1
516
F.-F. Li et al. / Chinese Chemical Letters 26 (2015) 1514–1517
Fig. 4. Single molecule catalysis of non-fluorescent molecule HCOONa on Pt/AC
catalyst. (a) TEM image of Pt/C catalyst. (b) Single molecule catalysis of non-
fluorescent molecule in TIRFM. The yellow arrow points out the location where the
reaction happens. (c) Trajectories of same location under the conditions with and
without 0.5 mol/L HCOONa. The HCOONa was dissolved in pH 7.30 phosphate
buffer.
Fig. 3. Working performance of a single microcell. (a) HCOONa electro-oxidation
reaction on working electrode, and fluorescent molecule generation reaction on
couple electrode with active carbon. (b) Fluorescent signal (from 7 Â 7 pixels) from
couple electrode when formic acid oxidation reaction happens on working
electrode. Green and black curves are for the electrodes with and without Pt
nano-catalyst on working electrode. (For interpretation of the references to colour
in this figure legend, the reader is referred to the web version of this article.)
achieve single molecule detection. In order to improve the signal
to noise ratio, we simplify the design in Figs. 1c and 2. We
simply use active carbon (AC) supported Pt nanoparticle as a
redox system. Since AC is not active to the electro-oxidation of
non-fluorescent molecule HCOONa, but active to the electro-
reduction resazurin, AC could be used to detect the single
molecule electro-oxidation of HCOONa on Pt nanoparticle.
Therefore, the isolator and Au wire were removed from the
design. The AC acts as the couple electrode, conductor and
catalyst for catalytic conversion from resazurin to resorufin.
Fig. 4a shows the TEM image of Pt/C catalyst with ꢀ3 nm size.
We directly immobilize Pt/C catalyst onto quartz slide to do
strong fluorescent signal, when HCCONa was flowed into the
flow cell. These two reactions couple each other. But the
fluorescent signal would disappear when the flow cell was
washed by water. In the control experiment, the electrode
without Pt nanoparticles has no signal at the same condition. So
our design works very well.
In addition, Fig. 3b shows some information of the stability of
the Pt nano-catalyst on the working electrode. As shown in Fig. 3b,
the signal keeps decreasing when HCOONa was flowed into the
flow cell. So the activity of Pt nano-catalyst keeps decreasing with
time due to the poisoning of CO-like intermediates. This result is
very similar as that from the measurement of conventional
technology. This simple method is estimated to measure the
single molecule catalysis of
a
non-fluorescent molecule
HCOONa. Since the oxidation of HCOONa on Pt and the reduction
of resazurin on active carbon do not disturb each other, the
solution of HCOONa and resazurin can be feed into the
microfluidic reactor together. Fig. 4b shows that the product
resorufin was detected when the solution including HCOONa
and resazurin was flowed into the microfluidic reactor. We can
see a lot of fluorescent signal from single product resorufin
molecules in Fig. 4c. In order to make sure that the generation of
product resorufin is coupled with the oxidation of HCOONa, a
control experiment without HCOONa was done. The trajectory of
control experiment in Fig. 4c shows that no product resorufin
was detected. As a result, this research shows that we can detect
the oxidation reaction of one HCOONa molecule through this
method. This paper also provides a general way to study the
SMECNFM for the molecules, such as formic acid, hydrogen,
oxygen, etc., on single nanoparticle.
À14
4
current as low as 10
A, which means we have detected ꢀ10
fluorescent molecules per second (or turnovers of HCOONa
electro-oxidation reaction).
As we all know that the electrochemical reactions like formic
acid oxidation, methanol oxidation, hydrogen oxidation and
oxygen reduction are very important in real applications. But
these reactions do not include any fluorescent molecule. So these
reactions cannot be studied by the previous research strategy
which needs the fluorescent molecule to take part into the target
catalytic reaction directly. In this paper, in order to study these
reactions at the single molecule/nanoparticle level, we couple
these reactions with a fluorescence reaction. Then we can study the
SMECNFM by measuring the fluorescence reaction on couple
electrode. Here, we use resazurin reduction reaction to generate
highly fluorescent resorufin. By detecting each single resorufin
molecule, we can detect the formic acid electro-oxidation at single
molecule level. This method is actually a general method, which
can be used to measure the single non-fluorescent molecule
reactions, as long as we have a fluorescence reaction with proper
redox potential.
4. Conclusion
In this paper, we use the concept of microcell in electrochemical
corrosion science to study the electrochemical reaction of non-
fluorescent molecule (SMECNFM) on nano-catalyst. In the micro-
cell, the SMECNFM is coupled with a fluorescence reaction. The
SMECNFM has strict stoichiometric relationship with the fluores-
cence reaction according to Faraday’s Law of Electrolysis. Then, the
SMECNFM is detected accurately by detecting the fluorescent
molecules. In this paper, we studied the oxidation reaction of one
HCOONa molecule through this method. This paper also provides a
3.2. Single molecule catalysis of non-fluorescent molecule
In principle, the above strategy is able to study the single
molecule catalysis of non-fluorescent molecule on nanoparticle.
However, the configuration in Fig. 2 has too high noise to