A Simple and Rapid Protocol for Measuring Neutral Lipids in Algal Cells Using Fluorescence

Abstract

Algae are considered excellent candidates for renewable fuel sources due to their natural lipid storage capabilities. Robust monitoring of algal fermentation processes and screening for new oil-rich strains requires a fast and reliable protocol for determination of intracellular lipid content. Current practices rely largely on gravimetric methods to determine oil content, techniques developed decades ago that are time consuming and require large sample volumes. In this paper, Nile Red, a fluorescent dye that has been used to identify the presence of lipid bodies in numerous types of organisms, is incorporated into a simple, fast, and reliable protocol for measuring the neutral lipid content of Auxenochlorella protothecoides, a green alga. The method uses ethanol, a relatively mild solvent, to permeabilize the cell membrane before staining and a 96 well micro-plate to increase sample capacity during fluorescence intensity measurements. It has been designed with the specific application of monitoring bioprocess performance. Previously dried samples or live samples from a growing culture can be used in the assay. Video LinkThe video component of this article can be found at

Multifaceted biofuel production by microalgal isolates from Pakistan

“…Fluorimetry techniques used in conjunction with confocal microscopy further validated the results. The fluorimetry method uses ethanol, a relatively mild solvent, to permeabilize the cell membrane before staining with the lipophilic dyes . High levels of dye‐conjugated lipid fluorescence were observed in Gloeocystis for both Nile red and BODIPY (11.06 and 9.04 RFU (relative fluorescence units), respectively) while, using the same dyes, relatively lower levels were observed in Dictyochloropsis (6.8 and 12.4 RFU, respectively) and Sphaerocystis (9.3 and 18.2 RFU, respectively) (Fig.…”

Mass‐spectrometry‐based lipidomics

“…C8 and C18 cartridges have been used to isolate cerebrosides, gangliosides, eicosanoids, steroid hormones and FAs from polar matrixes in biofluid samples [26,[29][30][31]. 96-well SPE plates enable pretreatment of 96 samples at once, which enables improved automation, throughput and parallel mode of lipid extractions [32,33]. The development of automated SPE that is coupled online with LC largely simplifies lipid extraction procedures [34][35][36].…”

Primitive red alga <i>Cyanidioschyzon merolae</i> accumulates storage glucan and triacylglycerol under nitrogen depletion

“…In algae, fluorescence determination of TAG in cells was reported us- ing Nile Red in 96-well plates (Storms et al, 2014); however, Nile Red undergoes a fluorescence wavelength change with solvent addition and can be masked by chlorophyll fluorescence. Therefore, we used BODIPY493/503 to evaluate the TAG content using fluorescence microscopy.…”