Traditional light microscopy suffers from the diffraction limit, which limits the spatial resolution to λ/2. The current trend in optical microscopy is the development of techniques to bypass the diffraction limit. Resolutions below 40 nm will make it possible to probe biological systems by imaging the interactions between single molecules and cell membranes. These resolutions will allow for the development of improved drug delivery mechanisms by increasing our understanding of how chemical communication within a cell occurs. The materials sciences would also benefit from these high resolutions. Nanomaterials can be analyzed with Raman spectroscopy for molecular and atomic bond information, or with fluorescence response to determine bulk optical properties with tens of nanometer resolution.;Near-field optical microscopy is one of the current techniques, which allows for imaging at resolutions beyond the diffraction limit. Using a combination of a shear force microscope (SFM) and an inverted optical microscope, spectroscopic resolutions below 20 nm have been demonstrated. One technique, in particular, has been named tip enhanced near-field optical microscopy (TENOM). The key to this technique is the use of solid metal probes, which are illuminated in the far field by the excitation wavelength of interest. These probes are custom-designed using finite difference time domain (FDTD) modeling techniques, then fabricated with the use of a focused ion beam (FIB) microscope. The measure of the quality of probe design is based directly on the field enhancement obtainable. The greater the field enhancement of the probe, the more the ratio of near-field to far-field background contribution will increase. The elimination of the far-field signal by a decrease of illumination power will provide the best signal-to-noise ratio in the near-field images. Furthermore, a design that facilitates the delocalization of the near-field imaging from the far-field will be beneficial.;Developed is a novel microscope design that employs two-photon non-linear excitation to allow the imaging of the fluorescence from almost any visible fluorophore at resolutions below 30 nm without changing filters or excitation wavelength. The ability of the microscope to image samples at atmospheric pressure, room temperature, and in solution makes it a very promising tool for the biological and materials science communities. The microscope demonstrates the ability to image topographical, optical, and electronic state information for single-molecule identification. A single computer, simple custom control circuits, field programmable gate array (FPGA) data acquisition, and a simplified custom optical system controls the microscope are thoroughly outlined and documented. This versatility enables the end user to custom-design experiments from confocal far-field single molecule imaging to high resolution scanning probe microscopy imaging.;Presented are the current capabilities of the microscope, most importantly, high-resolution near-field images of J-aggregates with PIC dye. Single molecules of Rhodamine 6G dye and quantum dots imaged in the far-field are presented to demonstrate the sensitivity of the microscope. A comparison is made with the use of a mode-locked 50 fs pulsed laser source verses a continuous wave laser source on single molecules and J-aggregates in the near-field and far-field. Integration of an intensified CCD camera with a high-resolution monochromator allows for spectral information about the sample. The system will be disseminated as an open system design.
展开▼
