Developers are pushing the limits of existing imaging technologies to create systems with three times the measurement speed, resolution and consistency.

Imaging used to be simply defined as the representation of an object's external form. That definition no longer holds true as researchers now look for more than just an image. They look for more information within an image, such as fluorescent tags, mechanobiological parameters, internal structures, fabrication while imaging and the characterization of materials as yet undefined. The images are increasingly becoming extremely small, most of them now being measured in nanometers. New imaging systems also have to be easier to use than even their less functional predecessors, able to process samples faster and able to process each sample less expensively.

Obviously, there are numerous types of imaging systems available for researchers, including those for chemical, optical, thermal, medical and molecular imaging and more. In this month's cover story, seven new imaging technologies and solutions for research and processing applications are explored.

Simplified fluorescence

The first imaging technique we'll examine is the FLoid Cell Imaging Station from Life Technologies in Carlsbad, Calif. Traditional fluorescent microscopes have historically been cumbersome to use. The set-up time alone for some devices often takes more than 30 min and the need for experienced technicians to run them can create scheduling constraints within the lab. Instead of conveniently stationing them on benchtops, each instrument must be placed within a darkroom, which takes up space that could be used for other research purposes.

The FLoid Cell Imaging station overcomes these limitations, as it was designed to eliminate the complexity of fluorescent microscopes. With this system, high quality images can be captured in less than five minutes and printed directly from the platform's printer or downloaded and emailed to serve as supporting documentation for the user's research. At a third the cost of traditional fluorescent microscopes, the FLoid Cell Imaging station is a fully integrated benchtop instrument with an intuitive user interface that enables quick detection and verification of fluorescently labeled cells with just a few clicks of a mouse.

The station includes protocols and Molecular Probes reagent selection guides to assist users in preparing their samples prior to imaging—steps that users would normally have to research through other sources. Depending upon the specific cellular structure or biological activity users are interested in imaging, the pre-loaded reference tool provides the precise protocol to follow and reagents to use for optimal performance on the instrument.

Customized inverted workhorses

The IX3 inverted research microscopes from Olympus, Center Valley, Pa., offer a line of three imaging instruments researchers can customize to meet their current research demands, while providing the flexibility to grow as their needs change. The top-of-the-line IX83 is a fully automated research microscope, while the IX73 is a modular system that can be configured in manual, semi-motorized or motorized versions to benefit a wide range of applications. The IX53 system microscope offers quick and efficient routine examinations of multiple tissue samples. The stands for these microscopes provide a choice of manual control, encoded manual, semi-motorized operation or fully automated high-speed motorization. A low-profile ultrasonic motorized stage provides quiet, smooth, precise movements. The IX83 and IX73 have a multi-story optical deck area that can be configured with one or two decks for the insertion of various optical components. These decks modularize the microscopes, allowing the rapid plug-and-play insertion of magnification changers, filter turrets and a right side port, among other options. Changing equipment and options is as easy as sliding out one deck insert and sliding in another. Both decks open out like cabinet drawers for easy access. The open architecture of the deck area allows researchers to insert optical components, while the large available space is ideal for user-designed modules.

A real-time controller in the IX3 delivers precision in high-speed imaging, with microsecond timing accuracy for high-speed devices such as light sources, triggered cameras and other TTL-controlled devices. A new Fly-eye fluorescence illuminator provides even, bright illumination across the specimen, allowing a much wider field-of-view and filling the imaging chips of even large-format cameras. Additionally, the IX83 is available with a third-generation zero drift focus compensation (ZDC) technology that allows accurate, consistent focusing during long-term time-lapse experiments. ZDC can even operate without a computer in standalone mode.

Nanoscale characterization

The PeakForce QNM imaging mode provides new functionality for the Bio-AFM (atomic force microscope) from Bruker, Santa Barbara, Calif. This includes expanded support for a lower modulus range covering live cells and tissues, making it the best mechanical property measurement tool for soft biological samples. The system's new PeakForce Capture function gives researchers full access to the PeakForce QNM data to explore alternate analysis models and validate their results against other techniques. New analysis capabilities in the Quantitative Force-Volume Mapping mode enable direct comparisons with this widely used legacy technique. Together, these features provide the fastest, highest resolution and most quantitative characterization of nanomechanical properties over a wide range of biological samples.

QNM maps and distinguishes between nanomechanical properties, including modulus and adhesion, while simultaneously imaging sample topography at high resolution. The forces applied to the sample are precisely controlled and a variety of probes can be used. This allows indentations to be limited to several nanometers in most cases, which both maintains resolution and prevents sample damage.

Nanofab and imaging

Carl Zeiss Microscopy, Jena, Germany, recently introduced the ORION NanoFab as the first multi-ion beam tool based on gas field ion source technology. As a major enhancement to its existing helium ion microscope, the NanoFab utilizes neon ions. The system is therefore capable of providing a complete sub-10-nm nanofabrication and sub-nm imaging solution for various research labs. An optional gallium focused ion beam (FIB) column can also be integrated.

The neon ion beam offers precise machining and nanofabrication capabilities due to higher sputter yields in ion beam milling and faster resist exposure in ion beam lithography. The helium ion beam allows sub-10-nm nanofabrication as well as high resolution imaging capabilities in the same instrument. The optional FIB column also allows massive material removal.

Eliminating error

Researchers at the National Institute of Standards and Technology (NIST), Gaithersburg, Md., have refined a method for measuring nanometer-sized objects that may help computer manufacturers more effectively size-up the myriad tiny switches packed onto chips’ surfaces. The method makes use of multiple measuring instruments and statistical techniques. These techniques minimize the uncertainty in measuring features smaller than a few nanometers, which optical microscopes are unable to distinguish.

In the past, metrologists have used indirect methods for these measurements, such as scatterometry—deducing shapes from sampling the patterns that light creates as it scatters off the features' edges. When scatterometry is insufficient, AFM is used, but while AFM gives distinct measurements of the height and width of nanoscale objects that troubles scatterometry, it's expensive and slow. Scatterometry can tell you the width of an object is 40 nm, but the range of error could be +/-3 nm, a relatively large variance. Also, the total uncertainty increases when measurement techniques are combined, making the object's image even hazier.

The solution to this issue was a combination of scanning techniques and statistical analyses. The NIST researchers first created a library of simulated data based on typical semiconductor chip feature dimensions to which they can compare actual measurements with AFM, scatterometry and other means. A complex statistical analysis of library values is then compared to actual measurements to extract valid measurement values. The elegant technique used by NIST statistician Nien Fan Zhang was the use of Bayesian analysis, which incorporates a few key additional measured values from other tools into the library model. The result is a reduction in the uncertainty of measurements by more than a factor of three. This approach is expected to be essential when measuring complex three-dimensional transistors that are 16 nm in size or smaller in the near future.

Fast production imaging

FEI, Hillsboro, Ore., recently introduced its new Helios NanoLab DualBeam systems for engineers that need to make vital process improvement decisions. The 450HP and 1200HP DualBeam systems include new capabilities that meet the critical requirements for semiconductor process development at the 28-nm device geometry node and smaller.

“TEM (transmission electron microscopy) samples must be ultra-thin, of the highest quality and generated in a routine and consistent manner across a range of tools,” says Rudy Kellner, the VP/GM of the Electronics Business Unit of FEI. “Typically, as samples get thinner, the process difficulty becomes the time to get results, operator skill level and subtle differences among equipment, which these systems overcome. Ultimately, the system's ability to yield more good samples at double the throughput allows for potentially significant reductions for both the time-to-answer and the cost-per-answer.”

The 450HP and 1200HP systems can prepare 15-nm-thick samples with less than a 2-nm damage layer in 90 min, two times faster than competitive alternatives. The systems' iFast automation software maximizes ease-of-use while ensuring consistency among multiple operators and systems. QuickFlip grid holders for the samples facilitate inverted sample preparation to improve sample quality while maintaining high throughput. Finally, Cell Navigation software allows automated navigation within non-unique memory arrays that can locate a single designated bit cell in a 50-nm lateral field.

Benchtop and MRI

We'd be remiss to leave out two other technologies that have made a significant impact on recent imaging procedures—benchtop SEM (scanning electron microscopy) and MRI (magnetic resonance imaging). A number of benchtop SEMs have been introduced over the past several years from varying manufacturers. The NeoScope, for example, was initially introduced by JEOL, Peabody, Mass., in 2008. Recently, the company introduced upgrades with higher magnifications, multi-touch screen controls and a sleek new design. As simple to use as a digital camera, the new NeoScope is a high-resolution SEM that produces images with a large depth of field at magnifications ranging from 10 to 60,000x. It features both high and low vacuum operation, three selectable accelerating voltages and secondary electron and backscattered electron imaging. The instrument accommodates samples up to 70 mm dia and 50 mm thickness. Both conductive and non-conductive samples can be examined and an optional EDS (energy dispersive spectroscopy) feature is available for elemental analyses.

The NeoScope's touchscreen interface has the familiar look and feel of today's smartphones and tablet computers. Automatic functions as well as pre-stored recipe files make it easy to use for a multitude of sample types. 

MRI's have become a mainstay of the medical community in their ability to image biological tissues quickly and reliably. Agilent Technologies (who in 2010 purchased Varian and its MRI product lines, which began in 1984) recently launched its nScope eMRI preclinical imaging system. The eMRI system features an improved interface for ease-of-use. 

Optimized packages uniquely align with specific user requirements, and a redesigned workflow ensures simple and ergonomic operation. This is particularly useful for new users who wish to generate biomedical results without being bogged down in technical details.