Quantum dots as FRET acceptors for highly sensitive multiplexing immunoassays

摘要

Homogeneous immunoassays have the benefit that they do not require any time-consuming separation steps. FRET is one of the most sensitive homogeneous methods used for immunoassays. Due to their extremely strong absorption over a broad wavelength range the use of quantum dots as FRET acceptors allows for large Foerster radii, an important advantage for assays in the 5 to 10 nm distance range. Moreover, because of their size-tunable emission, quantum dots of different sizes can be used with a single donor for the detection of different analytes (multiplexing). As the use of organic dyes with short fluorescence decay times as donors is known to be inefficient with quantum dot acceptors, lanthanide complexes with long luminescence decays are very efficient alternatives.rnIn this contribution we present the application of commercially available biocompatible CdSe/ZnS core/shell quantum dots as multiplexing FRET acceptors together with a single terbium eomplex as donor in a homogeneous immunoassay system. Foerster radii of 10 nm and FRET efficiencies of 75 % are demonstrated. The high sensitivity of the terbium-to-quantum dot FRET assay is shown by sub-100-femtomolar detection limits for two different quantum dots (emitting at 605 and 655 nm) within the same biotin-streptavidin assay. Direct comparison to the FRET immunoassay "gold standard" (FRET from Eu-TBP to APC) yields a three orders of magnitude sensitivity improvement, demonstrating the big advantages of quantum dots not only for multiplexing but also for highly sensitive nanoscale analysis.