Time-Tagged Time Resolved Measurement modes of the HydraHarp 400

Time-Tagged Time-Resolved (TTTR) mode allows the recording of individual count events directly to hard disk or computer memory. The timing of each photon is captured as an event record without any early data reduction (such as on-board forming of histograms). This mode is particularly interesting where e.g. the dynamics in a fluorescence process are to be investigated in depth. The availability of the full timing information permits photon burst identification, which is of great value e.g. for Single Molecule Spectroscopy (SMS) in a liquid flow. Other typical applications are Fluorescence Correlation Spectroscopy (FCS) and Burst Integrated Fluorescence Lifetime (BIFL) measurements. Together with an appropriate scan controller, TTTR mode is also suitable for ultra fast Fluorescence Lifetime Imaging (FLIM) with unlimited image size. Applications beyond fluorescence spectroscopy are e.g. time-interval analysis, quantum optics and related basic rearch. The HydraHarp 400 actually supports two different Time-Tagging modes, T2 and T3 Mode - a concept originally introduced with the PicoHarp 300. They differ slightly in their use of the input channels and by using the suitable mode, a very wide range of applications can be covered.

T2 Mode

In T2 Mode all signal inputs of the HydraHarp 400 are functionally identical. There is no dedication of one channel to a sync signal. All inputs can be used to connect photon detectors. The events from all channels are recorded independently and treated equally. In each case an event record is generated that contains information about the channel it came from and the arrival time of the event with respect to the overall measurement start. If the time tag overflows, a special overflow marker record is inserted in the data stream, so that upon processing of the data stream a theoretically infinite time span can be recovered at full resolution. Dead times exist only within each channel (80 ns typ.) but not across the channels. Therefore, cross correlations can be calculated down to zero lag time. This allows powerful new applications such as FCS with lag times from picoseconds to hours to be implemented with one instrument. Autocorrelations can also be calculated at the full resolution but of course only starting from lag times larger than the deadtime.

T3 Mode

The T3 Mode is specifically designed to use periodic sync signals from pulsed lasers with high repetition rate up to 150 MHz. This signal is connected to the dedicated sync channel. As far as the experimental setup is concerned, this is similar to classic TCSPC in histogramming mode. In addition to the picosecond start-stop timing, the channel number is recorded and each event is time tagged with respect to the beginning of the experiment. The time tag is obtained by simply counting sync pulses. From the T3 Mode event records it is therefore possible to precisely determine which sync period a photon event belongs to. Since the sync period is also known precisely, this furthermore allows to reconstruct the arrival time of the photon with respect to the overall experiment time. If the counter overflows, a special overflow marker record is inserted in the data stream, so that upon processing of the data stream a theoretically infinite time span can be recovered.

External Event Markers

Both TTTR modes support capturing up to four external marker events that can be fed to the instrument as TTL signals via the front panel. These events are recorded as part of the TTTR data stream. This allows to precisely synchronize the TTTR measurement with almost any experiment. The most important applications of this feature are FLIM and FRET imaging. This concept is used in the cutting-edge time-resolved mircroscope MicroTime 200.

Software Support

The acquisition software provided with the instrument comes with a rich set of demo programs that enable users to write their own analysis and display programs for TTTR data. Users who prefer to use standard data analysis algorithms out of the box may want to consider the powerful SymPhoTime software suite. It implements a wide range of state-of-the-art analysis algorithms for FLIM, FCS and FRET to name only a few.

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