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Making the Point

Thu, 07/26/2007 - 11:48am
The latest CLSMs pack surprises—stunning resolution, 3-D imaging, micro machines, and new market opportunities.

One constant in the development of microscopy is change, but sometimes technology is not the only evolution. In the case of a new confocal point-scanning microscope from Olympus Industrial America, Orangeburg, N.Y., for instance, the product could open the door to new laboratory environments.

Confocal imaging is benefiting from a renaissance in photonics. Major advances—efficient mirror units, thin film dielectrics, fiber optics, new noise reduction methods, microelectromechanical machines (MEMS), and stable laser light sources—have all contributed make confocal laser scanning microscopy (CLSM) in particular, a growing segment in laser-assisted imaging.

Olympus drew from experience in surface morphology studies in the semiconductor industry when designing a new microscope that uses low-wavelength laser scanning technology. The LEXT OLS 3100 series, a reflection-type CLSM, boasts one of the highest sub-1-mm resolutions on the market and is specifically directed at materials surface modeling by making use of point scanning to deliver high-resolution 3-D images in real time. This plus the flexibility of multiple imaging modes is intended to give Olympus’ confocal tool applications in a number of R&D industries.

Point scanning has long been a useful method for looking at fluorescence emission intensity changes in dynamic environments. These specimens usually involve living cells or other biological applications. As demand for high resolution and video imaging grows, more researchers in the semiconductor and materials industries also want to examine surface dynamics in non-biological settings.

The OLS 3100 is the second iteration of the LEXT series and incorporates several improvements. According to David Rideout, product manager of Olympus’ MicroImaging Division, the company saw a gap between scanning electron microscopy and high-end optical microscopy and decided to fill it with a new microscope.
Click to enlarge.
Click to enlarge.
Click to enlarge.
Olympus provides a number of software tools to manipulate 3-D images produced by the LEXT microscope. In addition to rotating and scaling, images such as these observations of the reverse side of a silicon wafer can be tailored to focus on surface characteristics or textures. Image: Olympus

Resolution is paramount
Why confocal? White light microscopy supplies real-time imaging, but the resolution is limited. Scanning electron microscopy can study nanoscale specimens, but at added expense, time, and the potential headache of preparing samples through vapor deposition of metals or vacuum pumpdown. Confocal microscopy, on the other hand, takes advantage of interference optics and photomultiplication to maximize the most desirable light information, and in turn yields spectra for a CCD to read.

Getting that information requires several sensitive components that must all work in perfect unison. This, plus traditional confocal design roadblocks, can lead to trade-offs for designers. When confocal is used with fluorescence, for example, the photonic limitations of acquiring fluorescence data at high magnification makes good signal-to-noise ratios difficult to acquire. Additional problems include galvanometers subject to vibrations and noise from accompanying electronics.

In addition, shorter wavelengths require tremendously sensitive optics, and assembling these such that they work together seamlessly (and on a repeatable basis) is a tall order for developers of confocal instruments.

Olympus calls the LEXT a “violet opt” system. Its focused light is near the ultraviolet (UV) range of the spectrum. Few applications in microscopy use wavelength lower than the 408 nm laser used for the OLS 3100.

“The lower the wavelength, the better the resolution,” says Rideout. “This is just about the lowest wavelength you can use before going into the UV range. Once you go into the UV, the optics get more complicated and expensive. The key to using the laser is the intensity and directivity of the light.”Olympus worked to minimize the aberrations associated with using this wavelength, resulting in a planar resolution of 0.12 µm and spatial patterns of 0.01 µm in height.

The confocal setup consists of a circular confocal pinhole, a laser light source, LED illumination, a photomultiplier, a combination MEMS/galvanometer scan stage, and apochromatic objectives designed specifically for use with a near-UV system.

“Objective lenses are tailored to magnification with as little optical aberration as possible. The lenses used on the LEXT have the highest level of aberration correction,” says Rideout.

Packaging useful 3-D
Analysis of wafer surface characteristics demands accomplished dimensioning capabilities. System setup and support software must measure up to the task of presenting useful imaging. Step height, line width, and point distance are all measurable, and final images can be rotated and displayed in several ways. Surface only, texture, wired frame, perspective, real color, and other variations can be rendered. Other features include real-time distance measurement, particle analysis, and simultaneous laser scanning and optical specimen observation.

The LEXT can be used in a number of different modes:

1) Brightfield observation—useful for detecting a flaw on a color filter or to locate corrosion on metal;

2) Differential interference contrast (DIC)—interference contrast observation helps identify flaws down to a few nanometers in height;

3) Laser confocal—high planar resolution is possible with sensitive optics and short wavelength laser light;

4) Laser confocal DIC—the combination of sharp contrast and high resolution yields wide application possibilities.

Two-dimensional measurement is best in the range of 1.5 mm to 1 µm widths. Both line-width and geometric measurements are possible. For 3-D study with LEXT, measurement of bumps in the range of 1 mm to 0.5 µm is ideal. Volume, capacity, surface area, and other characteristics can all be measured in 3-D, as well as thin transparent film thickness. In addition, analysis of line and measurement of mean roughness depth is possible to 0.1 µm. This function is assisted by a noise filter developed by Olympus.

Reflectance can cause problems for conventional laser microscopes, particularly in specimens with widely variant surface characteristics, such as those with multi-structure patterns and holes. LEXT has a five-step sensitivity switching function that allows an image to be captured by switching from one sensitivity level to another. The feature gives researchers a better chance to gather information about height and luminance on structures with highly variant reflectance.

Click to enlarge.
Click to enlarge.
A brightfield observation of a stud bump in real color, compared with the surface rendering. Image: Olympus

Confocal features evolve
Designed specifically for LEXT, the CLSM’s scanning stage uses MEMS technology designed by Olympus. Because it rasterizes the excitation scans and collects photon information, the scanning stage needs both linear and progressive performance at a high speed to keep up with the CLSM’s ability to capture many frames per second and to reduce the amount of dead time in the scan. Like other microscopes capable of true video-rate imaging, the LEXT uses a high-speed resonant galvanometer to both scan and descan focused light. However, designers eliminated the second mirror in favor of a MEMS device, an increasingly popular choice for 3-D microscopes.

“The MEMS device we use is in the x direction, and is quieter and faster, improving speed and reducing vibrational issues,” says Rideout.

The nosepiece of the LEXT features motorized switching, which helps efficiency. A built-in automatic light intensity adjusting function keeps brightness levels unchanged after magnification switching. For the OLS 3100, the speed of the automatic focus has been increased and a one-push gain was incorporated to help speed up 3-D image capture.

Putting LEXT to work
Olympus demonstrated the system’s flexibility by testing several specimens that typically pose challenges for study. For example, the LEXT can be used for observing complex structures resulting from the formation of polyamide resins. An injection-molded polyamide can be modeled in 3-D, clearly showing details of spiral-shaped spherocrystals, a few microns across. Other injection-molded materials, such as foam, can be sectioned and studied for features like air bubble distribution and volume.

According to Rideout, the LEXT is ideal for the precise measurement of micro-fabrication surfaces. In addition to polymers, surface properties can be characterized on wafers and metallic surfaces, and nanoscale structures in silicon wafers can be analyzed. Adhesives, too, can be quality-controlled with CLSM. Confocal methods also avoid the vapor deposition necessary for using a scanning electron microscope to observe reaction-ion etching, and it has the added advantage of viewing the object in its natural state.

“The best applications for the LEXT are materials research and semiconductor,” says Rideout. “We have had a great deal of success in both of these markets. The tool can act as a bridge tool between white light microscopy and SEMs. The LEXT is particularly good for failure analysis in the semiconductor and wafer market.”

Until electron microscopes become as portable, inexpensive, and easy-to-use as an optics-based microscope—confocal or otherwise—the market for a device capable of real-time 3-D imaging at resolutions below 1 µm will continue to strengthen. And with advanced laser-based optics continuing to change the microscopy playing field, there’s no reason to believe yet another new species of CLSM won’t appear on the horizon.

“LEXT really has allowed us to play in the higher end of existing markets and keep up with market changes. For instance, many semiconductor applications are now moving outside the scope of standard white-light microscopy. The LEXT’s resolution levels allow us to continue to offer solutions to this market,” says Rideout.

—Paul Livingstone
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