Use of methylene blue uptake for assessing cell viability of colony-forming microalgae

During the past few years, interest in microalgae has grown, mainly because of their potential for biofuel production. Botryococcus braunii, a green microalga that can accumulate more than half of its dry weight as hydrocarbons, is one of the most important examples. This microorganism grows in colonies and there has been no reliable viability protocol reported for this species as yet. Knowing the number of dead cells in cultures is essential for the development of efficient bioprocesses such as non-destructive extraction procedures (“milking”) to obtain lipid soluble substances from microalgal biomass. Our study presents a simple colorimetric method to determine the proportion of living to dead cells in cultures, based on the uptake of methylene blue in solution by dead B. braunii cells. The main parameters influencing this process were investigated and used to develop a protocol. This technique was validated using flow cytometry and Neochloris oleoabundans, and appears not to be limited to use with B. braunii.