Optical arrangements for single molecule detection

Next we shall describe these two common experimental geometries, but before we proceed we re-emphasize the key considerations of the experiment to maximize the signal-to-noise  

Clearly, single-photon far-field excitation in this manner provides no spatial reduction of the excitation/collection volume in the direction of propagation of the light and so additional optics are required in the detection path to minimize 

One-and two-photon epi-fluorescence permit similar experiments to be carried out, although one may have practical advantages over the other [19]. 

2 The PSF in single-photon confocal and two-photon epi-fluorescence illumination systems 

In confocal microscopy the PSF is a function of a number of physical components, namely the laser beam, the microscope objective, the confocal pinhole, and the detector. The PSF is a convolution of the effect of all of these components. Despite this apparent complexity it is perhaps surprising that a simple description of the PSF for these microscopes has been widely applied in which the sample volume is described by a three-dimensional Gaussian with a 1/e beam waist diameter 2coq and a length 2zq along the optic axis [6,20,21 ] and is given by, 

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Figure 9. (a) Diagram of the hyperbolic electromagnetic trap for levitating microdroplets in a laser beam. (b) Optical arrangement for single-molecule detection in microdroplets lens, L beam splitter, BS polarizer, P trap, T microscope objective, O optical filters, F spatial filter, SF photomultiplier, PMT desiccant chamber, D. (Adopted from [51].)... 

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