Professor Goldys with her team in the ARC Centre of Excellence for Nanoscale Biophotonics develops new “windows to the body”, new methodologies to probe and interrogate nanoscale biochemistry in living organisms. The program spans the follwing areas:
There is a shortage of automated and unbiased methods to rapidly characterize biological tissues and cells without the use of intrusive chemical procedures. Of specific interest are methods which are able to identify specific cell subpopulations, an area of intense interest in the life sciences as well as those able to be used for ongoing cell and tissue monitoring. Methods able to deal with complex biological heterogeneity are fundamentally important for biology and medicine.
We developed an automated reagent-free method for cellular characterization based on intrinsic fluorescence microscopy. The method provides a rapid, inexpensive and sensitive insight into sample biochemistry. It uses autofluorescence, light emitted by native molecules found in all cells. In our approach the fluorescence images of live cells are obtained at a number of selected excitation wavelengths and their emission is captured in a specified, longer wavelength range. These data are obtained using a standard microscope with a sensitive camera and a customised inexpensive light source comprising multiple light emitting diodes. Autofluorescence micrographs of cell populations are then analysed using custom-developed software to gather information about hundreds of quantitative features including size, circularity, intensity at each wavelength and texture. Multiple types of data analysis developed within our software enable us to:
1) Capture statistically meaningful differences between cell subpopulations. For example by using these specialised statistical tests we have been able to differentiate between healthy and diseased cells from patients suffering from a mitochondrial disease. We have also been able to distinguish the diseased cells from the cells treated with a specific drug and compare the treated cells to the healthy controls. This makes it possible to monitor progress of therapy.
2) Our method provides detailed insights into cell biochemistry, and we have been able to identify the abundances of several native fluorophores including free and bound NADH, flavins, retinoids and tryptophan. Their cellular maps and average cell values enabled us to detect subtle physiological features such as the metabolic rate. Trends over time, for example in stem cell differentiation experiments have been obtained.
3) We have been able to identify previously undetected cell subpopulations with respect to these abundances. These subpopulations of the cells previously regarded as homogeneous are sometimes completely separate.
4) The method is extremely sensitive, for example it is able to detect stem cell differentiation as early as 24 h after onset.
5) A suitable combination of autofluorescence features can be correlated with expression of specific surface biomarkers. Hence cells that are positive with respect to this marker do not need to be stained to be positively identified.
Our method has been successfully applied to the following types of cells and tissues:
• Olfactory neurospheres in metabolic disease
• Adipose-derived stem cells before and after osteogenic differentiation.
• Breast cancer MCF10A and MCF7 cells
• Pancreatic cancer and specific genetic mutants
• Diabetic tissue which we can distinguish from control tissue by their autofluorescence.
• Tissues (kidney and human placenta) affected by tobacco smoke
• Cattle embryos where we distinguish normal from compromised development
• Retina tissue and retinal cells
• Motor neurone disease cells
• IPS cells
Partnerships: This suite of projects are carried out in partnership with Kolling Institute in Sydney, Macquarie Cancer Institute and Macquarie Hospital.
As part of the program in the ARC Centre of Excellence for Nanoscale Biophotonics, our current work is targeting the following areas:
• Hardware development
• Multispectral image analysis
• Applications in medicine and integration with current medical practice
• Applications in biology and medical research
As part of the efforts within the Centre of Excellence in Nanoscale Biophotonics we develop technologies for biomolecular sensing on a nanoscale. Our programs in this area aim to develop sensing technologies that are able to be used in real time and which do not require washing steps. They need to be able to be used in physiological fluids. Of specific interest are nanoparticle sensors that are normally off and report fluorescence only in the presence of analyte as well as technologies with intrinsic amplification. Primary focus is on analytes of interest to CNBP including cytokines, metalloproteases, ROS and NO.
Partnerships: Regeneous Pty Ltd, University of Adelaide
Associated PhD projects
1. Nanoparticle based intracellular and extracellular immunosensors for the ultrasensitive detection of cell secreting molecules
2. Nanoparticles for intracellular and extracellular “switch-on” sensing with fluorescent reporters
3. Development of a high throughput affinity capture surface on living cells for selecting high secretors of molecules of interest
This program focuses on developing new assay formats. Building on and modifying assay biochemistry, we develop devices and approaches to be able to probe biochemistry in living organisms, as well as cells and tissues. Examples of projects include analysing cell secretions, monitoring progress of interventions into the spinal cord in living animals and building fluidic devices to examine ultrasmall samples of body fluids in real time.
Partnerships: CSIRO, University of Adelaide
Associated PhD projects:
1. Development of a sensing device to examine protein biomarkers based on ultrasmall samples of body fluids in real time
2. Intravital optical sensing of key analytes
3. Nanophotonics solutions for ultrasensitive biosensing
This program focuses on nanoparticles as tools to induce chemical change, with emphasis on targeted and/or triggered approaches. Within this context we develop diagnostic and therapeutic nanomaterials combining nanoparticles and specific molecular moieties. These are characterised and we develop new understanding of photophysical and photochemical mechanisms of nanomaterial action. We apply these theranostic nanomaterials in specific health contexts (diabetes, cancer, neurodegenerative diseases) and quantify and monitor the effectiveness of our nanomaterials in these applications.
Partnerships; Garvan Institute, Sydney Vital, Heart Research Institute, Sydney
Associated PhD projects:
1. Nanoparticles for gene delivery
2. Photodynamic therapy in deep tissue with nanoparticles
3. Multifunctional nanomaterials for medical early diagnostics