Quantitative characterization of light-harvesting efficiency in single molecules and nanoparticles by 2D polarization microscopy: Experimental and theoretical challenges R. Camacho, D. Thomsson, D. Yadav, I.G. Scheblykin ⇑ Chemical Physics, Lund University, Box 124, 22100 Lund, Sweden article info Article history: Available online 9 March 2012 Keywords: Light-harvesting Antenna Single molecule spectroscopy Anisotropy Fluorescence FRET Energy funnelling abstract General problem of extracting intramolecular energy transfer information from fluorescence and fluores- cence excitation polarization experiments at single molecule level is presented. A single funnel approx- imation is shown to be a very powerful approach to model the polarization data obtained by recently emerged 2-dimensional polarization single molecule imaging technique [O. Mirzov et al., Small 5 (2009) 1877]. Using this approximation a parameter characterising quantitatively light-harvesting effi- ciency of an individual light-harvesting antenna can be readily obtained. Technical details of 2D polari- zation imaging and practical methods of avoiding polarization artefact in fluorescence microscopy are discussed. Ó 2012 Elsevier B.V. All rights reserved. 1. Introduction Solar light is going to become one of the main sources of energy for human kind. Nature widely uses complex molecules and aggre- gates to collect solar energy. Remarkable examples are the photo- synthesis ‘‘machineries’’ of bacteria, algae and higher plants, where several different molecular structures assembled together in a spe- cial manner in order to use solar energy to run chemical reactions. A light-harvesting antenna is the primary part of any ‘‘device’’ uti- lizing light energy for its operation. A good antenna should have a large absorption cross-section, therefore being an ensemble of many chromophores. The excitation transport should be very effi- cient and directed towards special sites where the energy is to be used. Energy transfer (ET) or excitation transport in such nano-anten- nas like molecular aggregates, conjugated polymers, natural and artificial light-harvesting complexes has been an important topic for years [1,2]. Inhomogeneity of materials has always been a problem masking intrinsic properties of their individual entities (molecules, aggregates and particles) due to ensemble averaging. However, since the end of the 1990-s methods of single molecule spectroscopy (SMS) [3] have become available for studying organi- zation, electronic properties and energy transfer in particular in these systems at single molecule/particle/antenna level [4–7]. Although light-harvesting antennas are indeed molecular ensembles of tens or hundreds of chromophores the antenna’s individual properties do not vanish. This is due to differences in chromophore organizations/packing from antenna to antenna. These differences originate from inhomogeneity of the environ- ment and/or because artificial antennas are often prepared by self-assembling. It is very important that closely packed chro- mophores in an antenna provide a collective response to excitation and therefore they cannot be considered as independent. This ef- fect can be seen as the chromophores can ‘‘talk’’ to each other [8]. Indeed, light absorption, excited state delocalization and en- ergy transfer efficiency depend on interaction between individual chromophores, which in its turn depends on their mutual arrange- ment and the antenna conformation as a whole. Despite large physical size of an antenna there may be just a few places where charge transfer states (or other dark long-living states) are formed. Therefore, the exciton quenching becomes very sensitive to the organization/conformation of the whole system. Classical phenom- ena inherent to single quantum systems like blinking [9,10], spec- tral diffusion [7,11,12] and polarization fluctuations [13–15] have been already reported for several multi-chromophoric antenna systems. All these justify SMS as a very promising technique to study excited state dynamics and organization of light-harvesting antennas. Moreover, geometrical parameters of individual anten- nas such as number of light absorbing [16–18] and light emitting chromophores [19], their orientation and packing [18] and the shape of the whole antenna [20] become accessible when SMS ap- proach is applied. 0301-0104/$ - see front matter Ó 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.chemphys.2012.03.001 ⇑ Corresponding author. E-mail address: Ivan.Scheblykin@chemphys.lu.se (I.G. Scheblykin). Chemical Physics 406 (2012) 30–40 Contents lists available at SciVerse ScienceDirect Chemical Physics journal homepage: www.elsevier.com/locate/chemphys