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Jin D.,Macquarie University | Piper J.,Macquarie University | Yuan J.,Dalian University of Technology | Leif R.,Newport Instruments
Progress in Biomedical Optics and Imaging - Proceedings of SPIE | Year: 2010

In advanced cytometry, a fundamental challenge for rapid specific detection of rare-event micro-organisms is the autofluorescence noise from the complex biological samples. Time-gated luminescence can effectively discriminate labeled cells from autofluorescence background. Recently, a real-time true-colour time-gated luminescence microscopy system has been developed based on the synchronization of a solid-state excitation source and a super-fast optical shutter. We also developed a variety of ultra-bright silica nano-biolabels with multiple luminescence colours and controllable lifetimes in microsecond range. These developments allowed the development of an advanced cell analysis system for real-time background-free imaging and rare-event counting of microsecond-lifetime multi-colour labelled water-borne pathogens. © 2010 Copyright SPIE - The International Society for Optical Engineering. Source


Lu Y.,Macquarie University | Lu Y.,Tsinghua University | Jin D.,Macquarie University | Leif R.C.,Newport Instruments | And 5 more authors.
Cytometry Part A | Year: 2011

Many microorganisms have a very low threshold (<10 cells) to trigger infectious diseases, and, in these cases, it is important to determine the absolute cell count in a low-cost and speedy fashion. Fluorescent microscopy is a routine method; however, one fundamental problem has been associated with the existence in the sample of large numbers of nontarget particles, which are naturally autofluorescent, thereby obscuring the visibility of target organisms. This severely affects both direct visual inspection and the automated microscopy based on computer pattern recognition. We report a novel strategy of time-gated luminescent scanning for accurate counting of rare-event cells, which exploits the large difference in luminescence lifetimes between the lanthanide biolabels, >100 μs, and the autofluorescence backgrounds, <0.1 μs, to render background autofluorescence invisible to the detector. Rather than having to resort to sophisticated imaging analysis, the background-free feature allows a single-element photomultiplier to locate rare-event cells, so that requirements for data storage and analysis are minimized to the level of image confirmation only at the final step. We have evaluated this concept in a prototype instrument using a 2D scanning stage and applied it to rare-event Giardia detection labeled by a europium complex. For a slide area of 225 mm2, the time-gated scanning method easily reduced the original 40,000 adjacent elements (0.075 mm × 0.075 mm) down to a few "elements of interest" containing the Giardia cysts. We achieved an averaged signal-to-background ratio of 41.2 (minimum ratio of 12.1). Such high contrasts ensured the accurate mapping of all the potential Giardia cysts free of false positives or negatives. This was confirmed by the automatic retrieving and time-gated luminescence bioimaging of these Giardia cysts. Such automated microscopy based on time-gated scanning can provide novel solutions for quantitative diagnostics in advanced biological, environmental, and medical sciences. © 2011 International Society for Advancement of Cytometry. Source


N/A

Grant
Agency: Department of Health and Human Services | Branch: | Program: STTR | Phase: Phase II | Award Amount: 249.96K | Year: 1999

N/A


Trademark
Newport Instruments | Date: 2002-03-28

Laboratory Equipment, namely sample processor used for processing biological fluid samples used to manufacture cytologic or particle preparations.


Leif R.C.,Newport Instruments | Yang S.,Newport Instruments | Yang S.,Phoenix Flow Systems Inc. | Lu Y.,Macquarie University | And 3 more authors.
Progress in Biomedical Optics and Imaging - Proceedings of SPIE | Year: 2012

Time-gated luminescence images were obtained by analog summation of a series of sequential images that were obtained with a cooled modified interline CCD camera, and a fluorescence microscope modified to use a UV LED for illumination. The interline CCD camera obtains an analog sum of a multi-frame image by not reading out the storage line after each frame is acquired; instead, the charges from the acquisition pixels are transferred to the storage pixels, which adds them to those previously stored; subsequently, the sum of the images is readout from the storage pixels and digitized. The length of the exposure is limited by the capacity of the storage pixels and the rate of generation of background (noise). Previously, the quality of the images obtained with the room temperature camera was degraded by the buildup of thermal noise. The interline transfer, electronically shuttered, cooled astronomy CCD camera, which was modified for analog summation rapidly produced low noise images; yet permitted long exposures. The past problems with lanthanide dyes of low extinction coefficients and equipment cost have now been solved. © 2012 Copyright Society of Photo-Optical Instrumentation Engineers (SPIE). Source

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