Nanofabrication of Microfluidic Devices

As one of the most successful known pathogens of mankind, Mycobacterium tuberculosis, the causative agent of TB, kills two million people every year. The bacterium is often able to persist as a long-term asymptomatic infection, known as latent tuberculosis. Microfluidics can be defined as the science and technology of systems that process or manipulate minute amounts of fluids. As a distinct new field, it seeks to influence scientific areas from chemical and biological synthesis to engineering and information technology. Microfluidic devices recently investigated the growth kinetics of individual E. coli cells and demonstrated sub-populations of fast growing antibiotic-sensitive cells and slow growing ‘persistent’ cells that are antibiotic-resistant. This phenomenon is thought to be important in understanding latent tuberculosis.

Microfluidic devices are constructed using various lithographic techniques to investigate single-cell growth of E. coli and M. smegmatis cells. Confocal microscopy is used to visualise cell growth and mathematical modelling provides an insight into the nature of the persistence. The microfluidic devices consist of a polydimethylsiloxane (PDMS) layer embedded with small flow channels (1.5-0.5 µm), a cellulose membrane and a PDMS layer (~250 µm) with large flow channels (~25 µm) for introducing and draining the cell culture media and antibiotic solution.

E-beam lithography trials have produced chrome stamps, which are used to pattern PDMS through soft-lithography resulting in the formation of micro-channels. Individual E. coli cells are dispersed within the PDMS channels. The production of a complete device should demonstrate linear growth of the cells within the channel dimensions. Continuous supply of cell culture media and subsequent treatment with antibiotics should identify the ‘persistent’ strain.