ABSTRACT
In the fast-evolving eld of nano-biosciences, simple and sensitive methods for the detection and characterization of the matter at the nanoscale are needed. Electron microscopy or near-eld methods such as scanning tunneling microscopy and atomic force microscopy provide atomic resolution but need extensive sample preparations and are not fully suitable in complex environments such as biological matter. Far-eld optical methods have the advantage to be noncontact and minimally invasive. e most commonly used optical techniques are based on luminescence. e ultimate sensitivity in the optical imaging of live cells is achieved by single-molecule detection (SMD). First, by removing the average inherent to ensemble measurements, SMD yields a measure of the distribution of molecular properties, which is of importance in biological systems that display static or time-dependent heterogeneity. us, subpopulations, even minor ones, can be studied and statistical correlations between dierent distributions of parameters can be performed on dierent subpopulations. As an example, one can cite the study of the diusion of biomolecules in membranes (Schmidt et al. 1995; Sako et al. 2000; Schutz et al. 2000). ey not only have revealed the existence of dierent subpopulations in terms of membrane diusion in the plasma membrane of the live cells but also have permitted to study the characteristics of the diusion of the dierent populations (Vrljic et al. 2002; Tardin et al. 2003) and have even revealed a functional role of this diusion in the case of fast communication between neurons (Heine et al. 2008). Second, SMD gives access to the dynamic uctuations of one parameter and can thus unravel all steps
16.1 Introduction ......................................................................................501 16.2 Detection of Nonuorescing Nano-Objects .................................502 16.3 Photothermal Interference Contrast Method ..............................503 16.4 Photothermal Heterodyne Imaging ...............................................504 16.5 Single Metal Nanoparticle Absorption Spectroscopy .................505 16.6 DNA Microarrays .............................................................................507 16.7 Single-Nanoparticle Tracking in Live Cells .................................508 16.8 Photothermal Absorption Correlation Spectroscopy .................510 16.9 Conclusions........................................................................................ 511 Acknowledgments ........................................................................................512 References ......................................................................................................512
of its temporal evolution without the need to synchronize all molecules (which is in practice impossible for live organisms). Keeping with the previous example in neurobiology, the existence of exchanges of receptors between dierent cellular compartments such as synaptic and extrasynaptic spaces of live neurons has been revealed for the rst time (Tardin et al. 2003). Finally, SMD also provides an important advantage over ensemble measurements, namely, the possibility to track molecules with a position accuracy only limited by the signal-to-noise ratio at which the molecules are detected. is renders nanometer localizations possible far below the optical resolution (Schmidt et al. 1996; Yildiz et al. 2003). is superresolution feature is now widely applied to obtain superresolved images using photoactivable molecules (Betzig et al. 2006; Rust et al. 2006).
