Scientists keen on monitoring cellular events in real time can now add another imaging technique to their repertoire-the use of quantum dots.
Quantum dots (QD) are tiny nanoparticles of fluorescent inorganic semiconductors. About 10 nm in diameter, they are much larger than conventional organic fluorophores, but have superior characteristics, including resistance to degradation and very sharp emission spectra, the latter ensuring that several QDs can be measured simultaneously without cross-interference.
The widespread use of these fluorophores has been hindered, however, by some major obstacles, including difficulty coupling them to biological molecules and the question of toxicity. As reported in today’s Nature Biotechnology, two groups have successfully crossed these hurdles, using CdSe/ZnS nanocrystals to monitor subcellular organelles, and to label individual proteins.
Researchers from The Rockefeller University, New York, and the U.S. Naval Academy, Washington, DC, led by Sanford Simon, show that whole cells, either mammalian HeLa cells or cells from the slime mold Dictyostelium discoideum, can take up quantum dots by a variety of mechanisms including endocytosis, but do not suffer any deleterious effects. Furthermore, the amoeba cells responded normally to developmental stimuli and could be monitored for up to 18 hours with no adverse effects (See the Simon lab for time lapse video-Real Player required). In addition, the stability of quantum dots was demonstrated by the ability to monitor human cells for up to 12 days with no loss of signal.
First author Jyoti Jaiswal et al. were also able to label specific proteins by employing the immunoglobulin-binding power of avidin-conjugated dots. These conjugates were used to detect antibodies bound to the cell surface transporter, P-glycoprotein, in HeLa cells. The specificity of this detection method was proven by comparing quantum dot images with those of P-glycoprotein/green fluorescent protein chimeras in the same cell.
This type of conjugate approach was also used by Marcel Bruchez and colleagues at Quantum Dot Corporation, Hayward, California, and Genentech, San Francisco, to detect the breast cancer marker protein Her2 on live human cell lines, actin filaments in fixed fibroblasts, and nuclear antigens in fixed human epithelial cells.
First author Xingyong Wu et al. also compared the fluorescent intensity of their quantum dots with that of Alexa 568, reputedly the brightest organic fluorophore available. Fluorescence of the quantum dot, measured at the optimum excitation and emission wavelengths for the organic dye, was about four times brighter, while use of the optimum wavelengths for the QD more than doubled this difference. The QD was also much more stable than the organic reporter, which photobleached more rapidly.
"The paper by Wu et al is a lovely study which complements our own," commented Sanford Simon. "In both cases we have been able to show that quantum dots can be used to specifically label proteins for long periods of time." (See also comment below).—Tom Fagan
- Jaiswal JK, Mattoussi H, Mauro JM, Simon SM. Long-term multiple color imaging of live cells using quantum dot bioconjugates. Nat Biotechnol. 2003 Jan;21(1):47-51. PubMed.
- Wu X, Liu H, Liu J, Haley KN, Treadway JA, Larson JP, Ge N, Peale F, Bruchez MP. Immunofluorescent labeling of cancer marker Her2 and other cellular targets with semiconductor quantum dots. Nat Biotechnol. 2003 Jan;21(1):41-6. PubMed.