Thermal Lens Microscope TLM

FIGURE 7.20 Schematic illustration of the thermal lens measurement. The excitation beam was focused by the objective lens. After excitation of some analytes, their radiationless relaxation caused the thermal effect and the formation of a concave thermal lens. The probe beam, which was also coaxially focused by the same objective lens, was collected by the photodiode detector defined by the pinhole. Any change in the amount of heat produced by the radiationless relaxation is manifested as the change in the photodiode output as a TLM signal [733]. Reprinted with permission from Elsevier Science. 

The TLM detection method has been applied to detect labeled amino acids. For instance, after separation in a conventional capillary, DABSYL-labeled amino acids were detected in a Pyrex TLM detector chip. The LOD was 4.6 x l() 8 M, as compared to the LOD of 5.2 X 10-6 M obtained by the conventional absorbance method [343]. TLM has also been applied for the analysis of metal ions such as Co [319,734], Ni [735], Pb [736], Fe [734], and of organic molecules such as o-toluidine after its oxidation [737], 

The TLM method produces a spatial resolution of 1 [tm. Another advantage of TLM is that it is not strongly affected by light scattering (e.g., by the cell membranes in biological systems) [851], It has been demonstrated that the TLM method has a detection limit at the single-molecule level [425], 

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We have developed a novel ultrasensitive detection method, thermal lens microscopy (TLM), for nonfluorescent species [13]. TLM is photothermal spectroscopy under an optical microscope. Our thermal lens microscope (TLM) has a dual-beam configuration excitation and probe beams [13]. The wavelength of the excitation beam is selected to coincide with an absorption band of the target molecule and that of the probe beam is chosen to be where the sample solution (both solvent and solute) has no absorption. For example, in determination of methyl red dye in water, cyclohexane, and n-octanol, a 514-nm emission line of an argon-ion laser and a 633-nm emission line of a helium-neon laser were used as excitation and probe beams, respectively [21], Figure 4 shows the configuration and principle of TLM [13]. The excitation beam was modulated at 1 kHz by an optical chopper. After the beam diameters were expanded, the excitation and probe beams were made coaxial by a dichroic mirror just before they were introduced into an objective lens whose magnification and numerical aper-... 

Figure 4 Operating principles of thermal lens microscope (TLM).

Thermal lens microscopy (TLM) is a type of photothermal spectroscopy. TLM depends on the coaxial focusing of the excitation and probe laser beams (see Figure 7.20). Which is achieved using the chromatic aberration of a microscopic objective lens [731]. The excitation beam can be provided by a YAG laser (532 nm) [846,1021] or an Ar ion laser (514.5 nm [846] or 488 nm [732]).The probe beam can be provided by a He-Ne laser (632.8 nm) [846,1021], After optical excitation of the analyte molecules, radiationless relaxation of the analytes occurs,... 

Figure 3 MUOs and CFCP. Abbreviation TLM, thermal lens microscope (Tokeshi et al., 2002).

In order to solve these problems, differential interference contrast thermal lens microscope (DIC-TLM) was developed. The principle of DIC-TLM is shown in Fig. 8. The probe beam is separated into two beams and integrated again using a pair of DlC prisms. Contrary, the excitation beam is not separated and induces a local change in refractive index only for the one of the separated probe beams. Thus, phase contrast appeared between the probe beams is detected through an interference. 

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