Isolation of monovalent quantum dot-nucleic acid conjugates using magnetic beads.

Abstract

Control of the valency that is achieved in the decoration of quantum dots (QDs) remains a challenge due to the high surface area of nanoparticles. A population distribution of conjugates is formed even when reactions involve use of one-to-one molar equivalents of the ligand and QD. Monovalent conjugates are of particular interest to enable the preparation of multinanoparticle constructs that afford improved analytical functionality. Herein, a facile method for the formation and purification of QD-DNA monoconjugates (i.e., 1 DNA per QD) is described. Using diethylaminoethyl (DEAE) functionalized magnetic beads, a protocol was developed and optimized to selectively isolate QD-DNA monoconjugates from a mixture. Monoconjugates prepared with oligonucleotides as short as 19 bases and as long as 36 bases were successfully isolated. The monoconjugates were isolated in less than 5 min with isolation efficiencies between 68% and 93%, depending on the length of oligonucleotide that was used. The versatility of the method was demonstrated by purifying monoconjugates prepared from commercially available, water-soluble QDs. The isolation of monoconjugates was confirmed using agarose gel electrophoresis and single molecule fluorescence spectroscopy. Examples are provided comparing the analytical performance of monoconjugates to collections of nanoparticles of mixed valencies, indicating the significance of this separation method to prepare nanomaterials for bioassay design.