Which Windows version is the FluorCam 7 software compatible with?

It is compatible with Windows XP, Windows Vista, and Windows 7 (32 bit versions only).

Which LED panels are mounted in the standard FluorCam?

There must be two pairs of panels - each pair of the same color. The standard setup is: two red-orange panels 618 nm providing Measuring Light and Actinic Light 1. The second pair generates Actinic Light 2 and Saturating Pulse. These panels may be either white or blue 455 nm or red-orange 618 nm.

How should I make choice between white, blue or red color for the second pair of panels (Actinic Light 2 and Saturating Pulse)?

Blue actinic light is often used in the systems where GFP fluorescence is detected together with Chlorophyll fluorescence and blue LED panel (450nm) is used as excitation light. For chlorophyll fluorescence kinetic measurements both blue and white panels can be used. In case of blue panel one wavelength light source is used to active photochemistry (450+/- 10 nm), in case of white panel, the spectrum is covering large proportion of the PAR range. Thus it is up to your decision which of the light source would suite more to address your research questions.

Can you also add a UV panel to my FluorCam?

Yes, we can supply an additional UV panel that is mounted around the camera. It is constructed with 4 LEDs, their wavelength is 365 nm or 385 nm, according to the customer's preference.

What is the meaning of Fv/Fm, Kautsky, Quenching, Dyes and Fps?

Fv/Fm is a short protocol giving you Fo, Fm, and maximum quantum yield of Photosystem II = Fv/Fm. Kautsky kinetics is a fluorescent transient measured in continuous actinic light – so called Kautsky effect. Quenching analysis is a complex protocol giving full information of dark adapted as well as light adapted state of the plant. There exist some good reviews e.g. Maxwell, 2000 - http://jxb.oxfordjournals.org/content/51/345/659.full.pdf. Dyes and FPs is a special protocol for measuring GFP and other FPs.

What is usual value of Fv/Fm in a healthy plant?

Fv/Fm of a healthy plant is 0.83. If it is lower, the plant might be stressed. However, be aware of correct measurement of Fv/Fm. The saturating light must saturate photosyntesis and measuring light must be weak enough to determine correct Fo without actinic effect.

Is there any possibility to assess the chlorophyll concentration using the FluorCam?

Yes, this is possible with the PAR Absorbance Module (includes the filter-wheel, quartz filter and additional "PAR" light panel with 670nm and FAR LEDs. Using this module, you will be able to measure also Chl concentration heterogeneity. Easier and cheaper solution might be if you use the PSI's PlantPen https://handheld.psi.cz/products/plantpen-ndvi-and-pri/ (no heterogeneity comparison).

Using data export in the FluorCam, is it better to use "frame numeric" or "numeric average"?

In general, we would always recommend to use numeric average. FvFm measurement can serve as an example for explanation. If frame numeric is used, you use pixel by pixel fluorescence values to calculate FvFm and then you average the FvFm value over the area that you selected, means over all pixels selected. In case of numeric average, you take Fo value over all area selected (all pixels selected) and Fm value over all area selected and use this F0 and Fm value to calculate FvFm. If you analyse object with strong signal and you do not integrate any noise, you will not see any difference between the two results. If you have weaker signal and you have some noise in your measurement, the average numeric will be always more powerful with less noisy pixels included.

Sometime my fluorescence data from the FluorCam show "pixel overflow", what does it mean? How does it affect my data interpretation? How could I fix it?

Range of fluorescence units that can be detected by the camera is 0-4096 units. Any signal exceeding the maximum CCD well capacity (4096 levels) is indicated by the warning pixels overflow. Always prior initiating new experiment you need to adjust the settings in live window or protocol window as shutter, sensitivity and light intensity as such that when the given light used with the given settings (shutter and sensitivity) is for your imaged plant material giving fluorescence signal in 1/3 of the CCD well capacity meaning about 1500 units. It also must be taken into account that for different plants (for example less chlorophyll vs dark plants) different setting might be required and will have different response.

How does "electronic shutter" and "sensitivity" affect the imaging quality? Should I modify it for each condition? What combined settings do you recommend for best result?

Please refer to the manual provided and page 41, where you can find description how to define correct settings for measurements. In short: the shutter opening determines the duration of the interval during which the electronic shutter is opened during measurements. It controls also the duration of the measuring flashes and that cannot be longer than 20 or 30 ms. The apparently brighter measuring light with longer shutter opening time is due to the longer on-time rather than to an increased photon flux. The brighter fluorescence signal with longer shutter opening is due to longer integration in the CCD chip. The higher signal and the better signal-to-noise ratio with longer shutter opening time is unfortunately traded by increased actinic effects of the measuring light. The measured signal deviates from the true Fo if the measuring light is too strong. The problem can be alleviated by low repetition frequency of the measuring flashes. The sensitivity scale is linear and relative (0-100%): 0 meaning minimum sensitivity and 100% meaning maximum sensitivity. The maximum sensitivity range (80-100%) should be used only when necessary, because it can lead to a relatively low signal-to-noise ratio. Optimal setting for the sensitivity would be in range 0-50%.

We are using the FluorCam system for our measurements very successfully. Is there a way to export the fluorescence/time data from a Kautsky measurement beside the preset parameters (Fo, Fm, QY etc.)? The data is shown for each experiment in the software (FluorCam 1.0.6.14) very nicely. But we need to compare multiple data sets from different experiments.

There are two ways how to do it: (1) To export the raw data (fluorescence transient - graph), please go to: EXPERIMENT (top panel) -> Export -> Kinetic; (2) To export calculated parameters (Fo, Fv/Fm...): go to EXPERIMENT (top panel) -> Export -> Numeric. In both cases, it is recommended to select “All data“. For Numeric data: Frames numeric – are calculated for each pixel (e.g. Fv/Fm for each pixel) and then averaged over selected area. Averages numeric – first absolute parameters such as Fo, Fm, Fp... are calculated over selected area and then these values are used to calculate more advance parameters e.g Fv/Fm = (Fm-Fo)/Fm. The Frames numeric can give different (non-sense) values in the case of weak fluorescence signal with low signal/noise ratio. The data will be exported to txt file which can be easily imported to for example Excel.

How can I measure light intensity in the infrared LED Light Panel, which I use with my FluorCam?

For wavelengths above 700 nm, is not possible to use standard PAR light meter and measure light intensity in µmol(photon).m-2.s-1. Instead light can be measured for its radiated power (output power is given in milliwatts).

I am considering purchase of your GFP-Cam, closed version. Do I need to order a High-Resolution Camera as a separate item?

No, the High-Resolution Camera and motorized filter wheel with filters for GFP and Chl expression are included in the price of the GFP-Cam system.

Is it possible to use the FluorCam or GFP-FluorCam to detect Luciferase signal?

Yes, it is possible to detect Luciferase but the standard FluorCam instrument (FC 800-C, FC 800-C-GFP) is not needed for this purpose. In principle, no LED panels and filters are needed because Luciferase emits light based on chemical reaction in contrast to fluorescence emission. On the other hand, a very sensitive cooled CCD camera is needed for Luciferase detection (this camera is much more expensive than the camera used for fluorescence imaging). To sum up, PSI is able to deliver the system for Luciferase detection. It would consist of a mechanical construction used for the FC 800-C, 1 LED accessory panel - allowing easy sample manipulation under the camera, cooled CCD camera and modified software.

Where can I check FluorCam program compatibility with different Windows versions?

It can be checked at our web page - download.

How can I use the advanced Multiple function in the FluorCam software?

The Advanced Multiple function enables combination of two protocols and thus it is useful for circadian cycle study: one protocol runs at day time, the second protocol runs at night. This protocol is a paid one and must be purchased as an optional feature. The key to this protocol is then copied on delivered computer, directory: C:\Install...\Install CD\User Keys and on delivered installation flash disk, directory ...\User Keys. If the protocol is activated, you will see „double red flash“ icon in main tool bar of the FluorCam software.