Figure 1. The structure of a multifunctional QD probe. Schematic illustration showing the capping ligand TOPO, an encapsulating copolymer layer, tumor-targeting ligands (such as peptides, antibodies or small-molecule inhibitors), and polyethylene glycol (PEG).

Figure 2. Methods for conjugating QDs to biomolecules. (a) Traditional covalent cross-linking chemistry using EDAC (ethyl-3-dimethyl amino propyl carbodiimide) as a catalyst. (b) Conjugation of antibody fragments to QDs via reduced sulfhydryl-amine coupling. SMCC, succinimidyl-4-N-maleimidomethyl-cyclohexane carboxylate. (c) Conjugation of antibodies to QDs via an adaptor protein. (d) Conjugation of histidine-tagged peptides and proteins to Ni-NTA-modified QDs, with potential control of the attachment site and QD:ligand molar ratios.

Figure 3. Novel optical properties of QDs for improving the sensitivity of in vivo bioimaging. (a) Comparison of fluorescence light emission from the organic dye tetramethylrhodamine isothiocyanate (TRITC; left vial), green QDs (middle vial) and red QDs (right vial) under normal room light illumination and at the same molar concentration (1.0 μM). Bright fluorescence emission is observed from the QDs but not from the dye, owing to the large absorption cross-sections of QDs. (b) Photobleaching curves showing that QDs are several thousand times more photostable than organic dyes (e.g. Texas red) under the same excitation conditions. (c) A comparison of the excited state decay curves (monoexponential model) between QDs and common organic dyes. The longer excited state lifetimes of QD probes allow the use of time-domain imaging to discriminate against the background fluorescence (short lifetimes). τ_(dye) and τ_(QD) are the delay times for the fluorescence signals to decrease to 1/e of their original values, where e is the natural log constant and is equal to 2.718. (d) Comparison of mouse skin and QD emission spectra obtained under the same excitation conditions, demonstrating that the QD signals can be shifted to a spectral region where autofluorescence is reduced.

Figure 4. Fluorescence micrographs of QD-stained cells and tissues. (a) Actin staining (green QDs) on fixed 3T3 fibroblast cells. (b) Live MDA-MB-231 breast tumor cells labeled with a red QD–antibody conjugate targeting the urokinase plasminogen receptor. (c) Intracellular labeling of live mammalian cells using QD–Tat peptide conjugates [25^••]. (d) Frozen tissue specimens stained with QDs (targeting the CXCR4 receptor, red) and a nuclear dye (green).

Figure 5. In vivo targeting and imaging with QDs. (a) Ex vivo tissue examination of QD-labeled cancer cells trapped in a mouse lung [44^•]. (b) Near-infrared fluorescence of water-soluble type II QDs taken up by sentinel lymph nodes [49^••]. (c) In vivo simultaneous imaging of multicolor QD-encoded microbeads injected into a live mouse [25^••]. (d) Molecular targeting and in vivo imaging of a prostate tumor in mouse using a QD–antibody conjugate (red) [25^••].