New strategy for label-free and time-resolved luminescent assay of protein: conjugate Eu3+ complex and aptamer-wrapped carbon nanotubes.

We report here a carbon nanotube-based approach for label-free and time-resolved luminescent assay of lysozyme (LYS) by engineering an antilysozyme aptamer and luminescent europium(III) (Eu3+) complex The sensing mechanism of the approach is based on the exceptional quenching capability of carbon nanotubes for the proximate luminescent Eu3+- complex and different propensities of single stranded DNA and the DNA/protein complex to adsorb on carbon nanotubes. The luminescence of a mixture of chlorosulfonylated tetradentate beta-diketone-Eu3+ and the antilysozyme aptamer was efficiently quenched by single-walled carbon nanotubes (SWNTs) unless the aptamer interacted with LYS. Due to the highly specific recognition ability of the aptamer for the target and the powerful quenching property of SWNTs for luminescence regents, this proposed approach has a good selectivity and high sensitivity for LYS. In the optimum conditions described, >700-fold signal enhancement was achieved for micromolar LYS, and a limit of detection as low as 0.9 nM was obtained, which is about 60-fold lower than those of commonly used fluorescent aptamer sensors. Moreover, due to the much longer lifetime of the Eu3+ luminescence than those of the ubiquitous endogenous fluorescent components, the time-resolved luminescence technique could be conveniently used for application in complicated biological samples. LYS concentrations inhuman urine were thus detected using time-resolved luminescence measurement with satisfactory recoveries of 95-98%.