Determination of photosynthetic pigment composition in an individual phytoplankton cell in seas and lakes using fluorescence microscopy; properties of the fluorescence emitted from picophytoplankton cells

To develop a method for the determination of photosynthetic pigment species in individual phytoplankton cells, especially natural picophytoplankton cells, the fluorescence spectra of intact cells were studied with cultured phytoplankton. The study was made mainly with phycoerythrin-containing picophytoplankton collected off Japan in 1982 with reference to diatomal species and phycoerythrin-free cyanophycean species. The spectra were measured for cell suspensions with an ordinary spectrofluorometer, and for individual cells with a microscope spectrofluorometer, paying special attention to the effect of cell-fixation. Results indicated that: (1) the cell-fixation with the glutaraldehyde and paraformaldehyde mixture modified phycoerythrin emission from picophytoplankton markedly in its wavelength location and intensity, but (2) the emission from phycocyanin was affected far less, and (3) the emission from chlorophyll a was not altered. However, the phycoerythrin emission modified by the fixation was found to be easily distinguished from other emissions, and kept its intensity high enough for detection with a fluorescence microscope. The fluorescence properties after the fixation were kept unaltered for a long period of time. Based on the results, we propose a simple method for the determination of photosynthetic pigments in individual phytoplankton cells in seas and lakes using fluorescence microscopy. Results of our tests with natural samples of phytoplankton are presented, and problems for further improvement are also discussed.