Peptide-functionalized upconversion nanoparticles-based FRET sensing platform for Caspase-9 activity detection in...

In the present work, peptide-functionalized upconversion@silica nanoparticle (termed as UCNP@SiO₂@Cy5-pep)-based fluorescence resonance energy transfer (FRET) sensing platform has been fabricated for the detection of caspase-9 activity in vitro and in vivo. A Cy5 labeled peptide containing specific motif LEHD for caspase-9 cleavage was designed and conjugated with UCNP@SiO₂ through covalent attachment. The red upconversion luminescence (UCL) emission of UCNP@SiO₂ can be quenched by Cy5 while the green UCL emission of UCNP@SiO₂ remains undisturbed. After the cleavage of LEHD by caspase-9, the Cy5 departed from UCNP@SiO₂ surface, resulting in recovery of red UCL emission of UCNP@SiO₂. The UCNP@SiO₂@Cy5-pep has been successfully used to monitoring the changes of caspase-9 activity levels in apoptotic cancerous cells (MG-63 and SW480) by cisplatin-induction. Under same experimental conditions, it is found that the intracellular caspase-9 activity level of cisplatin-treated MG-63 cells is higher than that of cisplatin-treated SW480. This result is consistent with that of commercial caspase-9 activity kits. The UCL signal intensity ratio of red emission to green emission of UCNP (termed as R/G) is linearly dependent on the amount of MG-63 cells within the range of 5 × 10³ to 1 × 10⁶ cells (i.e., 0.5-100 U mL^-1 caspase-9) with a limit of detection (LOD) of 675 cells (i.e., 0.068 U mL^-1 caspase-9). Furthermore, the practicability of UCNP@SiO₂@Cy5-pep is demonstrated by detection of caspase-9 activities in tumor tissues in vivo, and satisfactory results are obtained.