We carried out a systematic exploration of lipophilic tag molecules that allow efficient anchoring of aptamers to live cell surfaces. Among the lipids tested, the C-16 dialkyl (dipalmitoylphosphatidylethanolamine) tag showed a good performance: a high anchoring yield and long retention on live cells. The 3'-C-16 dialkyl tag-labeled fluorescent aptamer sensor, targeting thrombin, was prepared. The aptamer sensor was anchored successfully to live cells, allowing fluorescence detection of thrombin on the cell surface.