Annu. Rev. Biomed. Eng. 2005. 7:55–76 doi: 10.1146/annurev.bioeng.7.060804.

100432 Copyright c 2005 by Annual Reviews. All rights reserved First published online as a Review in Advance on April 4, 2005

A. Paul Alivisatos,1,2 Weiwei Gu,2,3 and Carolyn Larabell2,4
1 Department of Chemistry, University of California, Berkeley, California 94720; email: 2 Materials Science Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720; email:, 3 Department of Anatomy, University of California, San Francisco, California 94143 4 Physical Bioscience Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720

Annu. Rev. Biomed. Eng. 2005.7:55-76. Downloaded from by Inflibnet N-LIST Programme on 09/03/11. For personal use only.

Key Words semiconductor nanocrystals, nanoparticles, fluorescent probing, imaging, biological application ■ Abstract Robust and bright light emitters, semiconductor nanocrystals [quantum dots (QDs)] have been adopted as a new class of fluorescent labels. Six years after the first experiments of their uses in biological applications, there have been dramatic improvements in understanding surface chemistry, biocompatibility, and targeting specificity. Many studies have shown the great potential of using quantum dots as new probes in vitro and in vivo. This review summarizes the recent advances of quantum dot usage at the cellular level, including immunolabeling, cell tracking, in situ hybridization, FRET, in vivo imaging, and other related technologies. Limitations and potential future uses of quantum dot probes are also discussed. CONTENTS
INTRODUCTION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . QUANTUM DOTS: FROM PHYSICAL CHEMISTRY TO BIOLOGY . . . . . . . . . . Synthesis and Optical Properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Biocompatibility . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Bio-Molecule Conjugation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Specificity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . QUANTUM DOTS AS IN VITRO PROBES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Immunolabeling—Molecular Localization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Immunolabeling—Signaling Pathways . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Live Cell Markers and Cell Lineage . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Cell Motility Assay . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . In Situ Hybridization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Fluorescence Resonance Energy Transfer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . QUANTUM DOTS AS IN VIVO PROBES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . OTHER APPLICATIONS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . PROSPECTIVE . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1523-9829/05/0815-0055$20.00 56 56 56 59 60 60 61 61 61 63 64 65 66 66 68 69 70






Fluorescence probes are widely used in cell biology. Organic fluorophores, the most commonly used probes, suffer from fast photobleaching and broad, overlapping emission lines, and therefore are limited in applications involving long-term imaging and multicolor detection. Advances in synthesis and biofunctionalization of colloidal semiconductor nanocrystals during the past decade have generated an increasingly widespread interest among investigators in the fields of biology and medicine. These nanometer-sized crystalline particles, also called quantum dots (QDs), are composed of periodic groups of II–VI (e.g., CdSe) or III–V (e.g., InP) materials. They are robust fluorescence emitters with size-dependent emission wavelengths. Their extreme brightness and resistance to photobleaching enables the use of very low laser intensities over extended time periods, making them especially useful for live-cell imaging, such as consecutive acquisition of z-stacks for high-resolution three-dimensional (3-D) reconstructions over time [four-dimensional (4-D) imaging]. The intense brightness is also particularly helpful for single-particle detection and an increasing number of biomedical assays. The tunable emission wavelength and distinct emission spectra of QDs facilitates data acquisition and analysis of multiple tagged molecules of interest. Quantum dot chemistry and its early biological applications have been reviewed elsewhere (1–7). In the past six years, the progress in synthesis and optimization of QDs for biological environments has opened the doors to an expanding variety of biological applications, such as serving as specific markers for cellular structures and molecules, tracing cell lineage, monitoring physiological events in live cells, measuring cell motility, and tracking cells in vivo. Therefore, in this review we summarize recent progress in QD biological and biomedical applications.

Annu. Rev. Biomed. Eng. 2005.7:55-76. Downloaded from by Inflibnet N-LIST Programme on 09/03/11. For personal use only.

The synthesis of monodisperse semiconductor nanocrystals, such as CdSe, CdS, or CdTe, can be achieved by injecting liquid precursors into hot (300◦ C) coordinating organic solvent (8, 9). Adjusting the amount of precursors and crystal growth time generates QDs of specific sizes (9). The quantum yield of the nanocrystal core synthesized as above is relatively low (less than 10%) (8, 10). Usually, a shell of high band-gap semiconductor material, such as ZnS, is epitaxially grown around the core to achieve the quantum yield of up to 80% (10, 11). QDs, which are only a few nanometers in diameter, exhibit discrete sizedependent energy levels. As the size of the nanocrystal increases, the energy gap also increases, yielding a size-dependent rainbow of colors. Extensive tunability, from ultraviolet to infrared (12), can be achieved by varying the size and the

The excitation wavelength was 476 nm and 355 nm for fluorescein and QD. small nanocrystals (∼2 nm) made of CdSe emit in the range between 495 to 515 nm.7:55-76. 2005. Furthermore. The figure is reprinted with permission from Reference 12. . Rev. For example. one of which is their unique optical spectra. they have asymmetric emission spectra broadened by a red-tail. copyright 1998 AAAS. organic dyes typically have narrow absorption spectra.annualreviews. enabling simultaneous examination of multiple molecules and events. with minimum signal overlap. composition of QDs (Figure 1). and their emission spectra are symmetric and narrow. QDs have broad absorption spectra. For personal use only. Eng. multicolor nanocrystals of different sizes can be excited by a single wavelength shorter than their emission wavelengths.QUANTUM DOTS AS CELLULAR PROBES 57 by Inflibnet N-LIST Programme on 09/03/11. Figure 2 Excitation (dotted line) and fluorescence (solid line) spectra of fluorescein (top) and a typical water-soluble QD (bottom). 12). QDs have several dramatically different properties compared to organic fluorophores. whereas larger CdSe nanocrystals (∼5 nm) emit between 605 and 630 nm (8. As illustrated in Figure 2. Biomed. respectively. Consequently. Downloaded from www. In contrast. enabling excitation by a wide range of wavelengths. which means they can only be excited within a narrow window of wavelengths.

Biomed. 22. there is limited evidence suggesting that QD blinking can be suppressed on some timescale by passivating the QD surface with thiol moieties (24). which imposes some limitations in QD applications requiring single-molecule detection. Another interesting characteristic of QDs is their fluorescence lifetime of 10 to 40 ns (4. The figure is reproduced with permission from Reference 14. Downloaded from www. 12). 21).7:55-76. making them quite well suited for examination of thick specimens and in vivo imaging using multiphoton excitation. This feature has been demonstrated in a number of biological labeling experiments where the photostability of QDs was compared with commonly used fluorophores. the use of QD labels can produce images with greatly reduced levels of background noise. QDs are also very stable light emitters owing to their inorganic composition. which is significantly longer than typical organic dyes or auto-fluorescent flavin proteins that decay on the order of a few nanoseconds (4). such as rhodamine. that is. There are also some photophysical properties of QDs that can. Combined with pulsed laser and time-gated detection. or when using QDs in free suspension (20).org by Inflibnet N-LIST Programme on 09/03/11. 2005. It has also been reported that QD fluorescence intensity increases upon excitation. However. fluorescein. One of these is the property referred to as blinking.annualreviews. an event referred to as photobrightening (25.58 ALIVISATOS GU LARABELL Annu. . This intermittence in emission of QDs is universally observed from single dots. Figure 3 Comparison of fluorescence intensities between QD 608 (emission at 608 nm) and Alexa 488 after continuous illumination. In addition. making them less susceptible to photobleaching than organic dye molecules (4. and Alexa-Fluor (Figure 3) (12–19). For personal use only. in some cases. 20. Rev. 23). Eng. be disadvantageous. QDs randomly alternate between an emitting state and a nonemitting state. This extreme photostability makes QDs very attractive probes for imaging thick cells and tissues over long time periods—a challenging task that necessitates collection of multiple optical sections without damaging the specimen. the two-photon crosssection of QDs is significantly higher than that of organic fluorophores (3.

Eng.7:55-76. . Rev. 25). dithiothreitol (36). For personal use only. During further silica shell growth. The easiest approach is to exchange the hydrophobic surfactant molecules with bifunctional molecules. which is not strong although it can be enhanced by using two thiol groups instead of one (33. which are hydrophilic on one side and hydrophobic on the other side. For CdSe/ZnS QDs. 25). 35). and polyethylene glycol (PEG)-silane ( by Inflibnet N-LIST Programme on 09/03/11. other types of silanes can be added to render a different charge and provide functional groups on the surface. Biocompatibility The core and core-shell QDs synthesized as described are only soluble in nonpolar solvents because of their hydrophobic surface layer. the particles in this case are coated with a cross-linked amphiphilic polymer. Recently other solubilization methods have been reported.QUANTUM DOTS AS CELLULAR PROBES 59 26). . Other approaches. The hydrophobic tails of the polymer intercalate with the surfactant molecules and the hydrophilic groups stick out to ensure water solubility of the particle. Many biological applications of QDs have been achieved by using mercaptohydrocarbonic acids (SH-. one of which involves coating the surface with amphiphilic polymers (14. it is problematic in fluorescence quantization studies. Although this is a general method for nanocrystals grown in different organic solvents. The trimethoxysilane groups can be cross-linked by the formation of siloxane bonds. Annu.and ZnS. . the surface must be hydrophilic. The long-term stability of the QDs depends on the bond between thiol. and although the prospects are good that they can be eliminated. for the time being these should be considered as limitations of QDs. 38). Those most frequently used are aminopropyl-silanes (APS). Most often. also called surface silanization (12. Instead of exchanging the hydrophobic surfactant. phospho-silanes. For QDs to be useful probes for examination of biological specimens. Although in most cases this property can be advantageous. 34). the final size of the particles after coating is rather large. Downloaded from www. thiols (-SH) are used as anchoring groups on the ZnS surface and carboxyl (-COOH) groups are used as the hydrophilic ends.annualreviews. Biomed. and oligomeric ligands (39). have also been reported. Because the silica shells are highly cross-linked. silanized QDs are extremely stable. An alternative approach is to grow a silica shell around the particle. 27–32). Several strategies have been used to stabilize core-shell nanocrystals in aqueous solutions.-COOH) to make QDs water soluble (13. the water solubility of the core-shell QDs capped in mercaptocarbonic acids is limited. The first step of this process involves exchanging the surface ligand with thiol-derived silane such as mercapto-trimethoxysilane (MPS). Therefore. organic dendron (37. such as coating the QDs with phospholipid micelles (19). which could place restrictions on many biological applications. the diameter is between 19 and 25 nm (35). 2005. This method’s drawbacks are that it is more laborious and the silica shell may eventually be hydrolyzed (4). Both blinking and photobrightening are linked to mobile charges on the surfaces of the dots. which bind to the ZnS shell.

and proteins. 50). 44. especially when they are used to study live cells and animals because they contain elements such as cadmium and selenium. 45). such as oligonucleotides (40. Downloaded from www. 25. the cross-linker 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) is commonly used to link -NH2 and -COOH groups. For example. The most sensitive . This adsorption is nonspecific and depends on ionic strength. Rev. mercapto (-SH) exchange. 38). 47). Annu.annualreviews. the biological functions of these molecules have not been affected by linkage of QDs. Unfortunately. peptides (42. Mattoussi and coworkers presented a method of conjugating proteins to QD surfaces using electrostatic interactions. 42–44). 40. oligonucleotides (19. In most cases. Biomed. If the surface of the nanocrystal bears -COOH. 44. biomolecules can readily detach from the surface. 52) and animal (20. Several successful approaches have been used to link biological molecules to QDs. A more stable linkage is obtained by covalently linking biomolecules to the functional groups on the QD surface using cross-linker molecules (6. 12. 41. 41) and various serum albumins (18). whereas 4-(N-maleimidomethyl)cyclohexanecarboxylic acid N-hydroxysuccinimide ester (SMCC) can be used to cross-link -SH and -NH2 groups (46). 34). and covalent linkage. temperature. -SH. pH. 51. by Inflibnet N-LIST Programme on 09/03/11. QDs need to be conjugated to biological molecules without disturbing the biological function of these molecules. are readily adsorbed to the surface of watersoluble QDs. 31. Using the above methods. including avidin/streptavidin (14.60 ALIVISATOS GU LARABELL Bio-Molecule Conjugation To make QDs more useful for probing live cells and other biological applications. and the surface charge of the molecule. 36. For personal use only. albumin (48). Toxicity Concerns have been raised about the toxicity of QDs. there have been numerous reports of conjugating QDs with various biological molecules. 42) models. including adsorption. 38. 2005. 14. including biotin (12). causing QDs to precipitate from the solution. 32. Biological molecules containing thiol groups can be conjugated to the QD surface through a mercapto exchange process (28. 45). The protein of interest was engineered to fuse with a positively charged domain. electrostatic interaction.7:55-76. 13. or -NH2 . It has been reported that simple small molecules. it is easy to link it to biological molecules that also have these reaction groups. The protein-QD conjugates prepared in this way were very stable and the fluorescence yield was even higher than that from the nonconjugated dots (33. and antibodies (13. 44. As a result. no acute and obvious CdSe QD toxicity has been detected in studies of cell proliferation and viability in live cells (17. the same problem of using thiol as anchoring group on a ZnS surface occurs because the bond between Zn and thiol is not very strong and is dynamic. 17. When properly capped by both ZnS and hydrophilic shells. Eng. which in turn interacted electrostatically with the negatively charged surface of the QD.

38. However. The toxicity of QDs was examined in Xenopus embryos (19). further slow down the photooxidation process. where injection of low concentrations of QDs had no negative effects. primary hepatocyte cultures were examined because the liver is the major target of cadmium injury (52). Downloaded from www. which have low nonspecific binding to biomolecules.annualreviews. Specificity Specificity is one of the most critical criteria for measuring the value of cellular probes. because of the low nonspecific adsorption offered by PEG (19). This happened when the QD surface coating was not stable. Eng. Coating the QD surface with ZnS eliminates toxicity owing to air exposure but provides only partial protection of the core from UV exposure. as the surface chemistry has been refined. However. Although some abnormalities were noted with increasing concentrations. 2005. Coating the QDs with a mixture of phospholipids and PEG polymer also prevents aggregation of dots and nonspecific binding. manipulating QD coating and surface charge affects their “stickiness. 36). Larger molecules. the potential effect of Cd2+ release over time has not been examined.” Both silica. Therefore.and mercaptohydrocarbonic acid–coated particles bind nonspecifically to cells (12. Biomed. such as bovine serum albumin (BSA) and polymer/streptavidin. environment in which to test QD toxicity is that of the live embryo because even slight cellular perturbances are manifested as measurable biological phenotypes. QDs were not toxic. has improved specificity of QD-tagged probes (36. Rev. There remains a pressing need for further investigations into QD toxicity in live animals and safety for diagnostic applications. exposing the CdSe to oxidization by air or UV damage. QDs are ideal probes in this area. QUANTUM DOTS AS IN VITRO PROBES Immunolabeling—Molecular Localization Fluorescence immunolabeling is widely used in cell biology for probing structure and locating signal transduction–related molecules. QD specificity has greatly improved and the number of biological applications has dramatically increased. Since the first experiment where semiconductor CdSe/ZnS nanocrystals were used to stain F-actin in fixed cells (12). 36.QUANTUM DOTS AS CELLULAR PROBES 61 Annu. Because the surface molecule and surface charge of the QDs play an important role in their binding properties (14. it was not clear whether they resulted from the QDs or changes in the osmotic equilibrium of the cell. In another case. Alternatively.7:55-76. For personal use only. 38). strategies to protect the QD surfaces from oxidative environments are critical for live cell and animal experiments. When QDs were used in early applications. Owing to their robust optical properties. investigators have performed a variety of . Under standard conditions of synthesis and water solubilization. 51). nonspecific binding was reported (12. 36). However. the use of hydroxyl groups. cytotoxicity was observed when Cd2+ was released (52).org by Inflibnet N-LIST Programme on 09/03/11.

Biomed.annualreviews. The microtubules were first incubated with mouse biotinylated antiα-tubulin antibody and then streptavidin conjugated QDs or Alexa. live cells. Ness et al. The scale bar is 20 µm. developed an immunohistochemical (IHC) protocol that combines conventional enzymatic signal amplification and QD labeling to detect intracellular antigens in rat and mouse brain tissue sections. As compared to Alexa. and tissue sections.62 ALIVISATOS GU LARABELL Annu.7:55-76. cytoskeleton fibers. They found alterations of plasma membrane organization and integrity upon heat stress. experiments in which QDs have been used to localize molecules in cells and tissues. 2005. Figure 4 Immunofluorescence labeling of microtubules for mouse embryo fibroblast cells (NIH/3T3) using streptavidin-coated QD 605 (left) and streptavidin conjugated Alexa 568 (right). mortalin. using QDs to show different staining patterns in normal and transformed cells (53). Minet and colleagues also examined breast tumor cells. and nuclear antigens in fixed cells. The excitation wavelength 488 nm was used to excite QD 605. Downloaded from www. this protein has been suggested as a reliable marker for normal versus transformed cells. Eng. QD labeling shows the same labeling pattern of microtubules. using QDs to label membrane glycoproteins to study heat stress effect (55). Rev. . The emission filter was LP 560 nm for both. Kaul and colleagues reported immunofluorescence labeling of the heat shock 70 protein. Their study showed that QD IHC labeling had greater sensitivity than similar IHC approaches using conventional dyes (16). Wu and coworkers developed reliable and specific QD probes to localize the breast cancer cell surface marker Her2. extremely high stability. high-resolution image of the membrane domain band 3 in erythrocytes (54). Taking advantage of the high photostability of QDs. with a substantial increase in brightness and photostability as compared to organic dyes (14). All these studies show that QDs have moved beyond the demonstration stage and are excellent probes with enhanced signal-to-noise ratio. Tokumasu & Dvorak were able to collect 40 consecutive optical sections using confocal microscopy and generated a 3-D by Inflibnet N-LIST Programme on 09/03/11. For personal use only. and improved specificity suitable for studying important biological problems. whereas 543 nm was used for Alexa 568. Consequently. both in live and fixed specimens (Figure 4).

and thus provided useful information for the design of artificial molecule delivery systems for gene and drug delivery. which triggers internalization of both EGF-QD and its receptor via endocytosis. the latter being the rate-limiting process (57). The process of cell endocytosis has also been studied using QDs (58). In this case. Owing to the photostability of QD. moves toward the cell body at a velocity of 10 nm/s. have also been examined using QDs (57). Biomed. The difference in size can significantly affect the molecule’s motion. .QUANTUM DOTS AS CELLULAR PROBES 63 Immunolabeling—Signaling Pathways Research of signaling pathways between and within cells also relies heavily on bright and sensitive fluorophores. QDs will be a valuable reagent for this kind of investigation. the endocytosis efficiency of 15 nm QD conjugated sugar balls was compared with the efficiency of 5 nm and 50 nm particles. Being more photostable than organic dyes and much smaller than beads. whereas a comparable Cy3 probe lasted only approximately 5 s. The QD conjugated sugar ball was an excellent size marker for studying the size effect of endocytosis in the viral size region. QDs have begun to play a major role in this field. reported using serotonin-linked QDs to target the neurotransmitter receptor on the cell surface. bridging the gap between large beads and small organic fluorophores. For personal use only.annualreviews. The 15 nm QDs were less effective at triggering endocytosis than were 50 nm particles. The QD labeling enabled the collection of sequential images for up to 20 min. In this experiment. after binding to the filopodium of the by Inflibnet N-LIST Programme on 09/03/11. the erbB1 receptor. the behavior of QD conjugates was similar to that of free serotonin. The GlyRs were linked to QDs through primary antibody and secondary F(ab ) fragment bridges.7:55-76. such as erbB/HER receptor-mediated signal transduction. EGF-QD binding and internalization kinetics were obtained. three different locations of GlyRs were characterized and corresponding diffusion coefficients were obtained. revealing that endocytosis is highly size dependent. Other signaling pathways. Although one to two orders of magnitude less potent at inhibiting the receptor than free serotonin. It is not surprising that the diffusion coefficients obtained are much larger than those measured with fluorescent beads because the QDs are extremely small (10–15 nm) compared with the beads (1 micron). 2005. Annu. Eng. but more effective than 5 nm particles (58). but also inhibited the serotonin transportation in a dose-dependent manner (43). examination of single QDs enabled discovery of a retrograde transport process in which the EGF-QD. Downloaded from www. Epidermal growth factor (EGF) conjugated to QDs is still capable of binding to and activating its receptor. QDs proved to be a suitable probe for single-molecule experiments in living cells. making QDs a valuable probe for exploring the serotonin transportation mechanism. Such quantitative understanding of the transduction mechanism is essential for receptor-targeted therapeutics. Rev. Dahan and coworkers studied the dynamics of individual glycine receptors (GlyRs) in neuronal membranes using QDs as fluorescent probes (56). Rosenthal et al. By tracking individual dots and analyzing the trajectories. The QD probes not only recognized and labeled serotonin-specific neurotransmitters on cell membranes.

they can be used as cell markers for long-term studies such as cell-cell interactions. and cell lineage tracking. If the original cell was preloaded with QDs. and subsequently undergo programmed cell death (60) (Figure 5). However. Cells can also be loaded with QDs by microinjection. extend invadopodia to contact the acini. The carrier for delivering QDs into the cells is the protein translocation domain Pep-1. Death is always contact dependent and is induced by polarized Annu.64 ALIVISATOS GU LARABELL Live Cell Markers and Cell Lineage Because QDs are extremely bright and photostable. they accumulate in vesicles in the perinuclear region (17. Compared to nonspecific endocytosis or peptide delivery. Eng. Internalized QDs are powerful probes for long-term studies of cell-cell interactions. However. One of these was developed by Matteakis and coworkers (59). cell differentiation. with the use of an endosome-specific marker. There are several other approaches for delivering QDs into live cells. 18. Biomed. Rev.annualreviews. 51). The exact pathway of incorporating QDs into cells under endocytosis is not understood. 18. Downloaded from www. human mammary epithelial cells form acini. an engineered 21-residue peptide composed of a hydrophobic sequence for QD binding and a hydrophilic lysine-rich sequence from the nuclear localization sequence for penetrating cells (59). we were able to unequivocally identify the tumor cells and normal cells in a coculture system. (19) microinjected phospholipid-coated QDs into early-stage Xenopus embryos and tracked the embryogenesis using fluorescence by Inflibnet N-LIST Programme on 09/03/11. such as fluorescein-labeled dextran (18) or pECFP (17). With different color-emitting QDs. When cultured in the 3-D matrix. QDs divided with the daughter cells at cell division (18. Our group examined the interactions of human mammary epithelial tumor cells with normal cells growing in a 3-D culture system using QDs as cell-type identification markers. who used a peptidemediated transportation method to incorporate different colored QDs into a variety of live mammalian cells. 51. Most experiments conducted to date have shown that QDs do not interfere with normal cell physiology and cell differentiation (19. For personal use only. it was confirmed that QDs were indeed accumulated in the endosomes or lysosomes and not coating them on the basis of colocalization of QDs and endosome markers.7:55-76. generating a unique and spectrally resolvable code for each cell type. 19. We observed that tumor cells migrate toward the acini. 51). Dubertret et al. Later. This enabled us to add tumor cells tagged with QDs of a different emission wavelength to the acini and clearly distinguish normal cells from tumor cells during two-week culture periods. 2005. Once internalized. all cells in the acinus will subsequently contain a subset of those QDs. Each acinus develops from a single cell during a 10–14 day time span. The idea to use QDs as cell markers is based on the discovery that QDs can be internalized by cells. 17. microinjection is more laborious and less efficient. it is worthwhile noting that microinjected QDs were homogenously distributed throughout the cell and were not compartmentalized (19). 59). after QDs enter cells. 51). polarized clusters of cells that resemble natural glandular tissue. . by either receptor-mediated (13) or nonspecific endocytosis (4. Perinuclear vesicles filled with nanocrystals were first seen by multiphoton microscopy (51).

leaving behind QD-free zones representing the pattern of phagokinetic uptake of QDs (Figure 6) (51). The scale bar is 200 µm. The figure was reproduced with permission from Reference 51. Eng. Biomed. Figure 6 QDs were used to study cell motility. which cannot be achieved by organic dyes. Present methods for measuring cell patterns and the extent of cell movement suffer from a number of practical limitations.annualreviews. Similar phagokinetic cell motility assays using gold colloids or polystyrene micro beads have been used with some degree of success (61. were grown on top of a collagen layer that had been coated with a thin layer of silanized QDs. those organized into acini. Downloaded from www. . both before and after perturbations.7:55-76. but typically require fixation of the cells. 2005. they engulfed the nanocrystals and left areas that were fluorescence-free. The use of QDs facilitates monitoring live by Inflibnet N-LIST Programme on 09/03/11. Human mammary epithelial tumor cells. such as the addition of potential chemotherapeutic agents to monitor Annu. 62). Cell Motility Assay Motility and migration of cancer cells are measurable properties that are associated with metastases and the formation of secondary tumors. While cells migrated across the layer. We developed an assay based on our finding that cells nonspecifically incorporate QDs as they crawl over them. Rev. For personal use only. MDA-MB-231.QUANTUM DOTS AS CELLULAR PROBES 65 cells. but not individual unpolarized cells. The high photostability of QDs enabled us to track and image these cocultures for up to two weeks.

These features make it a powerful new tool for cell motility studies. 28. gold (45. The experiment was performed with total genomic DNA as the probe. We also found that the quantity of QDs incorporated varies among different cell types. reported using a QD FISH probe to analyze human metaphase chromosomes. The use of organic dyes in multicolor FISH also has limitations. 40. For example. 51) and in vitro testing demonstrates that these conjugates retain their ability to form complementary sequences of Watson-Crick base pairs (51). Fluorescence Resonance Energy Transfer Fluorescence resonance energy transfer (FRET) is a process in which energy is transferred from an excited donor to an acceptor via a by Inflibnet N-LIST Programme on 09/03/11. In Situ Hybridization Annu. These drawbacks can be resolved with the use of QDs. and the ebrB2/Her2/neu gene as the target of hybridization. Later in 2004. 2005. high salt conditions inside the cells caused partial aggregation of the nanocrystals. and microbeads (69. which gave rise to varied signal intensities (36). near-field dipoledipole interaction (71. DNA or oligonucleotides can also be conjugated to QDs (19. However. In 2001. utilizes fluorescently labeled DNA probes for gene mapping and the identification of chromosomal abnormalities. Eng.annualreviews. using fluorescent detection in live cells. Typical probes that have been used include DNA conjugated to fluorescent molecules (67). and interactions. which is time-consuming and uses fixed cells. MDA-MB-231 tumor cells uptake more QDs than nontumorigenic MCF 10A cells (51). Although the results show the success of hybridization using QD probes. Fluorescent in situ hybridization. Pathak and coworkers used QD-based FISH labeling to detect Ychromosome-specific repeats in fixed human sperm cells. also called FISH. 41. 36. For personal use only. Xiao et al. use of the rapidly photobleaching organic probes compromises the experiments and requires immediate imaging. As might be expected.66 ALIVISATOS GU LARABELL changes in motility. 72). and a way to quantify gene copy numbers within tumor cells that have abnormal gene amplification. dynamics. Downloaded from www. FISH provides researchers with a method to visualize and map the genetic material in cells. FRET is very sensitive to the distance between donor and acceptor and has been used to study biomolecule conformation. Rev. Unlike the Boyden chamber assay. 70). 68). and it is much more sensitive than the conventional Boyden chamber assay with significantly reduced specimen processing (64–66). Texas-Red. The QD assay provides a useful measure of the invasive potential of cells (63). including color overlap and differential intensity of the fluorophores. They compared QD. 45. Some problems of using conventional dyes for FRET include . and FITC probe detection. The in situ hybridization procedure requires DNA-linked probes. Biomed. including specific genes or portions of genes.7:55-76. demonstrating that the QD probe was more photostable and provided higher signal-to-noise ratio than the organic probes. human metaphase lymphocytes as the hybridization substrate. the QD-based assay requires no fixation.

3-dimethyl6-nitrospiro-[2H-1-benzopyran-2. Enhanced TMR fluorescence was observed as energy transferred between the QDs and the TMR. Owing to the size of QDs. 74). which served as the FRET donor (77). Medintz and coworkers engineered a histidine tag on maltose binding protein (MBP) that finally bound QDs. BIPS was converted back to colorless spiropyran and QD emission was recovered. When exposed to white light. it is possible to use this type of system as a nanosensor device for sensing photochromical changes. This study demonstrated that QDs can be used as efficient energy donors in the FRET system and showed that by tuning their size. Later experiments demonstrated resonance energy transfer between QD donor/acceptor and organic dye acceptor/donor (75. (73.annualreviews. Annu.2. With a quencher bound to the maltose-binding site. After adding telomerase. One potential limitation of using QDs for FRET involves the physical dimensions of nanocrystals. In DNA telomerization. QD fluorescence was inhibited but was recovered by adding maltose to displace the quencher. In addition. or the degree of spectral overlap (by changing QD size). Quenching the luminescence of small dots was accompanied by an enhancement of the luminescence of large dots. reported using QDs as a FRET donor in a protein-protein binding assay (31). In 1996. C. In by Inflibnet N-LIST Programme on 09/03/11. Rev.QUANTUM DOTS AS CELLULAR PROBES 67 fast photobleaching and significant emission overlap between donor and acceptor. . BIPS was converted to colored merocyanine when exposed to UV light and quenched QD emission by acting as a FRET acceptor. By switching the QD emission on and off. These results indicate the potential use of the QD FRET process in fast and sensitive DNA detection and DNA array analyses.2. QDs were conjugated to a DNA template molecule and mixed with dNTP (N = A. resonance energy transfer between closely packed CdSe QD solids was reported by Kagan et al.7:55-76. QDs provide a potential solution to these problems. caused a substantial enhancement in energy transfer efficiency. 2005. the same group did an in-depth study using the MBP system by varying the QD size and the quantity of acceptor dye (78). In addition to protein binding assays. Increasing the amount of acceptor dye.2-(2H)-indoline]) (79). Downloaded from www. Biomed. 76). Willard et al. other processes like DNA replication and telomerization can also be probed by QD-based FRET as reported by Patolsky and coworkers (80). these same researchers demonstrated reversible modulation of QD fluorescence using FRET with the photochromic molecule BIPS (1.3-dihydro-1 2-(2-carboxyethyl)-3. A similar FRET process occurs when using QD conjugated primers to initiate DNA replication. In another paper. G) and Texas-ReddUTP. QDs were conjugated to BSA as the FRET donors and tetramethylrhodamine (TMR) was linked to the protein as the acceptor. caution must be taken with single-molecule experiments that might be complicated by the QDs blinking. This was consistent with resonance energy transfer from the small dots to the large dots. There are several examples of QDs used for FRET in biological systems. For personal use only. FRET transition occurred between the QD donor and Texas-Red acceptor. Eng. the distance between donor and acceptor is usually greater than 3 nm (77). QDs can transfer energy to a number of organic dye molecules.

Live animal imaging using QD fluorescence with multiphoton microscopy was achieved by Larson et al. In the other paper. multiphoton microscopy enables the imaging of structures deep within biological tissues with minimum photobleaching and photodamage. To optimize the conditions of in vivo experiments. extending into the infrared by Inflibnet N-LIST Programme on 09/03/11. in vivo imaging of QDs was much brighter and more sensitive than imaging with green fluorescence protein (GFP) (Figure 7). The nanoparticles. including light and electron microscopy on tissue sections and noninvasive whole-body fluorescence imaging (81). Biomed. A two-photon process requires about twice the excitation wavelength of a single-photon excitation. After injection into mice that had been transplanted with human prostate cancer cells. Eng. 2005. QDs provided a stable. who mapped sentinel lymph nodes (SLN) with near-infrared fluorescent QDs in mouse and pigs. tumor blood vessels and tumor lymphatic vessels. reported in vivo cancer targeting and imaging using QDs (21). Although the QDs showed no deleterious effects upon the animals in these studies. a more detailed evaluation of potential QD toxicity in the body is warranted prior to their long-term usage in higher organisms. Histological staining revealed that QDs were delivered to the appropriate site in vivo. QDs. QDs were injected intradermally into the animal. Owing to the QDs’ large absorption coefficient and long lifetime. Longer wavelengths. were injected into mice. microinjected phospholipid-coated QDs into early-stage Xenopus embryos to study the behavior of specific cells during embryogenesis (19). robust fluorescent probe that could be used in vivo over extended periods of time.7:55-76. Annu. Downloaded from www. Gao et al. Akerman and colleagues used QDs to target tumor vasculature in mice (42). Dubertret et al. were detectable through intact skin at the base of the dermis (∼100 micron) using an excitation wavelength of 900 nm. The use of QDs during surgical procedures was demonstrated by Kim and coworkers (82). Rev. They conjugated QDs to the antibody specific for the prostate cancer cell marker PSMA. which was totally buried in autofluorescence and background. Recently.annualreviews. Ballou et al. In one paper. can be utilized to excite chromophores in a single quantum event. intravenously injected into mice.68 ALIVISATOS GU LARABELL QUANTUM DOTS AS IN VIVO PROBES Colloidal semiconductor nanocrystals have a two-photon cross-section that is two to three orders of magnitude greater than organic dyes (20). Therefore. Two in vivo experiments appeared almost simultaneously in 2002. The high two-photon cross-section of QDs allows for more efficient probing of thick specimens and in vivo imaging when using multiphoton excitation microscopy. For personal use only. these QDs maintained their fluorescence even after four months in vivo. In both cases. Amazingly. guided by the peptides. the QD-tagged PSMA antibodies recognized and bound at the tumor site and were clearly imaged in vivo. . tested QDs with different polymer coating in vivo using various imaging techniques. in 2003 (20). conjugated to several peptides that differentially recognized blood vessels located in the lung.

which were then detected by fluorescence (90). the luminescence of CdS QDs selectively responded to the presence of zinc or copper ions. Downloaded from www. zinc. More research in this area needs to be completed before QDs can be used as probes in the human body. Chen et al. it is possible to use them as contrast or labeling probes in transmission electron and X-ray microscopy for high-resolution imaging of cells. The nanocomposite particles.annualreviews. The above studies have shown the great potential of using QDs as in vivo probes for cancer studies. Whether the QDs can ultimately be cleared from the body is not known. For personal use only. With some technical improvements. However.7:55-76. and were followed using an intraoperative imaging system. Biomed. and spleen (21. Combined with magnetic nanoparticles.QUANTUM DOTS AS CELLULAR PROBES 69 Annu. Rev. and noninvasive whole-body imaging. reported targeting QD conjugates to surface glycoproteins of bacteria and fungi (26). whereas copper ions quench their emission (89). reported that by changing the capping layer. Kloepfer et al. drug delivery. The surgeon followed the flow of QDs in real time with NIR QD image guidance. Eng. entered the lymphatic by Inflibnet N-LIST Programme on 09/03/11. The use of QDs as cell tracers after transplantation into mice has also been reported (83). Localization of specific biomolecules in cells and tissues at a high resolution provides both structural and quantitative information for molecular cell biology. 2005. QDs were also used as cell membrane permeable indicators for Escherichia coli (84). It is interesting to note that QDs here play a role as a size-defined cap and not as a fluorescent molecule (91). 42). were used to magnetically separate breast cancer cells. used CdSe QDs as probes in both conventional and energy-filtered TEM. popular immunofluorescence labeling is limited in spatial resolution. It is worth noting that the degradation and metabolism of QDs in the body remains to be investigated and there are reports that QDs injected can accumulate in the kidney. Nisman et al. as reported by Wang et al. Lai et al. liver. Because QDs are composed of heavy elements such as cadmium. this new technique might be useful for in vitro cancer diagnosis in the future. Cadmium. The luminescence of QDs can be affected by ionic environments and pH (85–88). QDs have also been investigated for use in drug delivery. and manganese ions increase the luminescence of CdS nanocrystals in basic solution. QDs have been demonstrated to be useful for cell detection and separation. demonstrating the feasibility of . and quickly identified the position of the SLN in a precise and rapid surgical procedure. OTHER APPLICATIONS QDs have also been found to be useful in the study of microorganisms. made up of magnetic Fe2 O3 superparamagnetic cores and CdSe/ZnS QD shells. The CdS cap ensures the drug is inside the system until triggered by disulfide bond–reducing reagents. used surfacemodified CdS QDs as chemically removable caps to keep pharmaceutical drug molecules and neurotransmitters inside a mesoporous silica nanosphere-based system.

A and B were reproduced with permission from Reference 92. Rev. One area of biological research that has not yet been addressed with these novel nanostructures is molecular polarity.70 ALIVISATOS GU LARABELL Annu. QDs are highlighted with arrows. Biomed. Tuning the chemistry to avoid . QDs. by Inflibnet N-LIST Programme on 09/03/11. The image was generated by dividing a cadmium postedge (510 eV) image by its preedge (415 eV) image after alignment by cross-correlation. To acquire chemical and structural information and to get better signal-to-noise ratio. Downloaded from www. Eng. PROSPECTIVE There are numerous unexplored possibilities to expand the repertoire of QD labeling in the future. with their intense luminosity and high photostability. optical confocal microscopy. Semiconductor nanorods (quantum rods). which demonstrate polarized emission properties. Figure 8 (A) Transmission electron microscopy detection of nuclear promyelocytic leukemia protein (PML) within a Hep-2 PML I cell section with QD labeling. (B) Electron spectroscopic imaging shows the distribution of discrete QDs within the PML body. The scale bar is 50 nm. unpublished data). and TEM. electron spectroscopic imaging (ESI) was used to identify the elemental map of cadmium without interference from other elements such as nitrogen (Figure 8).annualreviews. (C) Soft X-ray microscopy image of QDs in liposomes. Cellular probes that can identify a single gene.7:55-76. For personal use only. With the completion of the Human Genome Project. labeling the nuclear promyelocytic leukemia (PML) protein on cell sections to obtain correlative fluorescence and TEM images. Larabell. PET or SPECT. or protein at a specific time and location within a cell will facilitate an understanding of cellular function at the molecular level. 2005. Also. QDs are highlighted with arrows. might be potent tools for such experiments. X-ray microscopy. we have demonstrated the ability to detect liposome-encapsulated QDs using soft X-ray microscopy (C. The X-ray image was taken as described in Reference 93. The multilevel imaging on both the medical and cellular scales demonstrates the need for development of nanoparticles that can be used in all kinds of imaging techniques. it is timely to try to identify specific genes involved in disease and the relationship of gene regulation to cell function. are the best candidate for this research. such as magnetic resonance imaging.

Los Angeles. J. 6:365– 70 6.QUANTUM DOTS AS CELLULAR PROBES 71 sample aggregation within cells and optimizing imaging and detection conditions for single QD experiments will be the key to achieve this goal. 115:8706–15 Peng XG.S. et al. Sutherland AJ. Nie S. 8. Bailey RE. This work was supported (in part) by NIH National Center for Research Resources through the University of California. The Annual Review of Biomedical Engineering is online at http://bioeng. Bruchez M. Michalet X. Chem. 1993. Norris DJ. Wu X. there is an urgent need to understand QD toxicity and metabolism in the body. Han M. 9. and Christine Micheel for helpful discussions. Properties of fluorescent semiconductor nanocrystals and their applications to biological labeling. Biomed. Chem. Eng. Quantum dots as luminescent probes in biological systems. Am. Aihua Fu. Micheel C. Alivisatos AP. Solid State Mater. Phys. Gerion D. it will be more difficult to replace the toxic elements of QDs while keeping their desirable optical properties intact. Kadavanich AV. Lacoste TD. 22:47–52 2. Epitaxial growth of highly luminescent CdSe/CdS core/shell nanocrystals with photostability and Annu. 10. Biotechnol. Recent studies have shown the potential for the use of QDs in cancer diagnosis and therapy. 2:261–76 5. Thus. Dahan M.annualreviews. Te) semiconductor nanocrystallites. 1996. DE-AC0376SF00098. 2005. Luminescent quantum dots for multiplexed biological detection and imaging. Although it is possible to modify QD surfaces so that QDs are cleared from the body within a reasonable time. Dept. 2002. Yi Cui. Am. Biotechnol. Downloaded from www. Bruchez M. Alivisatos P. 13:40–46 Watson A. Single Mol. The use of nanocrystals in biological detection. 2002. Se. Curr. 1998.annualreviews. 2001. Sci. Maxwell DJ. Lighting up cells with quantum dots. Trends LITERATURE CITED 1. Zanchet D. 2003. 100:468–71 Peng XG. Rev.7:55-76. 2004. Gao X. J. 21:371–73 4. Opin. Biotechniques 34:296–300. et al. Biological applications of colloidal nanocrystals. For personal use only. Gao X. 7. Wickham J. 2–3 Murray CB. Nie S. Bawendi MG. Nat. 1997. Molecular profiling of single cells and tissue specimens with quantum dots. Synthesis and characterization of strongly luminescing ZnS-capped CdSe nanocrystals. Guyot-Sionnest P. J. . Synthesis and characterization of nearly monodisperse CdE (E = S. Alivisatos AP. Opin. Chem. We are grateful to Rosanne Boudreau and Benjamin Engel for proofreading the manuscript. 120:5343–44 Hines MA. 2003. Pellegrino T. ACKNOWLEDGMENTS We thank Dr. 11. Kinetics of II-VI and III-V colloidal semiconductor nanocrystal growth—focusing of size distributions. Soc. Parak WJ. 2003. Nanotechnology 14:R15–27 3. Curr. subaward agreement 0980GFD623 through the U. of Energy under Contract No. Chan WCW. Schlamp MC. by Inflibnet N-LIST Programme on 09/03/11. Pinaud F.

Mirkin CA. Am. Soc. et al. Liu H. Microminiaturized immunoassays using quantum dots as fluorescent label by laser confocal scanning fluorescence detection. Ding Y. Nealson KH. Kloepfer JA. Science 300:1434–36 Gao X. Norris DJ. 18. Cui Y. Lett. 14. J. 2003.72 ALIVISATOS GU LARABELL 22. Yet CP. 2001. 2003. Chen CC. Mauro JM. Noireaux V. Mitchell GP. 22:969– 76 . J. 15. Mattoussi H. electronic accessibility. 21:47–51 Hanaki K. Biotechnol. Phys. Cheng J. Momo A. 2003. Immunol. Quantum dots as strain. 69:4205–13 27. Weiss S. Long-term multiple color imaging of live cells using quantum dot bioconjugates. Zanchet D. In vivo cancer targeting and imaging with semiconductor quantum dots. Nie SM. Levenson RM. Roth KA. Pinaud F. 2002. J. Chao CY. Chung LWK. Am. Biotechnol. Am. Alivisatos AP. et al. 1998. Commun. Phys. Williams SC. Water-soluble quantum dots for multiphoton fluorescence imaging in vivo. 2003. Wang YA. Zhou Y. Simon SM. Nucleic Acids Res. Jin 12. Histochem. 1999. Stucky G. 51:981– 87 Jaiswal JK. Lett. Synthesis and properties of biocompatible watersoluble silica-coated CdSe/ZnS semiconductor quantum dots. Moronne M. Pinaud F. 123:8844–50 31. 1:469–74 32. Nadeau JL. Bruchez MP. Oku T. Letsinger RL. Langmuir 15:6845–50 28. 119:7019–29 Bruchez M. Treadway JA. 13. Lounis B. 329:399–404 by Inflibnet N-LIST Programme on 09/03/11. Peng XG. Barker PE. Nat. 17. Moerner WE. Biophys. Science 298:1759–62 Larson DR. Aldana J. Carillo LL. Photon antibunching in single CdSe/ZnS quantum dot fluorescence. Williams RM. Bechtel HA. Skourides P.annualreviews. J. Sun B. B 105:8861–71 26. 121:8122–23 29. Maenosono S. 126:1324–25 25.and metabolismspecific microbiological labels. Chem. Nat. Ma H. Cytochem. 2001. Environ. Akhtar RS. 21:41–46 Xiao Y. Am. Chem. 2004. Zipfel WR. Latham CB. 2001. Willard DM. Nie S. Wang HN. 2004. Chem. Microbiol. Soc. Jung J. 32:e28 Ness JM. Alivisatos AP. Libchaber A. Haley KN. Downloaded from www. Chem. Self-assembly of monolayers of cadmium selenide nanocrystals with dual color emission. Biomed. et al. Quantum dot bioconjugates for ultrasensitive nonisotopic detection. Hohng S. J. 2001. 26:825–27 24.7:55-76. 1999. 21. 1998. 16. Rev. Chemla DS. J. For personal use only. 2003. Appl. Opt. Biochem. Mielke RE. 19. Science 281:2016–18 Wu X. Nano Lett. 2004. Nat. 2005. Alivisatos AP. Immunofluorescent labeling of cancer marker Her2 and other cellular targets with semiconductor quantum dots. Liu J. Zhang CY. In vivo imaging of quantum dots encapsulated in phospholipid micelles. Time-gated biological imaging by use of colloidal quantum dots. Brivanlou AH. 2001. Wong MS. Nie S. Chen D. Laurence T. Xie W. Chem. Clark SW. Van Orden A. Annu. Parak WJ. Soc. Komoto A. Soc. Semiconductor nanocrystals as fluorescent biological labels. Gerion D. Dahan M. 20. CdSe-ZnS quantum dots as resonance energy transfer donors in a model protein-protein binding assay. 302:496–501 Dubertret B. Gerion D. Gin P. J. Science 281:2013–16 Chan WC. Ha T. Biotechnol. et al. Method. et al. 2003. Yi G. Photochemical instability of CdSe nanocrystals coated by hydrophilic thiols. Semiconductor nanocrystal probes for human metaphase chromosomes. Semiconductor quantum dot/albumin complex is a long-life and highly photostable endosome marker. 249:85–89 30. Res. 2000. Chem. Near-complete suppression of quantum dot blinking in ambient conditions. Eng. Combined tyramide signal amplification and quantum dots for sensitive and photostable immunofluorescence detection. Programmed assembly of DNA functionalized quantum dots.

2000. 124:4586–94 Winter JO. Targeting cell surface receptors with ligandconjugated nanocrystals. 48. 2002. Chem. Am. 4:703–7 Pathak S. Liu TY. 42. Soc. Harden HH. Ruoslahti E. Soc. 34. Am. 2004. Soc. Chen HY. 2002. Korgel BA. 43. Downloaded from www. Chan WC. Dent A. 13:1673–77 Gerion D. Bioconjugation: Protein Coupling Techniques for the Biomedical Sciences. Parak WJ. J. 2002. Am. Sci. Anderson GP. Mater. Am. 2004. Kudera S. Nanocrystal targeting in vivo. 124:7070– 74 Aslam M. et al. Nie S. 2002. 40. Am. Schmidt CE. Adams S. 229:407–14 Annu. 124:2293–98 Guo W. Nano Lett. King D. 46. Choi SK. Biochem. 1998. Eng. J. Zanchet D. Adkins EM. Bawendi MG. J. Chem. Stabilization of inorganic nanocrystals by organic dendrons. Analyst 125:1029–31 Mattoussi H. 36. Alivisatos AP. Anderson GP. Opt. Chem. 15:3125–33 Kim S. Mattoussi H. Rev. London: Macmillan Ref. Proc. Bioconjugation of highly luminescent colloidal CdSe-ZnS quantum dots with an engineered two-domain recombinant protein. Biomed. Murphy CJ. Pinaud F. Williams SC. et al. 2000. Sundar VC. Sorting fluorescent nanocrystals with DNA. 2001. tum dots: thermodynamics of “inorganic protein”-DNA interactions. Weiss S. J. Goldman ER. Chem. Phys. Li JJ. 41. Tran PT. Phys. Quantumdot nanocrystals for ultrasensitive biological labeling and multicolor optical encoding. 2001. Green TM. 2000. Anal. USA 99:12617–21 Rosenthal SJ. 2003. Biomed. Acad. Status Solidi B-Basic Res. Luminescent quantum dot-adaptor proteinantibody conjugates for use in fluoroimmunoassays. 2002. Mater. Soc. New York: Grove’s Dict. Gryczynski I. 47. Am.7:55-76. Chan WC. Chem. Bioactivation and cell targeting of semiconductor CdSe/ZnS nanocrystals with phytochelatin-related peptides. For personal use only. Oligomeric ligands for luminescent and stable nanocrystal quantum dots. 35. Manna L.QUANTUM DOTS AS CELLULAR PROBES L. Soc. 7:532–37 Goldman ER. Peng XG. Anderson GP. Micheel CM. Peng XG. Balighian ED.. Koktysh D. Gryczynski Z. 37. J. Adv. Tomlinson A. 2000. 39. Schroeter S. 2003. 125:14652–53 Lakowicz JR. 50. 126:6115–23 Gao X. J. Conjugation of luminescent quantum dots with antibodies using an engineered adaptor protein to provide new reagents for fluoroimmunoassays. Nowaczyk K.and salt-dependent binding of long DNA to protein-sized quan- 73 33. 38. Liedl T. Natl. Laakkonen P. Wang YA. Recognition molecule directed interfacing between semiconductor quantum dots and nerve cells. Soc. Temperature. Status Solidi B-Basic Res. 49. et al. 2002. 45. Chem. 74:841–47 Goldman ER. Am. 123:4103– 4 Wang YA. Li JJ. et al. Chem. Conjugation chemistry and bioapplications of semiconductor box nanocrystals prepared via dendrimer bridging. Chem. Tran PT. Am. 280:128– 36 Mahtab R. 122:12142–50 Mattoussi H. Chen DY. 122:14–17 Akerman ME. 2005. . Thompson ME. Anal. Kuno MK. Bhatia SN. Quantum dot-labeled trichosanthin. Self-assembly of CdSe-ZnS quantum dot bioconjugates using an engineered recombinant protein. 44. et al. Mauro JM. J. Labrenz S. Hydrophobic nanocrystals coated with an amphiphilic polymer shell: a general route to water soluble by Inflibnet N-LIST Programme on 09/03/11. Mauro JM. Charles PT. Hydroxylated quantum dots as luminescent probes for in situ hybridization. Mauro JM.annualreviews. Time-resolved spectral observations of cadmium-enriched cadmium sulfide nanoparticles and the effects of DNA oligomer binding. Goldman ER. 224:277–83 Pellegrino T. J. Soc. Soc. Chem. 2002. Chem. Arnheim N. J. 2001. Moore HP. Murphy CJ.

Wong EY. 2003. 77:311–16 Levsky JM. Beuthan J. 2003. Heintzmann R. Bruchez MP. Anal. 2004. Optical coding of mammalian cells using semiconductor quantum dots. Gerion D. Anal. Mattheakis LC. Singer RH. Dvorak J. et al. Cancer Inst. et al. Post JN. Pellegrino T.S. Soc. 1977. 126:6520– 21 59. Cell Sci. Luccardini C. 2004. Zanchet D. On the side effects of endocytosis in the subviral region. 22:198– 203 58. 47:3239–45 Kramer RH. Wong J. Taira K. Chem. Derfus AM. Le Gros M. Biotechnol. 2005. Xu Y. 65. et al. Nano Lett. For personal use only. Gu W. J. Multiplexed SNP genotyping using the Qbead system: a quantum dot-encoded microsphere-based assay. Fluoresc. et al. Parak WJ. Natl. Basement membrane and the invasive activity of metastatic tumor cells. Sha MY. Kanamori T. Sera T. 75:4766–72 Han MY. Annu. The phagokinetic tracks of 3T3 cells: parallels between the orientation of track segments and of cellular structures which contain actin or tubulin. 2003. 13:503–7 54. Diffusion dynamics of glycine receptors revealed by single-quantum dot tracking. Cancer Res. 2004. Dahan M. Mater. Martin GR. Invasion of reconstituted basement membrane by metastatic human tumor cells. Bensch KG.74 ALIVISATOS GU LARABELL mammary epethelial acini. Iwamoto Y. 46:1980–89 Terranova VP. 327: 200–8 60. 1987. Dressler C. Gao by Inflibnet N-LIST Programme on 09/03/11. Probing the cytotoxicity of semiconductor quantum dots. Uphoff J. 1978. A quantum dot conjugated sugar ball and its cellular uptake. Quantum dot-based cell motility assay. Proc. J. Kleinman HK. Riveau B. 69. 63. 1994. Biochem. Differentiation 71:542–48 Albini A. 66. Engel B. 2004. Boudreau R. 2003. Choi YJ. J. Chen FQ. Quantum-dot-tagged microbeads for multiplexed optical coding of biomolecules. Biomed. U. Gong J. Science 302:442–45 57. 2002. Fu AH. Microsc. 1986. Kannan B. et al. 64. Sci. 31:e43 Van Der Meer BW. New York: VCH Stryer L. Cell 11:395–404 Pellegrino T. Cell motility and metastatic potential studies based on quantum dot imaging of phagokinetic tracks. Dias JM. Nucleic Acids Res. Parak WJ. Breast tumor cells undergo apoptosis upon contact with 61. 14:241–47 56. Kaul SC. . Downloaded from www. J. 14:882– 85 52. Hujanen ES. Am. Nagy P. Cell Res. Osaki F. et al. Chem. ArndtJovin DJ. Martin GR. 1986. 2004. 71. Heat stress induced redistribution of fluorescent quantum dots in breast tumor cells. Mortalin imaging in normal and cancer cells with quantum dot immuno-conjugates. Yaguchi T. Fluorescence energy 51. Boudreau R. Room-temperature single-nucleotide polymorphism and multiallele DNA detection using fluorescent nanocrystals and microarrays. Nat. Gerion D. 211:256–61 55. Boudreau R. Fluorescence in situ hybridization: past. Aoyagi Y. Rostaing P. present and future. 116:2833–38 Gerion D. et al. 2001. J. Parak WJ. Quantum dot ligands provide new insights into erbB/HER receptor-mediated signal transduction. Parak WJ. 2003. Resonance Energy Transfer: Theory and Data. Araonson SA. 19:631–35 Xu H. Cancer Res. Minet O. 1977. 70. Biotechnol. Acad. Bhatia SN. Lidke DS. 2003. Chan WCW. Levi S. 72.A. Coker G III. Tokumasu F. Kaul Z. The phagokinetic tracks of 3T3 cells. Adv. 62. Su JZ. Nat. 2004. Sando S. Submitted Albrecht-Buehler G. 68. Natl. Wadhwa R. Nie S. Development and application of quantum dots for immunocytochemistry of human erythrocytes. 67. Cell 12:333–39 Albrecht-Buehler G. Chen S-YS. 4:11– 18 53. Le Gros MA. 2003. Hirano T. A rapid in vitro assay for quantitating the invasive potential of tumor cells. Eng. Rev.annualreviews. et al. Triller A.

Am. Matt. Rogach AL. Mokari T. 15:79–86 Kim S. Waggoner AS. Bruchez MP. 2001. J. 2003. Chem. Willner I. 77. Chem. Phys. Mattoussi H. Mauro JM. Chem. 2004. Medintz IL. Rosenzweig Z. Phys. Biotechnol. Chem. 2002. 1998. Nano Lett. Chem. Eychmuller A. Gill R. 91. Phys. Kirstein S. 2004. Applications of Tlymphoma labeled with fluorescent quantum dots to cell tracing markers in mouse body. Biochem. 76:1517–20 Kagan CR.QUANTUM DOTS AS CELLULAR PROBES transfer as a spectroscopic ruler. Patel K. Trammell SA. Enhanced Forster energy transfer in organic/inorganic bilayer optical microcavities. J. Biophys. 80. Methods 58:59– 66 Spanhel L. Lu Z. AM. 2004. Luminescent CdS quantum dots as selective ion probes. 54:8633–43 Finlayson CE. Res. Rosenzweig N. 1996. structure. Reversible modulation of quantum dot photoluminescence using a protein-bound photochromic fluorescence resonance energy transfer acceptor. BCondens. 79. Soltesz EG. Clapp AR. Anal. Yamamoto K. Suzuki K. Trewyn BG. Ernst LA. J. Annu. Surface modification and stability of strong luminescing CdS particles. Fluorescence resonance energy transfer between quantum dot donors and dyelabeled protein acceptors. Lee J. 2004. Murray CB. Mattoussi H.7:55-76. Strongly photoluminescent CdTe nanocrystals by proper surface modification. Mauro JM. Soc. Nano Lett. Biochem. 75. 90. Murphy CJ. Henglein A. Soc. Weller H. 2001. 1996. 2005. 82. Weizmann Y. Chem. Lighting-up the dynamics of telomerization and DNA replication by CdSe-ZnS quantum dots. Bioconjug. Superparamagnetic Fe2O3 beads-CdSe/ZnS quantum dots core-shell nanocomposite particles for cell separation. 74:5132–38 Wang DS. Eng. et al. Rosenzweig Z.annualreviews. Rev. 86. Q-CdS photoluminescence activation on Zn2+ and Cd2+ salt introduction. et by Inflibnet N-LIST Programme on 09/03/11. Mauro JM. 4:409–13 Lai CY. Chem. Goldman ER. 89. 83. 2003. Photochemistry of colloidal semiconductors. Lett. 81. Nat. Soc. 2003. Fisher B. Rev. A mesoporous silica nanosphere-based carrier system with chemically removable CdS nanoparticle caps for stimuli-responsive . 78. Albumin-CdTe nanoparticle bioconjugates: preparation. Commun. Long-range resonance transfer of electronic excitations in close-packed CdSe quantum-dot solids. Nirmal M. Greenham NC. Downloaded from www. 2001. Chem. Noninvasive imaging of quantum dots in mice. 2:630–38 Clapp AR. 2004. Kornowski A. Ou J. Am. Xie Z. Angel SM. Cullum BS. J. 76. Photophysical properties of ZnS nanoclusters with spatially localized Mn2+ . Self-assembled nanoscale biosensors based on quantum dot FRET donors. Xu S. 126:301–10 Medintz IL. Mattoussi H. Near-infrared fluorescent type II quantum dots for sentinel lymph node mapping. 125:13918–19 Ballou B. Fisher BR. Biomed. 126:30–31 Patolsky F. 85. Kotov NA. Lagerholm BC. Banin U. Xie H. Mater. 47:819–46 Kagan CR. B. 314:46–53 Li W. For personal use only. Nat. and interunit energy transfer with antenna effect. et al. 109:5649– 55 Sooklal K. Rev. Biophys. Phys. Bawendi MG. Soc. 87. Hanaki K. Electronic energy transfer in CdSe quantum dot solids. Jeftinija K. Murray CB. et al. 2004. Bawendi MG. 20. Exploring the mechanism of competence development in Escherichia coli using quantum dots as fluorescent probes. 22:93–97 Hoshino A. Bawendi MG. Annu. Hasse M. Jeftinija DM. Phys. Rogach AL. 2004. 1996. Biochem. Studer J. Lett. 74. Langmuir 17:2541– 44 Gao M. J. 100:4551–55 Moore DE. Chem. De Grand 75 73. 88. Lim YT. Ginger DS. Rev. 102:8360–63 Chen YF. J. 1:281–86 Medintz IL. J. Am. 84. Am. He JB. 338:83– 87 Mamedova NN. 1987.

76 ALIVISATOS GU LARABELL filtered transmission electron microscopy. Nair A. J. High resolution protein localization using soft X-ray microscopy. Am. Lelievre SA. J. 125:4451–59 92. Histochem. Microsc. . Nisman R. 52:13–18 93. Downloaded from www. conventional.annualreviews. 201:395–403 controlled release of neurotransmitters and drug molecules. 2004. For personal use only. et al. Eng. 2005. Ren Y. Cytochem. Soc. and energy- by Inflibnet N-LIST Programme on 09/03/11. Denbeaux G. J.7:55-76. Dellaire G. Li R. Meyer-Ilse W. Biomed. Chem. Bazett-Jones DP. Rev. Hamamoto D. 2001. Application of quantum dots as probes for correlative fluorescence.

org by Inflibnet N-LIST Programme on 09/03/11. Figure 1 Emission spectra of several semiconductor nanocrystals showing their size. Biomed. The figure is reprinted with permission from Reference 12. and blue series represent different-sized InAs.and composition-dependent emission character. respectively. For personal use only. copyright 1998 AAAS.QUANTUM DOTS AS CELLULAR PROBES C-1 Annu. The sizes of the nanocrystals are indicated above their corresponding spectra.annualreviews. green. . InP. and CdSe nanocrystals. Eng. Red. 2005. Downloaded from www.7:55-76. Rev.

The figure was reproduced with permission from Reference 60. (C) The contact was fatal to the tumor cells. Rev. After 14–16 h of incubation. (B) After the acini were formed. . it is a superimposition of all sections.7:55-76. leaving transparent ghosts loosely attached to the organoid. the tumor cells had attached to the acini’s basement membrane. Figure 5 QDs were used to study mixed cell interactions in a 3-D matrigel culture system. which were found dead surrounding the MCF 10A organoid. The scale bar is 10 m. displaying the sharp edge of each cell followed by a projection of color-coded depth information so that red is the uninvolved lower portion of the MCF-10A organoid and the tumor cells are shades of orange through green. Biomed. Eng. (D) The MCF-10A organoid and all invading tumor cells. human breast tumor MDAMB-231 cells (tagged with red-emitting silica coated QDs) were added to the culture. Downloaded from www. For personal use only.annualreviews.C-2 ALIVISATOS ■ GU ■ LARABELL Annu. (A) Human mammalian epithelial MCF 10A cells (tagged with green-emitting silica coated QDs) form acini structures after growing in growth-factor reduced matrigel for 10 days. Most of the tumor cells had lysed. 2005. but a few newly attached cells still retained red-emitting by Inflibnet N-LIST Programme on 09/03/11.

7:55-76. (a) Sensitivity comparison between QD-tagged and green fluorescence protein (GFP) transfected cancer cells. For personal use by Inflibnet N-LIST Programme on 09/03/11. Eng. Downloaded from www. whereas the same number of GFP-labeled cells (green) were injected on the left flank (circle) of the same animal. Figure 7 In vivo imaging using QDs. 2005. The figure was reprinted with permission from Reference 21. . Biomed.annualreviews.QUANTUM DOTS AS CELLULAR PROBES C-3 Annu. Rev. which were injected at three adjacent locations on a host animal. (b) Simultaneous in vivo imaging of multicolor QD-encoded microbeads. QD-labeled cells (orange) were injected on the right flank of a mouse.

and Carolyn Larabell BLOOD-ON-A-CHIP. Morse FUNCTIONAL ELECTRICAL STIMULATION FOR NEUROMUSCULAR APPLICATIONS. Simon and Chad E. A. and James W. Mehmet Toner and Daniel Irimia BIOCHEMISTRY AND BIOMECHANICS OF CELL by Inflibnet N-LIST Programme on 09/03/11.annualreviews. Stanley A. Mielke QUANTUM DOTS AS CELLULAR PROBES. King. 2005.7:55-76. Paul Alivisatos. Craig v . Benham and Steven P. Weiwei Gu. James D. Green 151 187 223 255 287 327 361 DETERMINISTIC AND STOCHASTIC ELEMENTS OF AXONAL GUIDANCE. Biomed. Rev.Annual Review of Biomedical Engineering Volume 7. and Shu Chien MOLECULAR MECHANICS AND DYNAMICS OF LEUKOCYTE RECRUITMENT DURING INFLAMMATION. Holmes. and Mark S. Jun-Lin Guan. Volumes 1–7 403 413 416 ERRATA An online log of corrections to Annual Review of Biomedical Engineering chapters may be found at http://bioeng. Knutson RETINAL PROSTHESIS. and Jack L. Robert J. Song Li.annualreviews. P. For personal use only. 2005 CONTENTS Annu. Eng. Lewis xii 1 21 55 77 105 DNA MECHANICS. Volumes 1–7 Cumulative Index of Chapter Titles. Wentai Liu. Thomas K. Weiland. Albert I. Borg. Werner Goldsmith WERNER GOLDSMITH: LIFE AND WORK (1924–2003). Covell INSTRUMENTATION ASPECTS OF ANIMAL PET. Yuan-Chuan Tai and Richard Laforest IN VIVO MAGNETIC RESONANCE SPECTROSCOPY IN CANCER. FRONTISPIECE. Gillies and David L. Jeffrey W. Downloaded from www. Humayun INDEXES Subject Index Cumulative Index of Contributing Authors. Berger. Hunter Peckham and Jayme S. Scott I. Susan Maskery and Troy Shinbrot STRUCTURE AND MECHANICS OF HEALING MYOCARDIAL INFARCTS.

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