Integrated Chip biosensors, designed to accurately measure small physical changes in the atmosphere using genetically engineered micro-organisms (bioreporters). The major challenge that now remains is to design a suitable matrix that can hold the bioreporters functionally active over a period of time. The project is an effort to develop and demonstrate alternative methods to favorably immobilize bioreporters without affecting its metabolic functions. The wide collection of literature indicates the successful use of hydrogels for cellular immobilization over the past few years. Hydrogels are inexpensive, easy to fabricate in laboratory conditions, chemically inert, biocompatible, structurally stable, optically transparent and more importantly permeable to target analytes. The equilibrium swelling properties and membrane potential of the hydrogels were studied to gain sufficient insight into its characteristic response over long periods. The knowledge was then used to control the mechanical properties such as stiffness, porosity and surface charge of the hydrogel scaffolds to exactly meet the design criteria of an ideal immobilization matrix. The study particularly involved tests to immobilize cells of Pseudomonas fluorescens 5RL, a genetically engineered bioluminescent bioreporter in the volume and surface of charged polyelectrolyte hydrogels and alginate gels. The bioluminescent light assays, Live dead assays, Electron microscopy were used to verify the viability of the immobilized bioreporters. The tests demonstrate the ability of the hydrogels in immobilizing the microorganisms without significantly affecting the physiology of the cells. The results indicate tremendous potential and a major role that hydrogels can play in the immobilization of iv microorganisms. Such successful techniques integrated with large scale commercialization could change the face of the conventional sensor technology in every possible areas like waste water remediation, medical diagnostics, sealed room gas analyzers in space shuttles and many more. v