Lasers for Flow Cytometry

Shapiro, Howard M.

doi:10.1002/0471142956.cy0109s27

Cited by 4 publications

(2 citation statements)

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“…Our protocol can discriminate prokaryotic cells based on excitation and emission of a high quantum yield fluorescent DNA stain (SYTOX-green) detected by a PTM tube, which can detect individual photons at low-light levels (weak signals) (Steen, 1986, 1992, 2000; Hadfield, 2009). In contrast, photodiode detectors gather average light and detect high-light levels (strong signals) (Wood, 1997, 1998; Shapiro, 2004). In addition to detector types, particle scattering can be increased by reducing flow velocity; the slower the cells move through the focus the more scattered light they emit and can be detected (Steen, 2000).…”

Section: Discussionmentioning

confidence: 99%

A flow cytometric method to measure prokaryotic records in ice cores: an example from the West Antarctic Ice Sheet Divide drilling site

2016

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ABSTRACT. Microorganisms were the earliest inhabitants on our planet that occupy nearly every environment, and play a major role in biogeochemical cycles. Despite their global importance, there remains a paucity of data on microbial responses to long-term environmental and climatic changes. Microorganisms are known to be immured in glacial ice, but no high-resolution temporal records of their density exist, owing in large part to the lack of appropriate clean methodology that allows for rapid analysis of samples over depth. We describe a clean and time efficient method that can produce a high-temporal resolution record of prokaryotic density archived in ice cores. The method combines acquisition of discrete samples using a continuous ice-core melting system coupled with flow cytometry (FCM) of DNA-stained samples. Specifically, we evaluate the performance of the FCM measurement technique in terms of specificity, precision, accuracy and minimum detection limits. Examples from the West Antarctic Ice Sheet Divide ice core are included to show the efficacy of the method.

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Section: Discussionmentioning

confidence: 99%

A flow cytometric method to measure prokaryotic records in ice cores: an example from the West Antarctic Ice Sheet Divide drilling site

2016

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“…Modern instruments capable of highdimensional analysis (10 or more simultaneous fluorochromes) are typically equipped with solid-state lasers emitting at 488 nm (cyan), 633-640 nm (red), 532 or 561 nm (green or yellow), and 405 nm (violet). These four wavelengths allow excitation of fluorescein, phycoerythrin and its tandem dyes (488 and 532/561 nm), allophycocyanin and its tandems (633-640 nm), and the Brilliant Violet (BV) series of polymer dyes (405 nm) (3). In particular, the addition of violet laser diodes and the availability of the BV dye series have allowed a dramatic expansion of detection capability, allowing the simultaneous analysis of up to 16 fluorescent probes (4,5).…”

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Deep ultraviolet lasers for flow cytometry

Telford

Georges²,

Miller

et al. 2018

Cytometry

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Modern flow cytometers require multiple laser wavelengths to excite the wide variety of fluorescent probes now available for high‐dimensional analysis. Ultraviolet (UV) lasers (typically solid state 355 nm) have become a critical excitation source for the Brilliant Ultraviolet (BUV) series of polymer fluorochromes. The BUV dyes have pushed the number of fluorescent probes available for simultaneous analysis to nearly 30, allowing an unprecedented level of precision for immune cell analysis. However, immunologists are already seeking analyze more than 30 simultaneous parameters, requiring both new fluorochromes and corresponding laser wavelengths. A group of polymer dyes requiring deep ultraviolet (UV) excitation (~280–300 nm) is currently under development, allowing the expansion of high‐dimensional cytometry beyond the current 30 color limit. In this study, we evaluated a newly available laser emitting at 280 nm as a possible laser source for exciting these dyes. Since deep UV polymer dyes are not yet available, we used quantum nanoparticles (Qdots) as a surrogate probe to assess the utility of this laser wavelength for flow cytometry. Deep UV laser light was found to excite Qdots as well as traditional UV sources. Deep UV 280 nm did not excite BUV dyes well, suggesting that BUV and deep UV polymers will be spectrally compatible with low crossbeam spillover issues. Deep UV excitation did excite considerable autofluorescence in the violet to blue range, a limitation that will need to guide deep UV fluorochrome development. A deep UV 280 nm laser may therefore be the next essential wavelength for high‐dimensional flow cytometry. © 2018 International Society for Advancement of Cytometry

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Lasers for Flow Cytometry

Cited by 4 publications

References 1 publication

A flow cytometric method to measure prokaryotic records in ice cores: an example from the West Antarctic Ice Sheet Divide drilling site

A flow cytometric method to measure prokaryotic records in ice cores: an example from the West Antarctic Ice Sheet Divide drilling site

Deep ultraviolet lasers for flow cytometry

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