An improved high-throughput Nile red fluorescence assay for estimating intracellular lipids in a variety of yeast species.
An improved high-throughput Nile red fluorescence assay for estimating intracellular lipids in a variety of yeast species.
复制标题
改进的高通量尼罗红荧光测定法，用于估计各种酵母菌中细胞内脂质。
DOI：
10.1016/j.mimet.2012.09.001
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发表时间：
2012-11
影响因子：
2.2
通讯作者：
Boundy-Mills, K.
中科院分区：
文献类型：
--
作者：
Sitepu, I. R.;Ignatia, L.;Franz, A. K.;Wong, D. M.;Faulina, S. A.;Tsui, M.;Kanti, A.;Boundy-Mills, K.
为了筛选大量的酵母样品，需要一种快速、廉价的方法来估计酵母中的脂肪含量。尼罗红是一种亲脂荧光染料，用于检测和定量各种生物系统中的细胞内脂滴，包括藻类、酵母菌和丝状真菌。然而，一种已发表的酵母菌检测方法受到细胞膜上可变扩散的影响，以及达到最大荧光发射所需时间的变化。在本研究中，改变了可能影响发射的参数，以确定最佳的测定条件。建立了一种具有高通量能力的改进的分析方法，包括添加二甲基亚砜(DMSO)溶剂以提高细胞渗透性，取消洗涤步骤，降低尼罗红浓度，动态读数而不是单一时间点读数，以及使用黑色96孔微孔板。通过与重量法测定多种子囊菌和担子菌酵母菌的脂肪含量的比较，验证了改进方法的有效性。
A rapid and inexpensive method for estimating lipid content of yeasts is needed for screening large numbers of yeasts samples. Nile red is a fluorescent lipophilic dye used for detection and quantification of intracellular lipid droplets in various biological system including algae, yeasts and filamentous fungi. However, a published assay for yeast is affected by variable diffusion across the cell membrane, and variation in the time required to reach maximal fluorescence emission. In this study, parameters that may influence the emission were varied to determine optimal assay conditions. An improved assay with a high-throughput capability was developed that includes the addition of dimethyl sulfoxide (DMSO) solvent to improve cell permeability, elimination of the washing step, the reduction of Nile red concentration, kinetic readings rather than single time-point reading, and utilization of a black 96-well microplate. The improved method was validated by comparison to gravimetric determination of lipid content of a broad variety of ascomycete and basidiomycete yeast species.
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影响因子：
2.2
作者：
Chen, Wei;Zhang, Chengwu;Hu, Qiang
通讯作者：
Hu, Qiang
影响因子：
2.9
作者：
COLE, TA;FOK, AK;ALLEN, RD
通讯作者：
ALLEN, RD
影响因子：
2
作者：
SUUTARI, M;PRIHA, P;LAAKSO, S
通讯作者：
LAAKSO, S
影响因子：
1.9
作者：
Cheng, Yu-Shen;Zheng, Yi;VanderGheynst, Jean S.
通讯作者：
VanderGheynst, Jean S.
影响因子：
2.9
作者：
SACKETT, DL;WOLFF, J
通讯作者：
WOLFF, J