Matthew McCluskey founded Klar Scientific after he designed a novel confocal microscope that has an auto-focus feature. Photo: McCluskey/Klar

Seeing is believing, right? But what if, as a scientist, you couldn’t actually see what you needed to? What if you couldn’t see the miniscule defect on the surface of a material, or get close enough to a living nerve synapse to understand how neurons communicate?

Despite today’s $6 billion microscope market, that is still a reality for some researchers. A reality that leaves them with only one option—design their own microscope.

Focus is important
Four years removed from a frustrating “out of focus” problem with his confocal microscope, Washington State University (WSU) physicist Matthew McCluskey finds himself in the unexpected position of founder and chief technology officer of his own startup company, Klar Scientific.

Klar Scientific specializes in the development of optical instruments for materials characterization—some of which arise from McCluskey’s improvisation while working on semiconductor characterization in his lab at WSU.

In 2012, he was studying miniscule defects on the surface of a sample using a confocal microscope, but it repeatedly lost focus as McCluskey scanned the sample.

“With a confocal microscope, if I want to examine a sample that has a rough or irregular surface, the laser spot used to investigate the surface constantly goes in and out of focus,” McCluskey told Laboratory Equipment. “If you happen to be in focus on one spot of the sample, but then translate the sample over, you go out of focus pretty quickly. This causes experimental artifacts.”

To rectify this, McCluskey built a microscope that used an off-the-shelf digital CCD camera as the detector, rather than a photomultiplier tube and scanning hardware that are traditionally used in confocal microscopes. This allowed two improvements: 1) without the photomultiplier tube, the instrument was portable and inexpensive; 2) the auto-focusing feature could now be extended over a wide sample area.

“By maintaining focus as we scan over the surface of our material, we aim to capture detailed images of surface defects with a vertical precision of 10 nanometers,” McCluskey said.

The physics professor knew right away he had designed something novel. WSU filed a provisional patent in 2012 and a non-provisional patent in 2013, which is still pending. In the meantime, McCluskey got to work. He applied for and received funding from the WSU Commercialization Gap Fund to develop several working models of his new microscope, and he teamed up with fellow physicist Rick Lytel—a startup specialist in Silicon Valley. Together, they formed Klar Scientific, LLC in February 2016.

The duo participated in WSU’s 8-week Innovation Corps program, which places researchers face-to-face with industry experts and potential customers to get feedback on their product.

“When I started, I thought we could make the Swiss Army knife of microscopes—it would do everything,” McCluskey said. “Now, I’ve completely changed my mind. Instead, I think it’s much better to make a specific microscope that doesn’t do everything, but is inexpensive, portable, simple and does a job well—not every job. That came directly from customer feedback during the program.”

The portability of McCluskey’s microscope really struck a chord with the I-Corps participants—much more so than he anticipated. In addition to costing upwards of $400,000, a conventional confocal microscope takes up a lot of bench space, and is immovable for the most part. While McCluskey’s microscope may not have as fast data acquisition, it’s inexpensive price point and portability offer researchers previously unheard of options.

Klar Scientific was recently awarded an SBIR (Small Business Innovation Research) phase 1 grant from the National Science Foundation to incorporate this newfound information into an improved version of the microscope. Part of that also includes teaming with Slade Jokela, a former Ph.D. student at WSU, to integrate the microscope’s topographic mapping capability with a diverse set of spectroscopic tools that normally require additional and expensive instrumentation.

McCluskey said he envisions applications in materials characterization, life sciences and even agriculture. And, of course, the microscope’s compact size and low cost make it perfect for academic research—at all levels.

“Our microscope could not only be used for research and quality control in academia and the private sector but also in schools at all levels,” Lytel said. “It could provide students with experience in measuring properties of complex objects and help prepare them for careers in various research, development and production settings.”

But Klar Scientific’s work is not done yet.

“Now, as always, we are raising money,” McCluskey said. “We are working with companies to solve their problems, which is great. The microscope is showing a lot of promise, so we’re going to continue to work with more and more companies and researchers. As far as a product goes, that is a year or two out. But we are definitely learning a lot and we’re improving things all the time.”

A world’s first
Size and affordability were also the driving factors behind Reza Moheimani’s development of a dime-sized, MEMS-based atomic force microscope. Research-grade AFMs can cost $500,000 or more, while the ones designed for education cost around $30,000.

“If AFMs could be in a price range that they could be found in every high school science lab, it would certainly have an impact on STEM education,” Moheimani, a mechanical engineering professor at the University of Texas at Dallas told Laboratory Equipment. “Another motivation is that AFMs are quite big and bulky, which makes them hard to use outside of a laboratory. A miniaturized AFM has the advantage that it could be packaged as a portable device. That is, instead of bringing the samples to the lab, one could take the AFM to the field.”

AFMs are used in research across scientific fields, including biology, chemistry, physics, materials science, nanotechnology, semiconductors, and more—in other words, they are extremely versatile. A typical AFM comprises an electromechanical device that interacts with the sample and performs imaging, electronics to drive the actuators and sensors, software for processing, and a control system. The electromechanical device uses a scanner that moves a tiny cantilever over a sample, using movement, lasers and optical sensors to precisely map the surface of a specific material, culminating in a 3-D image.

Alternatively, Moheimani’s MEMS-based AFM is about 1 square centimeter in size, or a little smaller than a dime. It is attached to a small printed circuit board, about half the size of a credit card, that contains circuitry, sensors and other miniaturized components.

“What we did was build a MEMS device that integrates the scanner with the cantilever,” Moheimani explained. “The scanner is typically heavy, bulky, difficult to build and expensive. They use piezoelectric actuators that require specialized low-noise, high-voltage amplifiers that themselves are very expensive. So, we integrated the sensing and actuation functionalities into the probe, which simplifies things quite a lot.”

In order to miniaturize such a complex instrument, Moheimani knew he needed to eliminate the optical scanner. So, on the prototype, he machined a layer of aluminum nitride onto the cantilever to act as the actuator. By measuring the electric current that goes into the piezoelectric actuator, one can determine the placement of the probe—thus allowing measurement capabilities.

“In a typical AFM, we put the sample on top of a scanner and move it while the probe interacts on top of the sample. The device we have designed puts the cantilever on the scanner itself. It’s just good engineering,” Moheimani said.

The MEMS-based mini-microscope is the culmination of seven years of research, with many complications to overcome along the way. And while the R&D phase was exhaustive and expensive, the fabrication phase will be the opposite. Batch fabrication of these devices is inexpensive—hundreds or thousands can be produced at once, driving the price down to only a few dollars per chip. Moheimani and his team have already adopted SOI-MEMS (silicon on insulator) technology that has the capability to produce an entire miniature AFM system for only a few thousand dollars.

“I would like to get the device to a point where we have all the functionalities of a conventional AFM,” Moheimani said. “That would change the research scene—even internationally. It opens up a lot of possibilities, and also ones we can’t imagine at the moment because this has never been done before. Once it’s out there in the market, it will make a lot of things possible.”

Earlier this year, Moheimani and his research team developed a first-generation prototype of the device. Since then, they have turned their attention toward demonstrating on-chip video-rate imaging. If successful, it would be a world first.

The design of the new device is completed, and it should be ready for testing before the end of this summer.

“Based on the results we have obtained thus far, we are confident that we can achieve video-rate scanning rates on our probe scanners,” Moheimani said. “We will still need to overcome significant hurdles as the new device will have a much smaller cantilever, which could make self-sensing and self-actuation with a single piezoelectric transducer quite challenging. But I hope a variant of this device can be commercialized and made available to researchers for a very reasonable price.

“This is one of those technologies where, as they say, ‘If you build it, they will come.’ We anticipate finding many applications as the technology matures,” he said.

Researchers designed a MEMS-based atomic force microscope that is about 1 square centimeter (center). Here, it is attached to a small printed circuit board that contains circuitry, sensors and other miniaturized components. Photo: UT Dallas

Novel detector, novel data
While some scientists need to reinvent the wheel for their research needs, others find they just need to adjust a hinge. Such was the case for Cornell physics professors Sol Gruner and David Muller when they designed the electron microscope pixel array detector (EMPAD). This custom detector can be fitted on old or new electron microscopes to enhance performance and resolution.

Although they didn’t design an entirely new microscope, Gruner and Muller’s detector proved so powerful that it caught the eye of FEI Co., a leading manufacturer of electron microscopes, only five months after the researchers’ work was published in the February 2016 issue of Microscopy and Microanalysis.

In the usual scanning transmission electron microscope (STEM), a narrow beam of electrons is fired down through a sample, scanning back and forth to produce an image. The detector underneath reads the varying intensity of electrons coming through and sends a signal that draws an image on a computer screen.

The EMPAD, however, goes beyond creating just the image. It comprises a 128 x 128 array of electron-sensitive pixels, each 150 microns square, bonded to an integrated circuit that reads out the signals—to not only form an image, but acquire a wealth of information. Its purpose is to detect the angles at which electrons emerge, as each electron hits a different pixel.

“What we’re able to do with this detector is record every scanning electron that makes it through the sample,” Muller explained to Laboratory Equipment. “If you have complete information about the scattering distribution both in real and reciprocal space, then in theory you could invert that information and recover fully the object that you scattered from. In other words, we can learn everything there is to know about the sample from the scattering experiment.”

By measuring where every electron is and where it’s going, researchers can build up a four-dimensional map of both position and momentum. Then, combining those pieces of information, scientists can see the atomic structure and forces inside a sample, such as magnetic field, strain, lattice constant, density, etc.

The EMPAD is novel in its speed, sensitivity and wide range of intensities it can record—from detecting a single electron to intense beams containing hundreds of thousands or even a million electrons. According to Muller, the detector images 1,000 times the dynamic range, and 100 times the speed of conventional sensors.

Due to this, the custom detector finds applications across industries. It’s ideal for looking at 2-D materials, especially magnetic and ferroelectric systems in the semiconductor industry. But it also brings improvements to life scientists, where it’s less-intense exposure does not damage living specimens. The EMPAD allows researchers to get a better look at the processes inside intact cells. And while it’s preliminary at this time, Gruner believes researchers may be able to look through thicker samples—normally a big limitation for biological materials.

“[The EMPAD] opened up a whole range of new opportunities, and we’ve discovered multiple new imaging modes that people didn’t think you could do before because the technology wasn’t there,” Muller said. “Now that we have it, there’s a huge number of new things we’ve been able to image we didn’t think we could previously.”

Muller and Gruner debuted their microscope at the Microscopy & Microanalysis meeting in July 2016. Numerous commercial vendors contacted Muller thereafter, with a few visiting the Cornell campus in the months that followed the trade show.

In the end, the scientists felt FEI was “the most capable of the batch,” having the most resources to commit to commercialization quickly.
A few months later, Thermo Fisher Scientific acquired FEI for $4.2 billion, explaining that electron microscopy is a new focus for the mega-company since it has a multitude of applications across industries.

FEI, now a division of Thermo Fisher Scientific, expects to complete the commercialization of the design and offer the EMPAD detector for new and retrofitted electron microscopes before year’s end.

“One of the big challenges as this is commercialized is, it’s all very well to let people take huge amounts of data in short times, but then the bottleneck becomes, how do you process it? We’ve had a lot of progress in developing software tools to handle these large datasets, let people visualize it. We’ve written a lot of software that lets you do that, and there’s a whole new way to think about these experiments,” Muller said.

Sol Gruner, left, and David Muller, right, designed a custom detector for electron microscopes that is being commercialized by FEI Co., this year. Photo: Chris Kitchen/Cornell University Photography