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Advisor(s)
Abstract(s)
Membrane integrity has been used as a criterion for
the definition of cell viability. In the present work, staining
conditions (time and dye concentration) for the evaluation of
membrane integrity in a fluorescence microplate reader, using
the membrane-impermeant nucleic-acid dye SYTOX Green,
were optimized. Incubating Pseudokirchneriella subcapitata
algal cells with 0.5 μmol/l SYTOX Green for 40 min allowed
a clear discrimination between live (intact plasma membrane)
and dead cells (with compromised plasma membrane). Algal
cell suspensions, labelled with SYTOX Green, exhibited a
green fluorescence proportional to the fraction of the cells
with a permeabilized plasma membrane. The optimized staining
conditions were used to assess the toxicity of 1-pentanol
on P. subcapitata in a short-term exposure (6 h) assay. The
loss of membrane integrity in the cell population increased
with the concentration of 1-pentanol. The 6-h EC10 and EC50
values were 7,617 mg/l 1-pentanol (95 % confidence limits
4,670–9,327) and 12,818 mg/l 1-pentanol (95 % confidence
limits 10,929–15,183), respectively. The developed
microplate-based short-term assay can be useful in the highthroughput
screening of toxics or environmental samples using
the alga P. subcapitata.
Description
Keywords
Membrane integrity Microplate assay Selenastrum capricornutum 1-Pentanol SYTOX Green Acute toxicity
Pedagogical Context
Citation
Publisher
Springer Verlag