为了实现三角褐指藻细胞内油脂含量的快速高通量测定，探索了尼罗红荧光染色法检测其油脂含量的检测条件.采用96孔板并配合酶标仪考察三角褐指藻的预处理液的种类及其用量、染色液尼罗红的用量、染色温度、染色时间以及细胞浓度对荧光强度的影响，确立最优染色条件，并研究荧光强度与油脂含量之间的对应关系. 结果表明，TritonX-100终浓度为0.05 %时，通透效果最佳，每100 μL藻悬液中加3 μL尼罗红染液(0.1 mg/mL)，温度40 ℃，时间10 min，细胞浓度低于3×10 6个/mL时，荧光强度（y）与油脂含量（x）之间呈现良好的线性关系：y=927.37x+15.913，R2=0.9247.方法优化后，90 min即内可完成对96个样品的处理及分析，极大提高了分析的通量.
In order to achieve the rapid high-throughput determination of intracellular lipid content of phaeodacylum tricornutum, the detection conditions were investigated by Nile red fluorescence staining. To establish the optimal dyeing conditions, the 96 well plates and microplate reader were used to examine the various pretreatment solutions and their usage, the amount of Nile red, the dyeing temperature, time, and the effect of cell concentration on fluorescence intensity. And the relationship between fluorescence intensity and oil content was further investigated. The results showed that the optimal conditions for detection were as follows: the final concentration of TritonX-100 at 0.05%, 3 μL of Nile red dye solution (0.1 mg/mL) to 100 μL of algal suspension，temperature 40 ℃ and the time 10 min. There was a good linear relationship between the fluorescence intensity (y) and the oil content (x) of the algal cells in the range of 3×10 6 cells/mL, y=927.37x+15.913，R2=0.9247.After optimization of the method, 96 samples can be processed and analyzed within 90 minutes, greatly improving the throughput of the analysis.
引用本文格式： 曹素娟,柳科欢,任玲萱,侯兴国,卿人韦,兰利琼. 快速高通量测定三角褐指藻油脂含量[J]. 四川大学学报: 自然科学版, 2018, 55: 1331.复制