355
It is important to note that the amount of light scattered or emitted as fluorescence by an
individual cell is very small, so signal sizes must be maximized by making the exciting light
beam very intense. This is typically done by focussing the beam to a very small spot,
sometimes only a few-fold larger than the cells of interest. This in turn necessitates confining
the sample flow stream to a very small diameter in the center of the flow channel and places
an upper limit on the sample volume that can be analyzed in a given time. This is a significant
constraint for many of the applications we will discuss.
APPLICATIONS TO ECOLOGY
Experimental Studies
Cell Cycle Analyses: One of the first applications of flow cytometry to the study of marine
organisms was a direct outgrowth of biomedical studies of the cell cycle: DNA distributions
in cultures of phytoplankton grown under different conditions were measured to investigate
the influence of environmental factors on the regulation of cell growth and division (e.g.
Chisholm et al. 1986; Olson et al. 1986; Vaulot et al. 1986). For this work it proved
necessary to remove the phytoplankters' autofluorescent pigments to get good signals from the
fluorescent DNA stains used. This was done either by photo-oxidation (Yentsch et al. 1983)
or by methanol treatment (Olson et al. 1983), making the phytoplankters quite analogous to
the non-pigmented cells for which the techniques had been designed. These laboratory studies
have improved our understanding of cell cycle regulation in different phytoplankton groups
but they did not initially lend themselves to field applications. Only through the use of
microspectrofluorometry (Chang and Carpenter 1988) could de-chlorophyllized cells belonging
to different species be distinguished from one another well enough to resolve the DNA
histograms in field samples (Chang 1989).
Recently, however, Boucher et al. (in press) have shown that when natural populations are
stained with DAPI, DNA histograms from the picoplankton can be resolved without extracting
the chlorophyll from the cells. Prochlorophytes in particular could be recognized from their
flow cytometric signatures and their cell cycle position could be determined. Moreover, the
distributions of DNA per cell reveal relative growth rates for cells growing under different
It is important to note that the amount of light scattered or emitted as fluorescence by an
individual cell is very small, so signal sizes must be maximized by making the exciting light
beam very intense. This is typically done by focussing the beam to a very small spot,
sometimes only a few-fold larger than the cells of interest. This in turn necessitates confining
the sample flow stream to a very small diameter in the center of the flow channel and places
an upper limit on the sample volume that can be analyzed in a given time. This is a significant
constraint for many of the applications we will discuss.
APPLICATIONS TO ECOLOGY
Experimental Studies
Cell Cycle Analyses: One of the first applications of flow cytometry to the study of marine
organisms was a direct outgrowth of biomedical studies of the cell cycle: DNA distributions
in cultures of phytoplankton grown under different conditions were measured to investigate
the influence of environmental factors on the regulation of cell growth and division (e.g.
Chisholm et al. 1986; Olson et al. 1986; Vaulot et al. 1986). For this work it proved
necessary to remove the phytoplankters' autofluorescent pigments to get good signals from the
fluorescent DNA stains used. This was done either by photo-oxidation (Yentsch et al. 1983)
or by methanol treatment (Olson et al. 1983), making the phytoplankters quite analogous to
the non-pigmented cells for which the techniques had been designed. These laboratory studies
have improved our understanding of cell cycle regulation in different phytoplankton groups
but they did not initially lend themselves to field applications. Only through the use of
microspectrofluorometry (Chang and Carpenter 1988) could de-chlorophyllized cells belonging
to different species be distinguished from one another well enough to resolve the DNA
histograms in field samples (Chang 1989).
Recently, however, Boucher et al. (in press) have shown that when natural populations are
stained with DAPI, DNA histograms from the picoplankton can be resolved without extracting
the chlorophyll from the cells. Prochlorophytes in particular could be recognized from their
flow cytometric signatures and their cell cycle position could be determined. Moreover, the
distributions of DNA per cell reveal relative growth rates for cells growing under different