the dream of the analytical chemistry single molecule...

2Ag, 183 cm­1 3Ag, 343 cm­1 4Ag, 367 cm­1

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Rapid development of Surface­Enhanced Raman Spectroscopy (SERS) techniques is stimulated by prospects of applications insuch fields as ultrasensitive detection, down to the single molecule limit, biomedical diagnostics, or molecular imaging.Understanding the mechanisms of SERS requires experiments in which the parameters relevant for the enhancement are varied ina systematic way. For instance, different surface materials/morphologies can be tried for the same chromophore, or, vice versa, aseries of appropriately modified molecules can be studied for the same support.Porphycene (Pc) was selected as a probe. The choice was justified by the fact that this molecule has been characterized bymethods of vibrational and electronic spectroscopy in our group [1]. The compound is not toxic, thus this single moleculeapplications in biology and medicine seem to be promising in the future.

1. Gawinkowski, S; Walewski, Ł; Vdovin, A; Slenczka, A; Ford, S;Johnson, M; Lesyng, B; Waluk, J; submited2. P. G. Etchegoin, M. Meyer, E. Blackie, and E. C. Le Ru, Anal.Chem. 2007, 79, 8411­84153. Naomi J. Halas, et al., Nano Lett., Vol. 8, No. 4, 2008

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Fig. 1. Structure (left) and absorption spectra (right) of Pc.

Fig. 4. Selected normal modes (from DFT calculation) of Pcwith symmetry and experimental Raman shifts.

Experimental setup

Fig. 5. Possible Pc­nanoparticle configurations.

Fig. 8. SERRS spectra of Pc measured on Au nanoparticlesdeposited on glass, registered from different points of thesample.

Au Au

Fig. 2. A 50x50 µm map of goldnanoparticles on silica support with smalladdition of Pc­d0 end Pc­d12 depositedfrom a low concentration (~10­10 M)ethanol solution.Fig. 3. Pc­d0 and Pc­d12 SM­SERRS spectraobtained from different hot­spots. Au Au

The SERS substrates were prepared by gold colloid (~20nm) drop evaporation on the silicon surface (2 cm2).Next, 10 mm3 of PC/EtOH ~10­10 mol/l solution wasdropped on the SERS substrate and evaporated. TheRaman spectra were taken with an InVia RenishawRaman spectrometer based on Leica microscope with100x or 50x objective. The laser power on the sam plewas kept below 0.1 mW. The spectral resolution was 5cm­1 and the wavenumber accuracy was ±2 cm­1 , bothcalibrated with the Rayleigh line and the 520.6 cm­1 lineof silicon.

Fig. 6. Time evolution of the Pc­d0 SERRS spectra deposited froma low concentration (~10­10 M) ethanol solution (b); timeevolution of the intensity of the 343 cm­1 peak of Pc (c); the first(d) and last (a) spectrum of the evolution. The peak at 520 cm­1 isdue to the support (silica).

Fig. 9. Extinction spectra of laser­ablated Ag (blue) and Au (orange,red) nanoparticles; theoretical extinction spectrum of a goldnanoparticle dimer (pink) [3]; the region of the registered SERRSspectra (green area).

Fig. 10. Time evolution of the intensity of the 343 cm­1 peak of Pcdeposited from "medium" concentration (~10­8 M) ethanol solution(a); SERRS spectra of Pc­d0 at different moments of the temporalevolution (b­d). The peak at 520 cm­1 is due to the support (silica).

Fig. 12. 1:1 mixture of Pc­d0 and Pc­d12 in ethanol. Time evolution ofthe intensity of the 343 cm­1 peak of Pc­d0 (red) and 325 cm­1 peak ofPc­d12 (blue) deposited from "medium" concentration (~10­8 M) (a);SERRS spectra of Pc­d0/Pc­d12 mixture at the beggining (t = 0) of theevolution (b). The arrows show representative peaks of Pc­d0 and Pc­d12 used for observation of the time evolution of the SERRS spectra.

SummaryI. Successful registration of the single molecule SERRS spectra ;)II. Single molecule SERRS spectra represent information aboutthe inner structure of species, as well as information about thelocal environment. The pattern of vibrations appearing in Ramanspectra may be significantly changed by the influence of at leasttwo factors: (1) local plasmon resonance distribution; (2) themolecular orientation with respect to the vector of theelectromagnetic field.III. The registration of time evolution of SERRS spectra allows usto separate spectra of different molecules and located in differenthot­spots at the same focal point.IV. SM­SERS spectroscopy may be used as an analytic qualitativeand quantitative tool.

(a)

(b) Fig. 11. Absolute values of correlation in 100 s time window.

Fig. 7: Histograms of the relative contribution pdye1 = I1 /(I1 + I2 ) to thetotal intensity of dye 1 for (a) N1 = N2 = 10, (b) 1000 molecules.Thehistogram passes from a regime where single molecule events dominatewith two singularities at pdye1 = 0 and pdye1 = 1 (a), to a Gaussiancentered at pdye1 = 0.5 at high concentrations (b) [2].

(a) (b)