Newport Instruments

San Diego, CA, United States

Newport Instruments

San Diego, CA, United States
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Lu Y.,Macquarie University | Zhao J.,Macquarie University | Zhang R.,Macquarie University | Liu Y.,Macquarie University | And 16 more authors.
Nature Photonics | Year: 2014

Optical multiplexing plays an important role in applications such as optical data storage, document security, molecular probes and bead assays for personalized medicine. Conventional fluorescent colour coding is limited by spectral overlap and background interference, restricting the number of distinguishable identities. Here, we show that tunable luminescent lifetimes τ in the microsecond region can be exploited to code individual upconversion nanocrystals. In a single colour band, one can generate more than ten nanocrystal populations with distinct lifetimes ranging from 25.6 μs to 662.4 μs and decode their well-separated lifetime identities, which are independent of both colour and intensity. Such 'τ-dots' potentially suit multichannel bioimaging, high-throughput cytometry quantification, high-density data storage, as well as security codes to combat counterfeiting. This demonstration extends the optical multiplexing capability by adding the temporal dimension of luminescent signals, opening new opportunities in the life sciences, medicine and data security. © 2013 Macmillan Publishers Limited. All rights reserved.

Spidlen J.,BC Cancer Agency | Moore W.,Stanford University | Brinkman R.,BC Cancer Agency | Almarode J.,FlowJo LLC | And 17 more authors.
Cytometry Part A | Year: 2015

The lack of software interoperability with respect to gating has traditionally been a bottleneck preventing the use of multiple analytical tools and reproducibility of flow cytometry data analysis by independent parties. To address this issue, ISAC developed Gating-ML, a computer file format to encode and interchange gates. Gating-ML 1.5 was adopted and published as an ISAC Candidate Recommendation in 2008. Feedback during the probationary period from implementors, including major commercial software companies, instrument vendors, and the wider community, has led to a streamlined Gating-ML 2.0. Gating-ML has been significantly simplified and therefore easier to support by software tools. To aid developers, free, open source reference implementations, compliance tests, and detailed examples are provided to stimulate further commercial adoption. ISAC has approved Gating-ML as a standard ready for deployment in the public domain and encourages its support within the community as it is at a mature stage of development having undergone extensive review and testing, under both theoretical and practical conditions. © 2015 International Society for Advancement of Cytometry.

Zheng X.,Macquarie University | Lu Y.,Macquarie University | Zhao J.,Macquarie University | Zhang Y.,National University of Singapore | And 9 more authors.
Analytical Chemistry | Year: 2016

Compared with routine microscopy imaging of a few analytes at a time, rapid scanning through the whole sample area of a microscope slide to locate every single target object offers many advantages in terms of simplicity, speed, throughput, and potential for robust quantitative analysis. Existing techniques that accommodate solid-phase samples incorporating individual micrometer-sized targets generally rely on digital microscopy and image analysis, with intrinsically low throughput and reliability. Here, we report an advanced on-the-fly stage scanning method to achieve high-precision target location across the whole slide. By integrating X- and Y-axis linear encoders to a motorized stage as the virtual "grids" that provide real-time positional references, we demonstrate an orthogonal scanning automated microscopy (OSAM) technique which can search a coverslip area of 50 × 24 mm2 in just 5.3 min and locate individual 15 μm lanthanide luminescent microspheres with standard deviations of 1.38 and 1.75 μm in X and Y directions. Alongside implementation of an autofocus unit that compensates the tilt of a slide in the Z-axis in real time, we increase the luminescence detection efficiency by 35% with an improved coefficient of variation. We demonstrate the capability of advanced OSAM for robust quantification of luminescence intensities and lifetimes for a variety of micrometer-scale luminescent targets, specifically single down-shifting and upconversion microspheres, crystalline microplates, and color-barcoded microrods, as well as quantitative suspension array assays of biotinylated-DNA functionalized upconversion nanoparticles. © 2015 American Chemical Society.

Spidlen J.,Cancer Research Center | Moore W.,Stanford University | Parks D.,Stanford University | Goldberg M.,Becton Dickinson | And 17 more authors.
Cytometry Part A | Year: 2010

The flow cytometry data file standard provides the specifications needed to completely describe flow cytometry data sets within the confines of the file containing the experimental data. In 1984, the first Flow Cytometry Standard format for data files was adopted as FCS 1.0. This standard was modified in 1990 as FCS 2.0 and again in 1997 as FCS 3.0. We report here on the next generation flow cytometry standard data file format. FCS 3.1 is a minor revision based on suggested improvements from the community. The unchanged goal of the standard is to provide a uniform file format that allows files created by one type of acquisition hardware and software to be analyzed by any other type. The FCS 3.1 standard retains the basic FCS file structure and most features of previous versions of the standard. Changes included in FCS 3.1 address potential ambiguities in the previous versions and provide a more robust standard. The major changes include simplified support for international characters and improved support for storing compensation. The major additions are support for preferred display scale, a standardized way of capturing the sample volume, information about originality of the data file, and support for plate and well identification in high throughput, plate based experiments. Please see the normative version of the FCS 3.1 specification in Supporting Information for this manuscript (or at in the Current standards section) for a complete list of changes. © 2009 International Society for Advancement of Cytometry.

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.

Lu Y.,Macquarie University | Lu J.,Macquarie University | Zhao J.,Macquarie University | Cusido J.,University of Miami | And 10 more authors.
Nature Communications | Year: 2014

Significant multiplexing capacity of optical time-domain coding has been recently demonstrated by tuning luminescence lifetimes of the upconversion nanoparticles called 'τ-Dots'. It provides a large dynamic range of lifetimes from microseconds to milliseconds, which allows creating large libraries of nanotags/microcarriers. However, a robust approach is required to rapidly and accurately measure the luminescence lifetimes from the relatively slow-decaying signals. Here we show a fast algorithm suitable for the microsecond region with precision closely approaching the theoretical limit and compatible with the rapid scanning cytometry technique.We exploit this approach to further extend optical time-domain multiplexing to the downconversion luminescence, using luminescence microspheres wherein lifetimes are tuned through luminescence resonance energy transfer.We demonstrate real-time discrimination of these microspheres in the rapid scanning cytometry, and apply them to the multiplexed probing of pathogen DNA strands. Our results indicate that tunable luminescence lifetimes have considerable potential in high-throughput analytical sciences. © 2014 Macmillan Publishers Limited. All rights reserved.

Jin D.,Macquarie University | Lu Y.,Macquarie University | Leif R.C.,Newport Instruments | Yang S.,Newport Instruments | And 2 more authors.
Current Protocols in Cytometry | Year: 2014

The sensitivity of filter-based fluorescence microscopy techniques is limited by autofluorescence background. Time-gated detection is a practical way to suppress autofluorescence, enabling higher contrast and improved sensitivity. In the past few years, three groups of authors have demonstrated independent approaches to build robust versions of time-gated luminescence microscopes. Three detailed, step-by-step protocols are provided here for modifying standard fluorescent microscopes to permit imaging time-gated luminescence. © 2014 by John Wiley & Sons, Inc.

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.

Leif R.C.,Newport Instruments | Yang S.,Newport Instruments | Yang S.,Phoenix Systems
Progress in Biomedical Optics and Imaging - Proceedings of SPIE | Year: 2010

Problem: Previous images of time-gated luminescence have been obtained with a cooled CCD camera by digitally summing a series of sequential images. The data acquisition rate of approximately 10 one millisecond exposure images per second was rate limiting and too slow for standard research and clinical use. An annoying undulating background was present, which could not be totally removed by subtraction of an unexposed, control image. Solution: An analog approach to this problem is to use an interline transfer, electronically shuttered camera. After each exposure, the storage line is not readout; instead, the electrons from the acquisition pixels are transferred to the storage pixels and thus are added to those previously stored. The length of the exposure is limited by the capacity of the storage pixels and the rate of generation of background (noise) electrons. This electronic concept was tested with a Point Grey Dragonfly2 640 by 480 pixel monochrome camera equipped with a Sony 1/3" progressive interline scan, electronically shuttered CCD, which since it did not have any cooling, was operated at room temperature. Pulsed excitation was from a Nichia UV LED. Results: Five and 0.5 micron uniform europium complex stained microspheres could at room temperature be imaged with time-gated excitation and acquisition times of 1 millisecond each and analog summation of 50 images. Conclusion: The analog integration solution apparently works; however, a cooled scientific grade camera with the same capacity for multiple transfers into storage pixels would be better suited for use with dimmer luminescent objects. © 2010 Copyright SPIE - The International Society for Optical Engineering.

Farkas D.L.,Cedars Sinai Medical Center | Leif R.C.,Newport Instruments | Nicolau D.V.,University of Liverpool
Progress in Biomedical Optics and Imaging - Proceedings of SPIE | Year: 2011

This PDF file contains the front matter associated with SPIE Proceedings Volume 7902, including the Title Page, Copyright information, Table of Contents, Introduction, and the Conference Committee listing. © 2011 Copyright Society of Photo-Optical Instrumentation Engineers (SPIE).

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