We will develop semiconductor quantum dot (QD) nanoparticles to be used as probes for multiplex detection of proteins. By varying the diameter of QDs, we will generate probes with tunable emission spanning the spectrum from the infrared to the ultraviolet. Western blots will be prove with primary antibodies against specific proteins, followed by addition of secondary antibodies conjugated to QDs of defined size and wavelength of luminescence emission. The QDS will be functionalized with Ni++-NTA chelating complex. Conjugation of secondary antibodies to QDs will be mediated by an IgG-binding affinity tag, which will contain an N-terminal hexahistidine tail for immobilization to QDs. The specificity of each reporting event will be realized by matching primary and secondary antibodies, and by coupling secondary antibodies to the tagged QD complex prior to probing the Western blot. The unique emission wavelength of each QD size- population will provide accurate identification of target proteins. QDs have desirable optical properties including resistance to photobleaching, non-overlapping emission profiles and stability at room temperature. Moreover, different size nanoparticles can be excited at a single wavelength, while they emit size-dependent luminescence at wavelengths spanning the visible spectrum, thus obviating the need for complex multi-laser instrumentation.
Highly luminescent semiconductor QDs can be used as probes in multiplex detection of proteins and nucleic acids, immunoassays, medical imaging, immunohistochemical procedures, analysis of clinical samples, simultaneous detection of multiple antigens to flow cytometry, highly sensitive staining of proteins and nucleic acids in electrophoretic separation techniques and high-throughput screenings of ligands using microarray platforms. The QD surface permits facile derivatization with a variety of tags, including antibodies, peptides and aptamers, enabling biosensor applications for rapid, multi-analyte detection of biomolecules and microorganisms.
