CARLSBAD, Calif.—Nanocrystalline quantum dots (Qdots) are seeing increased use in high-throughput binding experiments, but researchers from Invitrogen noted that most published work involved home-made systems, limiting their widespread utility. They therefore undertook a systematic examination of binding assays using commercially available Qdots and Alexa Fluor dyes, publishing their findings in Analytical Biochemistry.
Initially, the researchers conjugated small-molecule haptens (biotin, fluorescein and cortisol) to Qdots and monitored the binding of the fluorescent dyes conjugated to anti-hapten antibodies or natural binding partners using fluorescence resonance energy transfer (FRET) whereby low-wavelength light stimulated Qdot fluorescence that in turn stimulated dye fluorescence. They then used this system as the basis of a homogeneous competitive immunoassay of free haptens.
Using biotin-conjugated Qdots, the researchers found that streptavidin-conjugated Alexa Fluor had a 100-fold lower limit of detection for the biotin derivative biocytin than anti-biotin Alexa Fluor. Furthermore, they noted that fully biotinylated Qdots were 10-fold less sensitive than Qdots with 25 percent biotin coverage. They were likewise able to detect fluorescein and cortisol using antibody-conjugated dyes. As such, the researchers suggested that this study provides a first-step to widely applicable binding assays using Qdots.