Scanning optical microscopes close in on submicron scale

transcript

Lightwave microscopy and scanning technology

add a new dimension to semiconductor metrology

Scanning Optical Microscopes Close in on

Submicron Scale by G.S. Kino and T.R. Corle

S e v e r a l new microscope families with powerful resolv- ing capabilities are now here to aid the re- searcher in better visualizing submicron structures. Among these is the confocal scanning optical microscope (CSOM). Recent improvements in this type, first invented more than 20 years ago, presently equip it for demanding applications in imaging and measurements of semiconductor and biological materials.

Compared to a standard microscope, a CSOM offers the distinct advantage of eliminating defocused images rather than creating a blur of that image. In short, the CSOM’s image intensity drops as the image is defocused, whereas with a standard microscope the intensity does not change. This property allows the independent imaging of structures with height differences com- parable to a wavelength, thus permitting

profiles, three-dimensional reconstructions (Fig. l), and quantitative measurements of height. In addition, CSOM images tend to have sharper edges and more contrast than images obtained with a standard microscope.

The CSOM’s defocusing characteristic permits the optical cross-sectioning of transparent specimens, which is particularly useful when examining biological materials. The features of one layer of such a material can be observed without glare from layers in front or behind it, or by reflections from the glass slide on which the sample is mounted.

The principles of the CSOM are best understood in the context of conventional optical microscopy. A standard optical microscope consists of an objective lens and an eye-piece (Fig. 2). Generally, the entire sample is uniformly illuminated, a technique known as bright-field imaging. The objective lens forms a real inverted image of the object in aposition suitable for viewing with the eyepiece. The eyepiece functions as a simple magnifier, producing a virtual image of the object at a comfortable viewing distance for a “standard observer.” The total magnification is the product of the objective

28 CIRCUITS A N D DEVICES

and eyepiece magnifications. which allows high v n l u e ~ - l 0 0 0 to 2000x-to be achieved in ;I small space.

Unlike a conventional optical microscope. the CSOM illuminates and images the sample one point at a time through a pinhole (Fig. 3a). Light passes through the pinhole to a microscope objective, which focuses it to a diffraction-limited spot on the object. Light reflected from the object passes back through the objective and through another pinhole. If the object is moved out of focus. the reflected light reaching the pinhole is defocused and most ofthe light does not pass through it (Fig. 3b). Consequently. ifadetec- tor is placed behind the pinhole. the amplitude of the signal received drops off rapidly with the defocus distance. and the image disappears. The microscope is "cow focal" because the ob,jective lenj is used tu ice. both to illuminate and image the sample. Since only one point is illuminated at a time, the sample or beam must be raster scanncd to build up the image pixel by pixel. as in an ordinary television receiver. Thus the CSOM is ;I scanning optical microscope.

The three basic requirements for confocal scanning microscope$ are point illumination. point detection. and confocal imaging. These requirement5 lead directly to the need for image scanning, which is generally ap- proached in one of three ways. First, the sample or the optics can be mechanically scanned. The resulting raster image can then stored in acomputer anddisplayed as avideo image. This entire process typically takes a few seconds for each frame. The second, and somewhat faster, approach is to scan the beam using a galvanometer mirror or Bragg cells. The third approach is to use a Nipkow disk. which can produce real-time images at hundreds of frames per second.

History The idea for confocal scanning optical microscopes was examined by Young and Roberts in 1951 [ I ] and by Minsky in 1957 [ 2 ] . Minsky accurately described the salient feature5 of this type of imaging system in his patent, but lack of an adequate light source prevented full development.

The h e r solved many of the problems

encountered by early researchers. In 1969, Davidovits and Egger 131 were the first to develop a working laser-based CSOM. Much ofthe recent work has been stimulated by the development of the scanning acoustic microscope (SAM) by Quate and Lemons [ 3 ] because the SAM exhibited better transverse definition and far better range resolution than a standard optical microscope at the same wavelength. Kompfner. who was involved with the early acoustic work, suggested that this imaging technique might be excellent for observing internal features of translucent matcrials. such as body tissue. He started a group with Wilson and Shep- pard at Oxford to try out similar scanning microscope principles with optics.

At about the time that Davidovits and Egger were building their microscope, Petran and Hadravsky in Czechoslovakia invented a type of CSOM that operated in real time and produced an image that could be directly observed with the naked eye [6]. Their idea was to use a Nipkow disk to fulfill the requirements of point illumination and point detection. Petran and Hadravsky called their invention the tandem scanning reflected light microscope (TSRLM).

The Nipkow disk, invented in Germany in 1884, was used in the first form of mechanically scanned television and was easily adapted to provide the raster scan in a CSOM. It consists of an opaque disk into which many thousands of pinholes have been drilled or etched in a spiral pattern. Each illuminated pinhole on the disk is imaged by the objective to a diffraction-limited spot on the sample. The light reflected from the sample can be seen in the eyepiece after i t has passed back through the Nipkow disk. Several thousand points are simultaneously

illuminated on the disk achieving, in effect. several thousand confocal microscopes all running in parallel. Spinning the disk fills in the spaces between the holes and creates a real-time confocal image.

Since only about one percent of the area of the disk is transparent in most cases. a moderately intense light source is required. Initially, heroic efforts were required to obtain adequate illumination. In Petran and Hadravsky's first experiment, the microscope was carried to the top of a high moun- tain and "while one reflected the sun's rays into the microscope, the other had a few seconds glimpse which proved that it would work." [6] A major problem was to eliminate the reflected light from the top surface of the disk, which Petran and his colleagues ac- complished by passing the light through one set of pinholes in the disk and using the objective to produce diffraction-limited spots on the sample. The reflected light returned to the eyepiece through 21 series of mirrors and a conjugate set of holes on the opposite side of the disk.

One of the authors of this article (G.S. Kino) first saw an operating Petran microscope in Alan Boyde's laboratory at Univer- sity College in London. Its advantages over a mechanically-scanned CSOM became im- mediately apparent. As the focus knob was turned, different features came into view in real time. But its disadvantages a poor light budget, considerable difficulty in alignment, and mechanical complexity have kept all but a very few examples of this microscope from being constructed in the twenty years since its invention. In what may signal a basic change, however, a commercial ver- sion has been developed recently by Tracor Northern of Middleton, Wisconsin.

S t imula ted by Pe t ran ' s work, we developed a simplified form of the tandem scanning microscope at Stanford. We call it the Real-Time Scanning-Optical Micro- scope (RSOM) (Fig. 4). This microscope uses the same set of pinholes for illumination and imaging, which reduces the number of optical components and dramatically simplifiesalignment 171. The disk is made of low- reflectivity black chrome to reduce the light reflected from the top of the disk. The

CIRCUITS A N D DEVICES

illumination

eyepiece

numerical aperture (N.A.) =sin eo

2. A cmventional optic.ul mic,rost,ope wntains an ohjecti1.e lens and an eyepiece. Typic,ully. tht' ohjec,t to he iieM,ed is ei,enly illuminated. The resolution of the system is determined by the numerical aperture ( N . A . ) , i.e. the sine of the half- angle suhtended by the lens

illumination

out of illumination

focus I -:-

object scanned

eyepiece

photodetector

3. Unlike a coni.entiona1 micro.sc.ope. the c.onfac,al sc.unning optical mic,ru.sc.ope (CSOMI illuminates and imuges the sample one point atu rime through a pinhole ( a ) . l f the object is moi,ed out offocus. the reflected light reachin!: the pinhole i s defoc~used and most of the liSht does not pass thr-ough it (h ) . Conseyuenrly. if a detector- is plawd hehind the pinhole. the amplitude of the signal rec.eii,ed drops off rapidly w'ith the defoc,its distance. and the image disappears.

MARCH 1990

- _ _ _

disk is tilted so if any light is reflected, most of it strikes a stop in the microscope. In addition, the illumination is polarized, and an analyzer is placed in front of the eyepiece to eliminate any remaining light reflected from the disk. A quarter-wave plate placed near the objective rotates the polarization of the image so that it will be passed by the analyzer.

Comparing Microscopes The CSOM is but one of many types of scanning microscope. The best known of these is the scanning electron microscope (SEM). Other variations (Fig. 5 ) include the scanning acoustic microscope (SAM), scanning tunneling microscope (STM), scanning force microscope, and near-field scanning optical microscope (NSOM).

The scanning acoustic microscope (SAM) uses afocusedacoustic beam emitted from an acousto-electric transducer to scan an object [8]. The object is usually immersed in water, because water is an excellent propagating medium for acoustic waves. The waves reflected from the object to the transducer yield an electrical output. As in the scanning optical microscope, the transducer is scanned over the surface of the object and a raster display of the output is shown on a video screen. The definition of the SAM is on the order of the acoustic wavelength in both the transverse and range directions. Thus, for acoustic waves of frequency 3 GHz (h 0.5 p m ) propagating in water, the definition is on the order of .4 pm. The major advantage of the acoustic microscope is that it can be used for examining the properties and interiors of optically opaque materials. A particularly important applica- tion is the imaging of the internal structures of honeycombed and composite materials using low-frequency acoustic waves on the order of 3 MHz.

The scanning electron microscope (SEM) uses a focused electron beam to scan an object in a vacuum [9]. The image is obtained by collecting secondary or back- scattered electrons from the source on which the focused beam impinges. Since the number of electrons collected depends on the nature of the material and its geometry, ex-

cellent images of geometrical features are obtained. The SEM has better definition than a CSOM in the low submicron range because the diffraction wavelength of an electron beam is so small. For 2 keV electrons, typical wavelengths are on the order of 0.04 nm, compared with 500 nm for optics. But in many applications the resolution of an SEM is worse than the theoretical limits because of electron scattering as the beam penetrates the sample. Typical definitions are on the order of 100 8, for 2 keV electrons and 2.5 8, for 10 keV electrons. The major disad- vantage of the SEM compared with an optical microscope is that the sample must be in a vacuum. This complicates the measure- ment and makes the SEM unsuitable for use with live materials. Additional problems are the possibility of damage to biological and semiconductor samples by the high-energy electrons, and the need to coat surfaces with a metal so they do not become negatively charged. In recent years, some of these dif- ficulties have been obviated by working with beam acceleration potentials below I kV. With such low-voltage beams, the secondary emission coefficient of insulators is greater than unity, so charging does not take place.

One big difference between the SEM and CSOM is that, because of the SEM's small aperture, its depth of focus is relatively long, approximately lpm. Images remain in focus over a relatively large range and have a three-dimensional appearance. On the other hand, not much depth information is provided. To secure depth information the sample must be cross sectioned, which destroys it for further use.

The resolutions of the scanning-optical, scanning-acoustic, and scanning electron microscopes are determined by diffraction, and hence by the wavelength of the waves employed. Another class of microscopes, the near-field microscopes, have definitions which are based on the way that quasi-static fields fall off from a pinhole or probe. The most familiar of these is the scanning-tunneling microscope (STM). These microscopes are not suitable for measuring the internal properties of materials because the definition decreases so rapidly away from

the probe. They are, however, ideal for measuring surface properties and profiles by arranging for the probe to follow the surface.

The STM works on the principle that when a tungsten point is placed within a few angstroms of the object being examined, and a potential is placed between the tungsten point and the object, a tunneling current will pass from the point to the object [IO]. The tunneling current is exponentially dependent on the spacing between the point and the object. Generally, an image is obtained by scanning the point over the surface of the object and measuring the spacing required to maintain a constant current. The definition i s typically on the order of one or two angstroms because of the small spacing between the point and the object and the highly nonlinear dependence of the tunneling current on this spacing. The STM i s used for measurements at the atomic level, producing superb images of atomic structure at the surface of an object.

Scanning force microscopes are deriva- tives of the STM that measure the force between a point of tungsten (or other material) and the object [I 11. Many force microscopes have been developed. One type is based on using a magnetic wire sharpened to a fine point and measuring the force between it and a magnetic material, such as a magnetic disk in a disk memory. When the point probe is scanned over the disk, this technique provides superb profiles of the domain fields with definitions in the range of a few hundred angstroms.

The near-field scanning optical microscopes (NSOMs) either use the light collected through a pinhole when a sample is illuminated by an external source, or vice versa [ 121. In this case, the amount of light collected falls off rapidly with the spacing of the pinhole from the object. Because the pinhole is so small-less than 100 nm in diameter-the definition of this microscope depends on the pinhole size rather than the optical wavelength. It is typically in the range of a few tens of nanometers to a hundred nanometers.

Scanning optical and conventional optical microscopes, which are based on diffraction optics, have different properties from

eyepiece

4. By usinl: a spinning Nipkow disk topro\,ide both pinholes arid scanning. the real-time scannincq opt ical microscope (RSOMJ ,functions l ike thousands of conventional CSOMs operating in purullel.

CSOM 1 SEM I

18, 1008, 1Pn l00pm 10-

g lOmm1 .- I

Approximate Transverse Resolution

CSOM SAM SEM STM NSOM

5. The resolutions of d(f?r-erit types of .sc~urinin,q mii~r.osc.opes direc~tr0n.s.

in both the tiuiisi'erse undrange

CIRCUITS AND DEVICES

0 0.2 0.4 0.6 0.8 1.0 1 2 1.4

POSITION (urn)

standard optical

microscope,

confocal scanning

microscope

6 . The edge response of an RSOM is sharper thuii that of a c.oniwitionul microscupe. These C F perinientul plots of intensity for euch type oj microscope. MYth euch haling a 0.9 N . A . lens und utili5n~q light ut h = 550 inn, shon. the 50% intensity point ,for the stundurd micro.wope cor- responds to the 25% intensity pointfor the RSOM.

c - .. - -

"" Experiment Theory

uniform P(r) weighted P(r )

_ _ _ -

- ( 1 . S ~ m

7. Depth response (fa CSOM using an Olympus 8O.vIO.90 N . A . ohjec.ti1.e ut h = 633 nm provides the entirely typiml full hidth ut a halfmu.uimum (d/z/) cf0.5 m.

near-field microscopes. Their definition depends on the wavelength, and the beam may penetrate into a transparent material to examine its interior. Optical systems also have the great advantage of parallel pro- cessing, which makes it possible to form an image in real time. However, it is difficult to obtain better definition by scaling the wavelength even to the near UV, let alone to the soft X-ray range, because of the lack of suitable light sources and lens materials. For these and other reasons, scaling to shorter wavelengths might not always be the best way to obtain high definition despite the better theoretical diffraction limits of a shorter-wavelength beam. It is therefore vital to take advantage of any optical techni-

MARCH 1990

que which can help to improve the definition of a microscope in the standard optical range.

Imaging Properties The improved definition of the CSOM compared to a standard optical microscope can be understood by analyzing the point spread functions of both instruments. In any lens system be it electron, acoustic, or optical diffraction effects will tend to spread the image of a point object. The amplitude of the resulting image vanes with radius. This variation is known as the point spread function (PSF), and is simply the response of the imaging system to a d function. For an ideal lens the PSF is approximated by the equa- tion.

The optical systems of the CSOM and the conventional optical microscope are virtual- ly identical. Their main differences lie in the methods of illumination and detection. The standard microscope uniformly illuminates the entire sample with an incoherent light source, and the image is detected by a large- area detector such as the human eye or a TV camera. A point on the sample is imaged as a diffraction-limited spot on the detector by the objective lens. The resulting intensity PSF is equal to the square of the amplitude PSF of the objective.

In a CSOM, only one point at a time on the sample is illuminated through the objective. The amplitude of the field at the sample, rather than being uniform, is given by the PSF of the objective. In turn, the sample is imaged by the objective onto a point detector. Thus, for simple objects such as edges and points, the CSOM's intensity image is the square of a standard microscope's intensity image.

The CSOM's intrinsic resolution is not much better than that of the standard optical microscope, but it produces a visually sharper image (Fig. 6). The RSOM form of the CSOM also requires very little adjustment to achieve its optimum performance, whereas a standard microscope tends to require quite careful adjustment.

The major advantage of CSOMs over standard microscopes is that the image disappears when it is out of focus. This makes

8. Photographs if three images ( f a n integrated c,ircuit taken ut three difleer.entfoc.us levels with an RSOM

it possible to determine quantitatively the heights of surface features.

If a sample is scanned axially through the focal plane, the distance between half-power points of the detector response the range resolution of the microscope is given approximately by the formula 131:

d,= 0.45 h/ ( 1 - cos 8) where sin 8 is the numerical aperture N.A. of the objective lens. A typical CSOM range resolution is dz = 0.5 pm (Fig. 7).

Optical sectioning is carried out by imaging thick structures one layer at a time (Fig. 8). Only the layers of the circuit which are within approximately 0.25 pmof the focus level appear in each of the photographs. The confocal images can be combined in false

color or plotted as a three-dimensional reconstruction to show the entire circuit. The images in Fig. 8 used narrowband illumination of the sample. In the RSOM, however, there is a high-pressure arc lamp producing broadband illumination. Chromatic aberra- tion within the objective lens causes the different colors to focus at different depths. This broadens the depth response, but allows the resolution of height differences by color changes.

The same techniques may be used to obtain good-quality images of biological specimens and to carry out optical cross sectioning of these materials. A series of images of a mouse cerebellum, for instance, can be combined to make an extended-focus image of the same sample. The image could appear to be in focus over a range of almost 20pmbecause each of the individual images that were added together is unblurred. Such an image could not be obtained with a standard microscope.

It is also possible to obtain images more than 100pm beneath the surface of a translucent material such as bone despite the considerable scattering of light. This kind of image, also, cannot be obtained with a standard microscope.

Yet another important biological ap- plication of the CSOM is fluorescence imaging. Here the excellent range resolution of the scanning optical microscope eliminates glare from regions of the material where the beam is unfocused (Fig. 9).

There are now several commercial firms marketing CSOMs for use in the semiconductor industry, for biological applications, and for ophthalmology. In the semiconductor industry, CSOMs are particularly valu- able for their very good range definition and image contrast. The development of real- time confocal optical microscopes enables simple inspection for those familiar with standard micro-scopes, but with the additional advantages of excellent range definition and cross-sectioning ability for accurate metrology. The most important applications so far have been for semiconductors and for biology, but the range of as yet untapped applications for scanning optical microscopy is very large.

9. These matched fluorescence images of white blood cells helped convince biologists of the confocal m i i n m q w ‘ s importance. In the coni‘en- tional mic,roscope image (top), the glare from out-of-focus layers tends to ohscur-e the cellular features. In the f o c d image f tom the cwfocal microscope (bottom), the cross section of the blood cells is clear.

Acknowledgment This work was supported by the National Science Foundation on Contract No. ECS- 88-1 3558, the Semicon-ductor Research Corporation on Contract No. SRC 88-MJ- 120, International Business Machines on Contract No. IBM 645416, and the Office of Naval Research under Contract No. NOOO 1 - 4-84-K-0327.

K i n o Corle

Gordon S. Kino is Professor of Electrical Engineering, Professor by courtesy of Ap- plied Physics, and Associate Dean of Engineering for Facilities and Planning at Stanford University. He has published close

to 400 papers. Professor Kino is a Fellow of the IEEE, the American Physical Society, and the AAS, and is a member of the Nation- al Academy of Engineering.

Timothy R. Corle holds a joint appoint- ment as a research associate at Stanford University and an optical systems engineer at Prometrix Corp. His current interests include optical and other novel imaging techniques for semiconductor metrology.

References 1. J. Z. Young and F. Roberts, Nature 167, 231 (1951). 2. M. Minsky, U . S. Patent 3,013,346, Microscope Apparatus (December 19, 1961). 3. Paul Davidovits and M. David Egger, Nature 233,831 (1969). 4. R. Lemons and C. F. Quate, Proc. IEEE Ultrasonics Symp., IEEE Catalog No. 73CH08078SU, Eds: J. deKlerk & B. R. McAvoy (Institute of Electrical & Ele- ctronics Engineers, Inc., New York, (1973). 5. T. Wilson and C. J. R. Sheppard, Scanning Optical Microscopy (Academic Press, 1984). 6. M. Petran, M. Hadravsky, and A. Boyde, “The Tandem Scanning Reflected Light Microscope,” Scanning 7,97-I08 (1985). 7. G. Q. Xiao, T. R. Corle, and G. S. Kino, “Real-Time Confocal Scanning Optical Microscope,” Appl. Phys. Lett. 53 (8), 716- 7 18 (1988). 8. C. F. Quate, “Acoustic Microscopy,” Physics Today 38 (8), 34 (August 1985). 9. M. T. Postek et al., Scanning Electron Microscopy: A Student’s Handbook (Ladd Research Industries, 1980). 10. P. K. Hansma and J. Tersoff, J. Appl. i’hys. 61 (2), RlLR23 (15 January 1987). 11. P. K. Hansma et al., Science 242, 209- 216 (October 1988). 12. A. Lewis, “The Physics of the Optical Near Field,” Physics Today (to be published). 13. T. R. Corle, C-H. Chou, and G. S. Kino, “Depth Response of Confocal Optical Microscopes,” Opt. Lett. 1 I , 770-772 (1986). CD

CIRCUITS AND DEVICES

Scanning optical microscopes close in on submicron scale

Documents