Principles of Fluorescence Spectroscopy

678 NOVEL FLUOROPHORES
Figure 20.7. Labeling of the Her2 marker on breast cancer cells using
QDots emitting at 535 (left) or 630 nm (right). From [30].
(PC) were injected into Xenopus embryos, with no effect on
their development. 29
QDots can be used for specific labeling of fixed and
live cells. 30–31 Her2 is a cancer marker that is overexpressed
on the surface of some breast cancer cells. Fixed cells were
incubated with monoclonal antibodies against the external
domain of Her2. QDots emitting at 535 nm or 630 nm were
conjugated to anti-IgG antibodies. These protein-coated
antibodies bound specifically to the Her2 markers (Figure
20.7).
Figure 20.8. Photostability comparison between QDots and Alexa
488. Top row: the nuclear antigens were labeled with 630-nm QDots
and the microtubules with Alexa 488. Second row: the microtubules
were labeled with 630-nm QDots and the nuclear antigens with Alexa
488. The graph shows the intensities with continuous illumination.
From [30].
An important property of the QDots is their photostability.
Figure 20.8 shows cells labeled with both QDots and
the photostable fluorophore Alexa 488. In the top row of
images the QDots were localized in the nucleus. In the bottom
row of images the QDots were bound to the microtubules
in the cytoplasm. These images show that the signal
from Alexa 488 is quickly bleached but the QDots remain
fluorescent. These results also show that it is possible to
label internal cellular structures with QDots and that the
intracellular QDots can be highly photostable.
20.1.3. QDots and Resonance Energy Transfer
QDots appear to behave like any other fluorophore with
regard to energy transfer. RET occurs between QDots. 32–39
The schematic in Figure 20.9 shows a system used to test
for RET from a CdSe QDot to a rhodamine acceptor. The
QDot surface was coated with biotinylated BSA. Addition
of streptavidin labeled with tetramethylrhodamine (SAv-
TMR) resulted in quenching of the QDot emission.
Figure 20.9. RET from CdSe QDots to tetramethylrhodamine. The
QDot donor intensity decreased (top to bottom at 540 nm) as the concentration
of TRM-streptavidin increased. Reprinted with permission
from [39]. Copyright © 2001, American Chemical Society.