I need to image cyanobacteria using fluorescent microscopy, I have easily done this before with good results using the fluorescence of chlorophyl on a traditional fluorescent microscope. However, when I attempted to do this on a confocal system I had very poor results when imaging Cy5.5 along side the chlorophyl.
What excitation and emission wavelengths should I use for chlorophyl, specifically for cyanobacteria?
Should I use a different second dye like NBD or rhodamine instead of Cy5.5?
It seems that it would depend on the species of cyanobacteria.
Article Optimization of variable fluorescence measurements of phytop...
I did the following (CLSM): Autofluorescence of green
algae (chlorophyll a) and cyanobacteria (chlorophyll a and
phycobiliproteins) were excited by beams of 555 and 639 nm
wavelengths respectively. Using two laser beams made it
possible to differentiate between green algae and cyanobacteria:
superimposed images were generated by the outputs
of two channels. Emitted wavelengths were collected using
a band-pass filter 530/30 and cyanobacterial natural fluorescence
with a 590 nm long pass filter.
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