Compact TCSPC upgrade package for laser scanning microscopes based on 375- to 470-nm picosecond diode lasers

Ortmann, Uwe; Dertinger, Thomas; Wahl, Michael; Patting, Matthias; Erdmann, Rainer

doi:10.1117/12.536888

Cited by 9 publications

(3 citation statements)

References 4 publications

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“…The data are nearly free of redundancy and can therefore be transferred in real time, even if the scan speed is very fast, like, e.g., in laser scanning microscopes ͑LSM͒. 16 The image size is unlimited both in size and in count depth.…”

Section: Discussionmentioning

confidence: 99%

Dead-time optimized time-correlated photon counting instrument with synchronized, independent timing channels

Wahl

Rahn²,

Gregor³

et al. 2007

Review of Scientific Instruments

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Time-correlated single photon counting is a powerful method for sensitive time-resolved fluorescence measurements down to the single molecule level. The method is based on the precisely timed registration of single photons of a fluorescence signal. Historically, its primary goal was the determination of fluorescence lifetimes upon optical excitation by a short light pulse. This goal is still important today and therefore has a strong influence on instrument design. However, modifications and extensions of the early designs allow for the recovery of much more information from the detected photons and enable entirely new applications. Here, we present a new instrument that captures single photon events on multiple synchronized channels with picosecond resolution and over virtually unlimited time spans. This is achieved by means of crystal-locked time digitizers with high resolution and very short dead time. Subsequent event processing in programmable logic permits classical histogramming as well as time tagging of individual photons and their streaming to the host computer. Through the latter, any algorithms and methods for the analysis of fluorescence dynamics can be implemented either in real time or offline. Instrument test results from single molecule applications will be presented.

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Section: Discussionmentioning

confidence: 99%

Dead-time optimized time-correlated photon counting instrument with synchronized, independent timing channels

Wahl

Rahn²,

Gregor³

et al. 2007

Review of Scientific Instruments

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“…Basic requirement is a confocal microscope setup capable of conventional FCS measurement. There are several commercial products directly supporting FCS; most contemporary confocal laser scanning microscopes can also be upgraded for F(L)CS capability [ 34 ]. At least one picosecond pulsed laser source is necessary.…”

Section: Frequently Asked Questions About Flcsmentioning

confidence: 99%

Fluorescence Lifetime Correlation Spectroscopy (FLCS): Concepts, Applications and Outlook

Kapusta

Macháň

Benda

et al. 2012

IJMS

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Fluorescence Lifetime Correlation Spectroscopy (FLCS) is a variant of fluorescence correlation spectroscopy (FCS), which uses differences in fluorescence intensity decays to separate contributions of different fluorophore populations to FCS signal. Besides which, FLCS is a powerful tool to improve quality of FCS data by removing noise and distortion caused by scattered excitation light, detector thermal noise and detector after pulsing. We are providing an overview of, to our knowledge, all published applications of FLCS. Although these are not numerous so far, they illustrate possibilities for the technique and the research topics in which FLCS has the potential to become widespread. Furthermore, we are addressing some questions which may be asked by a beginner user of FLCS. The last part of the text reviews other techniques closely related to FLCS. The generalization of the idea of FLCS paves the way for further promising application of the principle of statistical filtering of signals. Specifically, the idea of fluorescence spectral correlation spectroscopy is here outlined.

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“…The data are nearly free of redundancy and can therefore be transferred in real time, even if the scan speed is very fast, e.g., in laser scanning microscopes. 25 In the original TTTR approach, the different time scales are processed and used independently. However, often it is of great interest to obtain high resolution timing on the overall scale, i.e., combining coarse and fine timing into one global arrival time per event, with picosecond resolution.…”

Section: Introductionmentioning

confidence: 99%

Scalable time-correlated photon counting system with multiple independent input channels

Wahl

Rahn

Röhlicke

et al. 2008

Review of Scientific Instruments

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Time-correlated single photon counting continues to gain importance in a wide range of applications. Most prominently, it is used for time-resolved fluorescence measurements with sensitivity down to the single molecule level. While the primary goal of the method used to be the determination of fluorescence lifetimes upon optical excitation by short light pulses, recent modifications and refinements of instrumentation and methodology allow for the recovery of much more information from the detected photons, and enable entirely new applications. This is achieved most successfully by continuously recording individually detected photons with their arrival time and detection channel information (time tagging), thus avoiding premature data reduction and concomitant loss of information. An important property of the instrumentation used is the number of detection channels and the way they interrelate. Here we present a new instrument architecture that allows scalability in terms of the number of input channels while all channels are synchronized to picoseconds of relative timing and yet operate independent of each other. This is achieved by means of a modular design with independent crystal-locked time digitizers and a central processing unit for sorting and processing of the timing data. The modules communicate through high speed serial links supporting the full throughput rate of the time digitizers. Event processing is implemented in programmable logic, permitting classical histogramming, as well as time tagging of individual photons and their temporally ordered streaming to the host computer. Based on the time-ordered event data, any algorithms and methods for the analysis of fluorescence dynamics can be implemented not only in postprocessing but also in real time. Results from recently emerging single molecule applications are presented to demonstrate the capabilities of the instrument.Scalable time-correlated photon counting system with multiple independent input channels Time-correlated single photon counting continues to gain importance in a wide range of applications. Most prominently, it is used for time-resolved fluorescence measurements with sensitivity down to the single molecule level. While the primary goal of the method used to be the determination of fluorescence lifetimes upon optical excitation by short light pulses, recent modifications and refinements of instrumentation and methodology allow for the recovery of much more information from the detected photons, and enable entirely new applications. This is achieved most successfully by continuously recording individually detected photons with their arrival time and detection channel information ͑time tagging͒, thus avoiding premature data reduction and concomitant loss of information. An important property of the instrumentation used is the number of detection channels and the way they interrelate. Here we present a new instrument architecture that allows scalability in terms of the number of input channels while all channels are synchronized ...

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Compact TCSPC upgrade package for laser scanning microscopes based on 375- to 470-nm picosecond diode lasers

Cited by 9 publications

References 4 publications

Dead-time optimized time-correlated photon counting instrument with synchronized, independent timing channels

Dead-time optimized time-correlated photon counting instrument with synchronized, independent timing channels

Fluorescence Lifetime Correlation Spectroscopy (FLCS): Concepts, Applications and Outlook

Scalable time-correlated photon counting system with multiple independent input channels

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