Marie Dominique, Rigaut-Jalabert Fabienne, Vaulot Daniel
Sorbonne Universités UPMC (PARIS 06), UMR 7144 and FR2424, Station Biologique de Roscoff, Place Georges Teissier, 29688, Roscoff, France; CNRS, UMR 7144 and FR2424, Station Biologique de Roscoff, Place Georges Teissier, 29688, Roscoff, France.
Cytometry A. 2014 Nov;85(11):962-8. doi: 10.1002/cyto.a.22517. Epub 2014 Aug 25.
Preservation of cells, choice of fixative, storage, and thawing conditions are recurrent issues for the analysis of phytoplankton by flow cytometry. We examined the effects of addition of the surfactant Pluronic F68 to glutaraldehyde-fixed photosynthetic organisms in cultures and natural samples. In particular, we examined cell losses and modifications of side scatter (a proxy of cell size) and fluorescence of natural pigments. We found that different marine phytoplankton species react differently to the action of Pluronic F68. In particular, photosynthetic prokaryotes are less sensitive than eukaryotes. Observed cell losses may result from cell lysis or from cell adhesion to the walls of plastic tubes that are commonly used for flow cytometry analysis. The addition of the surfactant, Pluronic F68, has a positive effect on cells for long-term storage. We recommend to modify current protocols for preservation of natural marine planktonic samples, by fixing them with glutaraldehyde 0.25% (final concentration) and adding Pluronic F68 at a final concentration of 0.01% in the samples before preservation. Pluronic F68 also appears effective for preserving samples without fixation for subsequent sorting, e.g. for molecular biology analyses. © 2014 International Society for Advancement of Cytometry.
对于通过流式细胞术分析浮游植物而言，细胞保存、固定剂的选择、储存及解冻条件一直是反复出现的问题。我们研究了在培养物和天然样品中，向戊二醛固定的光合生物添加表面活性剂普朗尼克F68的效果。具体而言，我们研究了细胞损失以及侧向散射（细胞大小的一个指标）和天然色素荧光的变化。我们发现不同的海洋浮游植物物种对普朗尼克F68的作用反应不同。特别是，光合原核生物比真核生物更不敏感。观察到的细胞损失可能是由于细胞裂解或细胞粘附到常用于流式细胞术分析的塑料管管壁上所致。表面活性剂普朗尼克F68的添加对细胞长期储存具有积极作用。我们建议修改当前天然海洋浮游生物样品的保存方案，即在保存前用0.25%（终浓度）的戊二醛固定样品，并在样品中添加终浓度为0.01%的普朗尼克F68。普朗尼克F68对于保存未经固定的样品以供后续分选（例如用于分子生物学分析）似乎也有效。© 2014国际细胞计量学促进协会。
