Designing activatable aptamer probes for simultaneous detection of multiple tumor-related proteins in living cancer cells.

We report a novel strategy for high specific and simultaneous detection of multiple tumor-related proteins in cancer cells based on the activated fluorescence signal, which is triggered by specific-binding-induced conformation alteration of the designed activatable aptamer probe. The activatable aptamer probe consists two fragments: a target-protein-recognized aptamer sequence for specifically recognizing the protein, and an extending spacer making the aptamer in hairpin structure to enable the close proximity of quencher to fluorophore, which is labeled at 5'- and 3'-terminus, respectively, of the probes. Before interaction with cell, the fluorescence of the probe is quenched due to fluorescent resonance energy transfer (FRET) between the fluorophore and quencher. After interaction events, the fluorescence signal is activated through specific binding of the probe with target protein in cell, causing the conformation alteration and forcing the separation of fluorophore from the quencher. We achieve simultaneous detection of multiple tumor-related proteins in cells by designing the different activatable aptamer probes with various fluorophore/quencher combinations. Moreover, it can also achieve a high detection sensitivity (for example, detecting MCF-7 cells at a low abundance of ~(10±5) cells mL(-1)) and specific discrimination of the subtype of cancers. The advantage of this approach is that it has high detection sensitivity because of the significant suppression of background with use of the designed activatable aptamer probe. In addition, it has ability of avoiding false signals arising due to the nonspecific adsorption of interferents because it operates via monitoring the activated fluorescence signals of the designed activatable aptamer probe.