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A high throughput nile red method for quantitative measurement of neutral lipids in microalgae
Chen, wei1,2; Zhang, chengwu1; Song, lirong2; Sommerfeld, milton1; Hu, qiang1
SourceJournal of microbiological methods
2009-04-01
Volume77Issue:1Pages:41-47
ISSN0167-7012
KeywordBiofuel Fluorescence Green algae Microalgae Nile red Neutral lipids
Corresponding AuthorHu, q, arizona state univ, dept appl biol sci, 7001 e williams field rd, mesa, az 85212 usa
AbstractIsolation of high neutral lipid-containing microalgae is key to the commercial success of microalgae-based biofuel production. the nile red fluorescence method has been successfully applied to the determination of lipids in certain microalgae, but has been unsuccessful in many others, particularly those with thick, rigid cell walls that prevent the penetration of the fluorescence dye. the conventional "one sample at a time" method was also time-consuming. in this study, the solvent dimethyl sulfoxide (dmso) was introduced to microalgal samples as the stain carrier at an elevated temperature. the cellular neutral lipids were determined and quantified using a 96-well plate on a fluorescence spectrophotometer with an excitation wavelength of 530 nm and an emission wavelength of 575 run. an optimized procedure yielded a high correlation coefficient (r-2 = 0.998) with the lipid standard triolein and repeated measurements of replicates. application of the improved method to several green algal strains gave very reproducible results with relative standard errors of 8.5%, 3.9% and 8.6%, 4.5% for repeatability and reproducibility at two concentration levels (2.0 mu g/ml and 20 mu g/ml), respectively. moreover, the detection and quantification limits of the improved nile red staining method were 0.8 mu g/ml and 2.0 mu g/ml for the neutral lipid standard triolein, respectively. the modified method and a conventional gravimetric determination method provided similar results on replicate samples. the 96-well plate-based nile red method can be used as a high throughput technique for rapid screening of a broader spectrum of naturally-occurring and genetically-modified algal strains and mutants for high neutral lipid/oil production. (c) 2009 published by elsevier b.v.
English AbstractIsolation of high neutral lipid-containing microalgae is key to the commercial success of microalgae-based biofuel production. the nile red fluorescence method has been successfully applied to the determination of lipids in certain microalgae, but has been unsuccessful in many others, particularly those with thick, rigid cell walls that prevent the penetration of the fluorescence dye. the conventional "one sample at a time" method was also time-consuming. in this study, the solvent dimethyl sulfoxide (dmso) was introduced to microalgal samples as the stain carrier at an elevated temperature. the cellular neutral lipids were determined and quantified using a 96-well plate on a fluorescence spectrophotometer with an excitation wavelength of 530 nm and an emission wavelength of 575 run. an optimized procedure yielded a high correlation coefficient (r(2) = 0.998) with the lipid standard triolein and repeated measurements of replicates. application of the improved method to several green algal strains gave very reproducible results with relative standard errors of 8.5%, 3.9% and 8.6%, 4.5% for repeatability and reproducibility at two concentration levels (2.0 mu g/ml and 20 mu g/ml), respectively. moreover, the detection and quantification limits of the improved nile red staining method were 0.8 mu g/ml and 2.0 mu g/ml for the neutral lipid standard triolein, respectively. the modified method and a conventional gravimetric determination method provided similar results on replicate samples. the 96-well plate-based nile red method can be used as a high throughput technique for rapid screening of a broader spectrum of naturally-occurring and genetically-modified algal strains and mutants for high neutral lipid/oil production. (c) 2009 published by elsevier b.v.
SubjectBiochemical Research Methods; Microbiology
WOS HeadingsScience & technology ; Life sciences & biomedicine
WOS SubjectBiochemical research methods ; Microbiology
WOS Subject ExtendedBiochemistry & molecular biology ; Microbiology
WOS Keyword PlusRapid screening method ; Biotechnology ; Cells ; Dye
Indexed TypeSci
Language英语
WOS IDWos:000265314000007
Available Date2010-10-13