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Please use this identifier to cite or link to this item: http://hdl.handle.net/1959.3/76950
- Nanometre localisation of quantum dots based on their stochastic blinking properties and (F)PALM rendering algorithms
- Kouskousis, Betty; van Embden, Joel; Morrish, Dru; McMahon, Kerrie-Ann; Russell, Sarah; Gu, Min
- Fluorescence imaging is routinely used to obtain information on the spatial organisation of proteins in cells and other biological tissues. However due to the limit of diffraction imposed by conventional optical techniques, images can only be resolved within 250-300 nm. In response to this, several methods capable of breaking the diffraction barrier have been developed, in particular (fluorescence) photoactivatable localisation microscopy (F)PALM [1, 2]. (F)PALM is based on the serial photoactivation and subsequent bleaching of numerous sparse subsets of photoactivatable fluorescent protein (PA-FP) probes within a specimen. Due to the small subset of PA-FP probes activated, individual molecules can thereby be precisely localised to obtain a highly resolved map (image) with an effective resolution of tens of nanometres. These exciting new technologies are likely to revolutionise cell biology, but are yet unavailable to most biologists. We describe here experiments towards developing a high-resolution system for biological imaging. As a first step towards developing the algorithms needed for such a system, we have utilised semiconductor nanocrystals or quantum dots (QDs). The attractive optical properties of QDs include narrow emission bands, emission wavelength tuneability with size, photostability and enhanced brightness. The stochastic blinking behaviour of QDs provide opportunities for superresolution without the need for serial activation and bleaching of FA-FPs . In addition QDs provide a simple model for developing the algorithms needed for (F)PALM, as an alternative to PA-FPs, has been explored in this work. A final high resolution spatial map (image) of randomly dispersed QDs is obtained using independently written localisation and rendering algorithms based on the techniques described by [1, 2]. Furthermore, the results indicate that localisation and photon summing precision rely heavily on the signal to noise (SNR) ratio of the detection system; where the localisation precision was found to rely heavily on the particle density of the sample. We will describe progress in applying (F)PALM and (QD)PALM to biological imaging.
- Publication type
- Conference paper
- Paper presented at the Inaugral Light in Life Sciences Conference (LILS09), Melbourne, Australia, 24-27 November 2009
- Publication year
- FABLS Research Network
- Publisher URL
- Copyright © 2009.