Few-Atom Silver Clusters as Fluorescent Reporters - Advanced Fluorescence Reporters in Chemistry and Biology - page 324

affected during the reduction of silver either by a chemical reductant or by an
irradiation source. For example, silver salts can be easily reduced by sodium
borohydride, but this reductant may also reduce other functional groups, includ-
ing disulfide bonds to sulfhydryl groups and could destabilize the folding and
function of proteins.
To overcome the difficulty of finding a proper scaffold, which provides the
required environment for the formation of silver clusters being at the same time
stable during reduction of silver, Dickson et al. proposed a new approach where
silver clusters are first prepared using a polymer and then transferred, or shuttled, to
the desired biomolecule [ 69 ]. These two steps for labeling of cellular components
introduce additional advantages to those already mentioned. For instance, the
production of silver clusters can be optimized in the first step by adding 3-(2-
aminoethylamino)propyltrimethoxy silane (APTMOS) to complex silver ions
before stabilization with the proper organic polymer, in this case poly(acrylic
acid) (PAA) [ 69 ]. The silver clusters prepared in this way have tenfold increased
emission intensity (Fig. 11b ) and the formation of nanoparticles, was eliminated. In
the second step, the transfer efficiency of fluorescent clusters to high-affinity
ssDNA sequences (oligocytosines) can be as well optimized for the specific
sequence by adjusting the temperature, the pH and the buffer in the transfer step
and by varying the ratio of Ag + /APTMOS in the reduction step.
The shuttle of silver clusters from PAA to oligocytosine due to the stronger
interaction of clusters with oligonucleotides was confirmed by a pronounced shift in
the excitation spectra from 515 to 570 nm (Fig. 11b ). The specific conditions
required in each system for the successful shuttle of silver clusters represent an
enhancement in the selectivity of the labeling process. Using the cluster transfer, it
is possible to produce silver clusters in a polymer and then label ssDNA or
oligopeptides [ 77 ]. In a similar manner, antibodies conjugated with ssDNA were
labeled with fluorescent silver clusters and then the antibody-DNA-Ag clusters
were applied to tag live cells. Incubation of live cells at 4 C results in cell-surface
staining (Fig. 11c ) whereas incubation at 37 C results in internalization of silver
clusters and staining inside the cells (Fig. 11d ).
The concept of dynamic silver clusters capable to transfer between molecules
was also pointed out recently by Ras et al. for silver clusters prepared by photo-
activation using PMAA as scaffold [ 20 ]. Every specific initial ratio of silver ions to
methacrylate unit, Ag + :MAA, results in distinct spectral bands (Fig. 12a , b). Thus,
an initial ratio 0.5:1 gives an absorption band at

503 nm, whereas a ratio 3:1 gives

a band at 530 nm. The shuttle effect was proven when for a given silver cluster

solution with ratio 3:1 and absorption at 530 nm, a blue shift was achieved by the
addition of pure PMAA. For instance if the added amount of polymer decreases
the ratio Ag + :MAA from 3:1 to 0.5:1, the new optical band will match exactly with
the band corresponding to a solution with initial ratio 0.5:1, that is 503 nm
(Fig. 12c ). The explanation given for this blue shift was the redistribution of
the existent silver clusters in PMAA chains over the newly available PMAA
chains, in other words that the clusters shuttle from partly clusters-filled chains
to empty ones.