## 2 Brief History of TERS  
Normal optical microscopy can only achieve a resolution of 200 nm in the visible region owing to limitations caused by optical diffraction, which can be expressed by [PERSON]'s equation as follows:  
\[\Delta x=0.61\lambda/NA \tag{1}\]  
Another disadvantage of optical microscopy is that only an average signal from the target molecules can be obtained. Hence, extensive investigations have been performed to improve spatial resolution through the use of techniques such as scanning near-field optical microscopy (SNOM) with a small aperture [40] and scanning tunneling microscopy (STM) with a narrow tip [41]. By combining these technologies [7; 42], four groups are dedicated to developing the spatial resolution of TERS [25; 26; 27; 28]. To make the best use of TERS, it is crucial to develop a method that can easily focus the incident laser on the apex of the tips while simultaneously maintaining the intensity and sensitivity of the signal. As shown in Figure 1, TERS is based on SPM technologies such as STM, atomic force microscopy (AFM), shear force microscopy (SFM) and scanning near-field optical microscopy (SNOM). Using a sharp metal tip (approximately 10-50 nm) and when the focused laser illuminates the apex of the metal or metal-coated tip, a 'hot spot' can be excited in the vicinity of the tip apex due to LSPR [37; 43]. Furthermore, as shown in Figure 1a, the intensities of the Raman signals can be enormously enhanced. The most prominent advantage of TERS compared to SERS is the position of the hot spot and the fact that the signal enhancement factor can be effectively controlled by moving the tip and monitoring the distance between the tip and substrate through a feedback system. As previously introduced, SERS usually requires rough metal surfaces or aggregations of metal nanoparticles to create the hot spot. This roughness is difficult to control and will ultimately lead to randomness in the experimental results. TERS can overcome one of the most severe restrictions in the application of SERS by providing higher accuracy. The spatial resolution of TERS can reach a few nanometers [44; 45], even down to the sub-molecular level [46]. With the STM technology shown in Figure 1b, the spatial resolution can reach an atomic level of 0.1 nm and the distance between the tip and substrate can be accurately manipulated by adjusting the tunneling current and sensitive feedback system. The marriage between STM and a ultrahigh vacuum can improve the performance of TERS. However, this type of TERS system is limited by the need for conductive substrates and thin samples. AFM and SFM are also suitable for use with TERS, as shown in Figure 1c,d [47]. With AFM measurements [48], the samples are not limited and a surface roughness on the micrometer level is acceptable [49]. AFM-TERS can be applied in aqueous environments and may become a suitable technique for biological investigations [35; 50; 51; 52; 53]. However, the spatial resolution is one nanometer and the tip must be coated with a noble metal, which can easily break during measurements. It should be noted that a dielectric coating on the tip can prevent contamination from the environment or target species, but the coating decreases the plasmonic enhancement. The coating is limited to the nanometer scale (1-2 nm) and must be pinhole-free. For the SFM-TERS shown in Figure 1d, a metal tip is attached to a high-quality tuning fork as an ultrasensitive feedback system [54; 55]. The resulting device is suitable for investigations in aqueous environments as with AFM [56; 57], however, more stable tips for SFM are made of metal rather than coated [58; 59]. On the other hand, the resolution of SFM is limited by the tuning fork [58; 60]. The SNOM shown in Figure 1e is designed to investigate evanescent waves on a surface and includes two types of feedback systems. These are: Constant force (AFM) and shear force (SFM). The resolution of SNOM is strongly dependent on the size of the detector aperture rather than the laser.