Luminescent Biocompatible Quantum Dots: A Tool for Immunosorbent Assay Design

We have developed several conjugation strategies based on noncovalent self-assembly for the attachment of proteins and other biomolecules to water-soluble luminescent colloidal semiconductor nanocrystals (quantum dots [QDs]). The resulting QD-protein conjugates were employed in designing a variety of bioinspired applications, including single and multiplexed immunosorbent assays to detect toxins and small molecule explosives. In these studies we showed that QD fluorophores offer several important advantages. In particular, their tunable broad excitation spectra combined with narrow fluorescence emission peaks permit single-line excitation of multiple color nanocrystals, with facile signal deconvolution to extract individual contributions from each population (e.g., size) of QDs in multiplexed assays. Furthermore, the QDs strong resistance to photobleaching under continuous illumination relative to many organic dyes makes them ideal fluorophores for long-term cellular imaging studies. This chapter details the materials and methods for the synthesis of surface-functionalized CdSe-ZnS core-shell QDs, the construction and preparation of recombinant proteins, the conjugation of antibodies (and antibody fragments) to QDs, and the use of antibody-conjugated QDs in fluoroimmunoassays.