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- intensity_time_trace_analysis
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- recording_a_fluorescence_lifetime_image_flim_stack_with_a_lsm_upgrade_kit_on_a_nikon_a1
- symphotime_tips_and_tricks
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- lifetime_fitting_using_the_flim_analysis
- hows step-by-step, how the FLIM analysis of SymPhoTime 64 can be used to perform a lifetime fit and how to extract and read the results. In d... s on the right of the plot.\\ **Lower left:** Lifetime fitting parameters. Here lifetime fitting models can be defined. Fitting is then ap... gging the limit bars in the color scale "Fast Lifetime" histogram or by considering the peaks of the "Lifetime Histogram". Entering fixed values has the advanta... now the calculated intensity weighted average lifetime "tau_Av_Int" is displayed, in contrast to the "Fast Lifetime", which is just the mean arrival time and contain
- flim_fret_calculation_for_multi_exponential_donors
- portant to use the amplitude weighted average lifetime. While the intensity weighted average lifetime corresponds to the real average arrival time of the photons and is therefore also close to the Fast lifetime in case of low background noise, the amplitude weighted average lifetime reflects the quenching of the donor due to the FR... nential_donors_Image_3.png }} * Start the "Lifetime FRET Image" script by clicking on "Start". {{ fli... _donors_Image_25.png }} **Response:**\\ The “Lifetime FRET Image” script is applied to the file ''CENP-
- lifetime-fitting_using_the_rapid_reconvolution_algorithm
- tains now the fitted, intensity weighted mean lifetime, instead of the "fast Lifetime", based on the photon arrival time only. The Histogram on the right of the image con... d distribution of the intensity-weighted mean lifetime. The distributions for lifetime component 1 and lifetime component 2 appear as bars, as these parameters h... the result image after the fit with the fast lifetime image above, you will see that the lifetime distribution of the lifetime within a single bead is much more uniform, as eff... orrected for.\\ * If you want to export the lifetime distribution, open the context menu via right mou
- intensity_time_trace_analysis
- {{tag> tutorial howto time-trace analysis SymPhoTime}} ~~NOTOC~~ ======Intensity Time Trace Analysis====== ===== Summary =====... start the script ==== * Start [[products:SymPhoTime64|SymPhoTime 64]] software. * Open the "Samples" workspace v... ript_Image_4.png }} **Response:** The "Intensity Time Trace" script is applied to the file ''Cy5_immo_Lifetime_Trace.ptu''. Thereby, a new Window opens: {{ usin... Plotting options. - **Center/right:** Intensity time trace. As no time traces have been calculated yet, this area is sti
- lifetime-fitting_using_the_flim_script
- {{tag>howto tutorial FLIM analysis SymPhoTime MicroTime LSM_upgrade}} ====== Lifetime-Fitting Using the FLIM Analysis (updated for SymPhoTime V 2.5 and above) ====== ===== Introduction ===== This tutorial shows step-by-step, how the FLIM script of SymPhoTime 64 (V2.5) can be used to perform a pixel-by-pixel lifetime fit of a FLIM image and how to extract and read t... \\ **4: Decay Fitting Parameters**\\ Here the lifetime fitting model can be selected. Fitting is then applied to the lifetime graph(s) on the lower center ([[glossary:TCSPC]]
- lifetime_fitting_using_the_tcpsc_fitting_script
- {{tag>howto analysis lifetime tutorial SymPhoTime}} ~~NOTOC~~ ====== SymPhoTime Lifetime Fitting ====== ===== Summary ===== This tutorial shows step-by-ste... and start the script * Start [[products:SymPhoTime64|SymPhoTime 64]] software. * Open the "Samples" workspace v... SPC histogram it can be clearly seen that the lifetime of this dye is significantly shorter than the lifetime of ATTO655. {{ lifetime_fitting_using_the_tcpsc
- flim-fret_calculation_for_single_exponential_donors
- This tutorial shows step-by-step, how the "Lifetime FRET Image" analysis of [[products:SymPhoTime64|SymPhoTime 64]] can be used to calculate pixel-by-pixel the ... ce of FRET (τ<sub>D</sub>), while the shorter lifetime (τ<sub>DA</sub>) is varied. This lifetime is considered to be the donor lifetime with a FRET acceptor present. The FRET efficiency... ws a single peak, which is in contrast to the lifetime variations in the FLIM image. The explanation can... ram plots the amplitude fraction of the donor lifetime in presence of FRET. As the amplitude of a donor
- visualizing_dynamics_using_the_multiframe-flim_script
- \\ **4: Decay Fitting Parameters**\\ Here the lifetime fitting model can be selected. Fitting is then applied to the lifetime graph(s) on the lower center ([[glossary:TCSPC]] ... cale by typing the limit values for the "Fast Lifetime [ns]" and "Events[cnts]". Additionally set the "Lifetime Histogram" range on the right hand.\\ {{ howto:f... whether you have the scalebar active in the SymPhoTime. When measuring with a scanner controlled by SymPhoTime, e.g. a MicroTime system, or you transfer the image size informatio... . If your scanner is not controlled via the SymPhoTime, and no handshake transfers the correct image siz
- roi_fitting_using_the_flim_script
- [[glossary:flim|FLIM]] script of [[products:SymPhoTime64|SymPhoTime 64]] can be used to fit several regions of intere... start the script ==== * Start [[products:SymPhoTime64|SymPhoTime 64]] software. * Open the "Samples" workspace v... the right of the plot.\\ - **Lower left:** Lifetime fitting parameters. Here lifetime fitting models can be defined. Fitting is then ap... gging the limit bars in the color scale "Fast Lifetime" histogram or by considering the peaks of the "Lifetime Histogram". Entering fixed values has the advanta
- update
- \\ **4: Decay Fitting Parameters**\\ Here the lifetime fitting model can be selected. Fitting is then applied to the lifetime graph(s) on the lower center ([[glossary:TCSPC]] ... cale by typing the limit values for the "Fast Lifetime [ns]" and "Events[cnts]". Additionally set the "Lifetime Histogram" range on the right hand. * Tick the ... p, as an example, we mark an ROI with similar lifetime range using the "Magic Wand" tool. The sensitivity of the tool to lifetime differences can be adjusted in a box below the in... \\ The parts of the image with a similar fast lifetime range will be selected and the lifetime histogram will get updated accordingly. If you wa
- visualizing_dynamics_with_the_multi_frame_flim_analysis
- re (i.e. the piezo scanner in the [[products:MicroTime#MicroTime 200|MicroTime200]]), the image dimensions are automatically cor... ded, a procedure according to the tutorial: [[Lifetime fitting using the FLIM Analysis]] must be applied... 17 images have to be analyzed, which can be quite time consuming. Therefore, it can be helpful to limit
- burst_duration_and_fcs_diffusion_time @howto:burst_duration_and_fcs_diffusion_time
- = 4.2 Physical interpretation ==== The diffusion time $t_D$ is not the mean residence time of an individual molecule. Instead, it is a corre... entally different observables. The FCS diffusion time $t_D$ is a correlation-derived geometry time scale governed by quadratic spatial weighting of ... relation. Flow introduces an additional transport time scale that leaves the FCS diffusion time unchanged when modeled explicitly, while directly... ely confined to slow non-stationarity. Diffusion time and burst duration should therefore be regarded a
- symphotime_tips_and_tricks
- time) in the TCSPC fitting panel * the fast lifetime in an ideal system should equal the intensity weighted average lifetime [τ av. int.]. Usually, fitted lifetimes tend to b... ', and also the smoothing does not affect the lifetime histogram * Take care that you record the data... d.png?200 |}} marks areas with a similar fast lifetime ==== Undo ROI selection ==== Using ''<CTRL>+<Z... Example: marking several spots with a similar lifetime: ==== |1. Click on the file ''FAST_FLIM.pqres'',
- separation_of_2_species_with
- start the script ==== * Start [[products:SymPhoTime64|SymPhoTime 64]] software. * Open the "Samples" workspace v... dye is analyzed that changes its fluorescence lifetime under certain conditions, a lifetime fit can be performed of the TCPSC histogram of th... LCS filters can be calculated from the fitted lifetime components. This approach called "LT (lifetime) fitting" can only be employed, if the two compon... graph. These filter curves correspond to the lifetime component of ~0.8 ns (Cy5), the lifetime component of ~1.8 ns (ATTO655) and the background
- calculate_ratiometric_single_pair_fret_distributions_using_the_pie-fret_script
- {{tag> tutorial howto FRET PIE analysis time-trace SymPhoTime}} ~~NOTOC~~ ====== Calculate Ratiometric Single Pair FRET ... tart the script ==== * Start [[products:SymPhoTime64|SymPhoTime 64]] software. * Open the "Samples" workspace v... ** Fitting and analysis options\\ - **Center:** Time traces. As the time traces are not yet calculated, this graph is stil... , calculate the FCS curve and check the diffusion time to see the average residence time of the sample in the laser focus. {{ calculate_r