Introduction
Chlorophyll Fluorescence Imaging from Individual Cells to Plant Populations
Introduction

Chlorophyll fluorescence imaging device FluorCam is used to monitor fluorescence kinetics in pulse-amplitude modulated mode and saturation pulse method, which provides a wealth of information about a plant’s photosynthetic capacity, physiological and metabolic condition, as well as its susceptibility to various stress conditions. Chlorophyll fluorescence yield is estimated after the application of short saturating flash in dark-adapted plants or when plants are light-adapted or illuminated with photosynthetically active actinic light. The changes in chlorophyll fluorescence are used to describe plants capacities for photochemical and non-photochemical quenching of light energy supplied to plants surface.

FluorCam device consists of a CCD camera and different LED panels according to the type of the device:

  • Closed FluorCam FC 800-C/1010, FC 800-C/1010-GPF has fixed 4 to 5 LED panels (4 + 1 additional, which is not included in the standard setup) and, optionally, of a filter wheel equipped with up to 7 different emission filters.
  • Open FluorCam FC 800-O/1010, FC 800-O/2020 has 4 to 5 LED panels (4 + 1 additional, which is not included in the standard setup), when 1 pair of the LED panels can be optionally changed to LED panels with different wavelenght. Also a filter wheel equipped with up to 7 different emission filters can be optionally included.
  • Handy FluorCam FC 1000-H, FC 1000-H/GFP has 4 fixed LED panels and, optionally, 2 manually changeable emission filters.
  • Customized FluorCam versions consist of LED panel with 3 to 4 different LED wavelenghts panels (3 + 1 additional, which is not included in the standard setup) and, optionally, of a filter wheel equipped with up to 3 different emission filters.
  • Fluorescence Kinetic Microscope can be equipped with 10-LED light module or 3-LED light module.

For more information about each type of the FluorCam device, please refer to respective FluorCam chapters.


LED WAVELENGTHS OPTIONS


The available LED wavelengths are as follows:

Standard version


  • Red-orange 617 nm for measuring flashes and Actinic light 1
  • Cool white 6,500 K for saturating pulses and Actinic light 2

Other available wavelengths


  • Royal-blue 447 nm
  • Blue 470 nm
  • Cyan 505 nm
  • Green 530 nm
  • Amber 590 nm
  • Red 627 nm
  • Deep-red 660 nm

Additional lights


  • UV 385 nm
  • Green 530 nm
  • Amber 590 nm
  • Deep-red 660 nm
  • FAR red 740 nm

GFP measurement


  • Royal-blue 447 nm
  • Blue 470 nm
  • The GFPCam consists of a CCD camera, four to five fixed LED panels (4 + 1 additional, which is not included in the standard setup) and of a filter wheel equipped with up to 7 different emission filters. To be used in all FluorCam configurations.

NDVI measurement


  • Deep-red 660 nm
  • FAR 740 nm
  • High-end option for measuring PAR absorptivity and NDVI reflectance index. It includes 7-position filter wheel, light panel 13 × 13 cm (installed round the camera and mounted with FAR and deep-red LEDs) and a glass filter. PAR-measurement is based on reflectance of plants in near infrared region and in the visible red range of spectrum. NDVI is used as vegetative index for predicting photosynthetic activity. To be used only with FC 800-C/1010, FC 800-C/1010-GPF , FC 800-O/1010 or FC 800-O/2020.

Multicolor fluorescence measurement


  • UV 385 nm
  • Multicolor Plant Module is High-end option for measuring emission spectra in plants (according to Lichtenthaler and Bushmann). It includes 7-position filter wheel, UV LED light panel (installed round the camera and mounted with 4 UV LEDs), four filters - BLUE (FF01-440/40), GREEN (FF02-520/28), RED (FF01-690/8), FAR-RED (FF01-747/33) and a special software module.
  • To be used only in the FC 800-C/1010, FC 800-C/1010-GPF, FC 800-O/1010 or FC 800-O/2020. This module cannot be used with the PAR-Absorptivity and NDVI Module in the FC 800-C/1010 or FC 800-C/1010-GPF.

Please note, that not all of the offered LED wavelengths are available for all FluorCam types. For more detailed information please refer to respective FluorCam chapters.



EMISSION FILTERS

All the FluorCam devices are standardly equiped by chlorophyll fluorescence emission filter. For different applications, such as fluorescence proteins detection or multicolor fluorescence detection, the 7-position filterwheel with different filters can be used:

Multicolor fluorescence measurement


  • Blue 440/40
  • Green 520/23
  • Red 690/8
  • IR 747/33

Fluorescence proteins detection



CCD CAMERAS


Three CCD cameras are available, high sensitivity CCD camera TOMI-1, high resolution CCD camera TOMI-2 and high speed CMOS camera TOMI-3.

Please note, that not all of the offered CCD cameras are available for all FluorCam types. For more detailed information please refer to respective FluorCam chapters.


TOMI-2

High resolution CCD camera, which is intended for combined measurement of ChlF and detection of weak steady-state fluorescence signals (such as GFP) with long integration times or applications where the higher spatial resolution is of highest importance (microscopy).

  • Resolution: 1360 × 1024 pixels
  • Binning option 2 × 2: 680 × 512 pixels
  • A/D converter resolution: 16 bit (65 536 grey levels)
  • Pixel size: 6.45 µm × 6.45 µm
  • 20 frames per second for both resolution options
  • Operating temperature: 0–50 °C
  • Operating humidity: 0–95 %
  • Interface connector: Gigabit Ethernet
  • Operating modes: video (ChlF kinetics measurement), snapshot (long integration time for FPs detection)
  • CCD Detector Wavelength Range: 400–1000 nm
  • Spectral Response: QE max at 540 nm (~72 %), 50 % roll-off at 350 nm and 800 nm
  • Read-Out Noise: Less than 8 electrons RMS - typically only 6 electrons
  • Full-Well Capacity: Greater than 22 000 electrons (unbinned)

TOMI-3

Intended for high speed fluorescence imaging, fast fluorescence kinetics, OJIP protocol and other applications where high speed imaging is beneficial

  • High-speed camera
  • Global shutter CMOS, 1.3 M pixel sensor
  • A/D: 12 bit (4096 grey levels)
  • 6.6 µm x 6.6 µm pixel size
  • Frame rate:
  • - 1280 x 1024 pixels; 1000 images per second
    - 640 x 512 pixels; 4000 images per second
    - 640 x 256 pixels; 8000 images per second
    - 640 x 128 pixels; 16000 images per second
    - Faster frame rates with smaller resolution

  • Binning option: 2x2 - 640 x 512 pixels, 4000 images per second
  • Operating Temperature 0 to +50 °C
  • Operating Humidity 0- 90% (non-condensing)
  • Interface connector: Gigabit Ethernet
  • CMOS detector 400-1000nm
  • Spectral Response: QE max 50% at 550nm
  • Read-Out Noise: Less than 25 electrons RMS
  • Full-Well Capacity: Greater than 17 000 electrons (unbinned)