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.
