Report
NEAR-FIELD FSS s analytical chemists continue U M to push the limits in microscale » 7 1 ^ analysis, the need for new methods capable of probing samples with high spatial resolution and low detection limits has become increasingly important. The explosion in research aimed at reducing sample size, miniaturization and integration of analysis systems onto a single chip, and the massive multiplexing seen in applications such as combinatorial chemistry, all place new demands on the tools available to analytical chemists. For many of these applications optical techniques are attractive given their noninvasiveness single-molecule detection limits high temporal resolution low cost and ease of use However, fundamental restrictions in optical resolution have placed a limit on the usefulness of these techniques for applications requiring high spatial resolution. This limitation motivated the development of higher resolution techniques such as electron-based microscopies or, more recently, the many scanning-probe techniques such as atomic force microscopy (AFM) and scanning tunneling microscopy (STM). Together, these techniques have revolutionized the way we view the "micro world" and have allowed the study of sample erties at the atomic level The unique spectroscopic information and high temporal resolution available with optical techniques, however, provided the necessary incentive to explore new ways of increasing the spatial resolution in optical microscopy. These efforts have culminated in the development of near-field scanning optical microscopy (NSOM or SNOM), which can probe the optical properties of samples at the nanometric level. NSOM has successfully been used to study the spectral properties of single molecules (1-4) as-
SCANNING
OPTICAL MICROSCOPY
The nanometricfluorescenceand force-imaging capabilities of NSOM provide new perspectives onsampleproperties. gregates and thin films (5-9), solid-state devices (10-12), and biological samples (13-17), all with subdiffraction limit spatial resolution. In this Report, we briefly describe the concepts behind NSOM and discuss recent applications of the technique that illustrate its unique capabilities. Basics
In the NSOM technique, high spatial resolution is achieved by scanning a small llght source (or collector) close to a sample surface. The light source is formed with special probes that funnel light down to an aperture that is smaller than the optical wavelength. By positioning the aperture close to a sample, the emerging radiation is forced to interact with the sample before diffracting out.
The spatial resolution in NSOM therefore is only limited by the size of the aperture and its proximity to the sample and not the wavelength of the light, as is the case in conventional microscopy (Figure ea). This elegantly simple idea can be traced back to the early 20th century in a series of papers written by E. H. Synge (18,19). With remarkable foresight, Synge outlined the basic ideas that would eventually lead to the development of NSOM and listed many of the challenges that would have to be overcome to implement the technique. Although simple in concept, the technical Hitoshi Shiku Robert C. Dunn University of Kansas
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Report vance resulted from the introduction of the fiber-optic NSOM probe by Betzig and co-workers in 1991 (23). The majority of NSOM microscopes used today deliver the nanometric spot of light with these fiberoptic probes, which are vastly more efficient than their micropipette forerunners. The probes are fabricated by heating and pulling a single-mode optical fiber down to a fine point (—100 nm). As the fiber narrows, its waveguide properties breakdown and light escapes from the sides, preventing the formation of a well-defined aperture. To circumvent this problem, probes are coated with an opaque metal such as aluminum, which effectively confines the radiation within the probe so that it only exits the very end of the tip enabling high-resolution measurements. The effects of the aluminum coating can be seen by comparing NSOM fluorescence images shown in Figures lb and lc Figure lb shows an image of fluorescent l l'-dioctadecyl-3 3 3' 3'-tetramethylindocarbocyanine perchlorate (diIC ) disnpt-cfvl in a lipirl monolayer taken with an unrnated NSOIVT nmhe RntVi the licrVit exil" Figure 1 . Effects of tip and feedback conditions in near-field fluorescence imaging. (a) Pulled fiber-optic probe coated with 50-100 nm of aluminum around the sides. By placing the probe (white) near the sample surface (gray), the emerging light (green) interacts with the sample before diffraction effects degrade the resolution, (b) 15 um x 15 urn NSOM fluorescence image of dilC18 dispersed in a lipid monolayer taken with an uncoated NSOM probe. The light escaping from the sides of the taper forms diffraction patterns, which are mapped in the fluorescence image, (c) A 10 urn x 10 um fluorescence image of a similar sample imaged using an aluminum-coated probe. High-resolution features are now visible, the size of which reflects the dimensions of the particular aperture used, (d) Same sample area and conditions as (c) with larger tip-oscillation amplitude, illustrating degradation in resolution and sensitivity. (Adapted with permission from Ref. 29.)
challenges were daunting, and progress awaited developments in lasers and piezoelectronics before becoming feasible today. Experimentally, the first demonstration of subdiffraction-limit resolution was accomplished in the microwave region of the spectrum, in which the long wavelength of the radiation (centimeters) relaxed the constraints on aperture formation and aperturesample distance regulation (20). These successful results sparked interest in pushing the technique into the visible part of the spectrum. The much shorter wavelength of the light, however, seriously complicated the technical aspects of implementation and another decade would pass before subdiffraction resolution was achieved in Dieter Pohl's laboratory at IBM Zurich (21) Pohl and another group working inde24 A
pendently at Cornell University first broke the diffraction limit at visible wavelengths in the early 1980s (22). These impressive achievements illustrated the feasibility of NSOM and marked the beginning of subdiffraction limit optical imaging. Although still not routine, NSOM has been used successfully to gain new insights into the properties of samples at the nanometric level. Not surprisingly, as Synge originally pointed out, the key issues involved in implementing the technique revolve around aperture formation and positioning (discussed next) Since the initial reports, the steady development of NSOM has been punctuated by notable technical advancements that have led to dramatic improvements in performance. Perhaps the greatest ad-
Analytical Chemistry News & Features, January 1, 1999
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