Chlorophyll Fluorescence Imaging and PAM Fluorometry
The HEXAGON-IMAGING-PAM represents the largest Walz Imaging System available. Despite the large measuring area, the instrument is still flexible enough to measure a wide variety of samples. These include individually potted plants, seedlings in larger plant trays, or harvested samples of higher or lower plants.
It can measure areas of 20 x 24 cm with a resolution of up to 1000 x 1200 px (2x2 binning), according to the PAM principle with highest accuracy.
In contrast to the significantly smaller MAXI Imaging System, the HEXAGON-IMAGING-PAM offers 4 times the measuring area of the smaller MAXI-IMAGING-PAM and up to 1.2 MP image resolution (binning switched off).
This high resolution results in a pixel size of 100 x 100 µm.
During the design process, we placed special emphasis on the shadow-free, uniform illumination of the measuring area and designed the cooling of the panels very efficiently for a long service life of the LEDs.
The HEXAGON-IMAGING-PAM can now also be used to determine the Fo' value of the samples under investigation. For this purpose, the instrument has an additional, dual-circuit far-red LED panel with which FR light and also FR light with a higher red component can be very finely dosed.
The name HEXAGON-IMAGING-PAM is derived from the shape of the individual panels that make up the main panel. The chosen shape offers the best possibility to achieve the ideal illumination of the sample area. Smaller hexagon shaped panel sub-units allow better compensation of LED imbalances to achieve even more reproducible results.
In the HEXAGON-IMAGING-PAM, the control electronics are integrated into the measuring head so that, despite the large measuring surface, a quite compact device is available.
Outstanding Properties of the HEXAGON-IMAGING-PAM
- Homogeneous illumination of measured area
- Image resolution up to 1,2 MP (2x2 pixel binning)
- Large variety of possible sample morphologies
Chlorophyll Fluorescence and PAM Fluorometry
Chlorophyll fluorescence is a very sensitive indicator of photosynthesis. Quantitative information on the quantum yield of photosynthetic energy conversion is obtained by PAM fluorometry and the saturation pulse method (Schreiber U (2004) Pulse-Amplitude-Modulation (PAM) Fluorometry and Saturation Pulse Method: An Overview, pp. 279-319. Kluwer Academic Publishers, Dordrecht, The Netherlands).
A wide range of photosynthetic parameters can be derived from fluorescence measurements, giving insight into the physiological state of all photosynthetically active organisms, including higher plants, mosses and ferns as well as various types of algae, phytoplankton and biofilms.
Chlorophyll Fluorescence Imaging
With the advance of highly sensitive CCD cameras, followed by modern CMOS type camera chips, together with extremely strong light emitting diodes (LED), development of IMAGING-PAM fluorometers has become possible that not only measure images of chlorophyll fluorescence but are also fully competent in providing all relevant chlorophyll fluorescence parameters, using the saturation pulse method. In this way, images of photosynthetic activity and its spatio-temporal variations can be obtained.
In recent years, the trend has increasingly been towards very low-noise CMOS cameras offering high sensitivities. LED technology is also making continuous progress so that high irradiation intensities can be achieved with lower power.
An often-neglected problem of LEDs is their heating, which leads to a change in their performance. With the new HEXAGON-IMAGING-PAM, we have therefore focused in particular on compensating for these power changes and thus opening up new dimensions of measurement accuracy in imaging and illumination homogeneity. This also opens the door for new measurement methods to be integrated into the software.
All IMAGING-PAM fluorometers provide images for 17 different parameters. The fluorescence parameter Ft is continuously monitored and kinetic values can also be exported with 150 ms clock speed. Fo and Fm are assessed after dark adaptation, serving as reference for fluorescence quenching analysis by the saturation pulse method.
Besides Fv/Fm, the PS II quantum yield after dark acclimation, also the PS II quantum yield during illumination, Y(II), and the quantum yields of regulated and non-regulated energy dissipation, Y(NPQ), Y(NO) as well as the apparent electron transport rate (ETR) can be imaged.
General Features HEXAGON-IMAGING-PAM
In the development of the HEXAGON-IMAGING-PAM, we have followed the wishes of our customers and have, in addition to an enlarged surface, also revised the LED control to create space in the technical specifications for new applications. The housing was also adapted to the versatile applications of our customers and allows the measurement of leaves, algae in vessels and benthic organisms but also whole, potted plants or plant trays without long conversion times.
Safety has also been taken into consideration. The strong light pulses with which PAM instruments work should not harm the eyes of the experimenter. Therefore, we have integrated a safety shutdown into the HEXAGON-IMAGING-PAM, which suppresses all strong light intensities if the door is opened during an experiment.
After starting the software, the HEXAGON-IMAGING-PAM works in 2x2 pixel binning mode on an area of 20 x 24 cm. However, a special zoom function allows the user to zoom to a freely selectable quarter of the measuring area without changing the image resolution. Thus, without loss of sensitivity, it is very easy to work with a higher magnification on a smaller measuring area.
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