Biological Studies with Advanced Microscopy

Second Harmonic Imaging of Collagen
Collagen, the most important structural protein in the body, is a strong generator of optical second harmonics - that is, light at half the wavelength of the illuminating light. This can be used to identify and characterise collagen in tissue. Out work is based around measuring the directional and polarization properties of the signal from different types of collagen, and on finding biological and medical applications for the technique.
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Properties and Function of Coral Fluorescent Pigments
Corals contain brightly coloured fluorescent proteins, which are involved in protection against excessive sunlight. These proteins have found many uses in cell biology since they can be expressed in other living cells and used as molecular tags. We are studying their function in coral ecology and physiology and also investigating the behaviour of newly-identified proteins which change colour when irradiated at specific wavelengths.
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Structural Organization of the fts-Z Protein in Bacillus
fts-Z is a protein with some homolgy to the tubulins, which are important in cell organization, division and motility in eukaryotic cells. fts-Z seems to have similar functions, expecially in cell division, in the much simpler bacterial cell. We will be using colour-changing coral fluorescent proteins to follow the dynamics of fts_Z through the cell cycle.
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Single-Molecule Electrical Studies of DNA on Silicon Surfaces
Understanding how electrons flow through single molecules is important in several areas: fabricating nanoelectronic devices and sensors; specially rationalising electron transfer (ET) in biological molecules. The nature of electrical transport along the DNA duplex is important in understanding DNA damage in vivo and in exploring the possible use of DNA in molecular electronics and direct detection of DNA hybridisation. This project aims to investigate ET properties of single DNA molecules as 'molecular wires' on silicon surface – and increase our understanding of the role of surfaces on the ET behaviour. This study will also attempt to exploit the fundamental knowledge gained to fabricate novel molecular electronic devices that utilise ET.
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Collaborators: Prof. Justin Gooding (School of Chemistry, University of New South Wales), .

Live-cell Imaging in 3-D Matrices
Live confocal and DIC imaging into 3-D matrices require careful calibration of matrix and coverslip thicknesses that enable imaging within a certain depth of view. Live imaging allowed us to record the parameters of cellular invasion into dense collagen gels such as speed and directionality. A physical mechanism of traction was discovered whereby cells utilised curved protrusions to hook around collagen fibres, gaining stronghold on the matrix during cancer cell invasion.
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Measurement of Molecular Dynamics Using a Photoconvertible Protein
Phamret is a photoconvertible protein that when highlighted by UV light stimulation, induces a change in fluorescence emission from cyan fluorescent protein (CFP) to photoactivated GFP (PA-GFP). Coupling Phamret to ROM, we have been able to confirm plasma membrane localisation of ROM and calculate the diffusion rate of the protein.
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Vesicle Tracking Using Total Internal Reflection Microscopy (TIRFM)
TIRFM allows high-resolution imaging at the ventral cell surface with several fold improvement in resolution over confocal microscopy. We have been able to track four kinds of vesicle transport including vesicle rocketing by de novo actin polymerisation with this microscopy technique. Furthermore, using GFP-ROM transfected cells, it was possible to associate ROM with vesicles that were transported via the rocketing mechanism.
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