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Imaging and diffraction characterize particles, from medical treatments to objects in outer space
Scientists and engineers must analyze particles to understand natural events such as volcanic eruptions, and to optimize industrial processes such as powder manufacturing. That analysis almost always includes the size of the particles, but other features also matter. “There is a huge uptick in customers who realize that size is not enough,” says Jason T. Noga, brand manager at Microtrac (Montgomeryville, Penn.). “They are looking to understand materials in ways that they couldn’t before.”
Many scientists use particle analysis in research and development, and gathering more data allows a deeper dive into a material’s properties. This might include statistics about outlier particles, as well as characterization of the average ones.
Some changes make more improvements than others in particle analysis. “In terms of developments that could be considered revolutionary, single-particle-counting techniques using the resonant mass measurement, RMM, and nanotracking particle analysis, NTA, have spurred new products in the marketplace, delivering additional information to researchers in the biopharmaceutical arena,” says Alan Rawle, applications manager at Malvern Instruments (Westborough, Mass.) and co-chair of the E56.02 Characterization Subcommittee of ASTM E56 Committee on Nanotechnology. He adds that “modern techniques find general acceptance by being incorporated into international standards and there are now a much larger number of relevant standards in ISO and ASTM than there were 5 or 10 years ago.”
Particle analysis based on light scattering started with experts, but now it is used more broadly. “Light-scattering techniques have moved from being pure research tools to quality control and particle property-predictor tools,” Rawle says. The technique is now used by astronomers and meteorologists, and in industrial sectors—from ceramics and cement to paints and pharmaceuticals.
Boosting biological research
A range of biological applications can benefit from better use of particle analysis. “Needs in the biopharmaceutical fields from oncology to exosomes and assessing the stability of protein formulations have been an area actively explored by many companies utilizing many techniques and leading to a plethora of product offerings,” Rawle says. But this is not the easiest application area. “Once living entities are incorporated into any system, then things can get much more complex from a scientific perspective,” notes Rawle.
That complexity means that biological research often requires more than one way to analyze particles. As Rawle says, “It is clear that a combination of tools is necessary to deal with the many complex issues and problems faced by researchers in the biological area.” He adds, “I would point to our Archimedes product, using the RMM technique, for protein aggregation studies, and our Nanosight, using the NTA technology, range of products for exosome applications as two important developments within the Malvern portfolio.”
Seeing the samples
Analyzing the same particle in more than one way comes in handy in some cases. Microtrac’s SI is a dynamic image-analysis instrument that works by directly integrating with a laser-diffraction sample loop. It can measure particles that are 5–1500 microns across, and can collect information on 24 size and shape parameters. The SI captures 100 images per second.
Some data capture and analysis systems, such as the SI, include software that helps researchers characterize particles. (Image courtesy of Microtrac.)
With the NanoSight system, researchers can examine individual particles or study the features of many particles. (Image courtesy of Malvern.)“This platform gets applied to lots of powders,” Noga says. “3-D imaging is becoming more popular, and those systems require each particle to be highly spherical.” He adds, “Imaging as the first principle measurement helps to optimize a powder and make sure that it’s very consistent.”
The precision required for a powder’s size distribution depends on the application, but some require the materials to meet tight standards. “In a regulated environment, like pharmaceuticals, the size specification is very important,” Noga says.
A team of scientists applied imaging-based particle analysis to study enterovirus 71 (EV71), which causes hand, foot and mouth disease.1 By reconstructing images from cryo-electron microscopy single-particle analysis, the scientists characterized the bond between EV71 and a monoclonal antibody, D5, that neutralizes the virus. The technology revealed crucial aspects of the structure at angstrom-level resolution. Giving insight into the details behind the findings, the authors wrote: “The structures reveal a bivalent binding pattern of D5 antibody across the icosahedral 2-fold axis on mature virion, suggesting that D5 binding may rigidify virions to prevent their conformational changes required for subsequent RNA release.” Studies like these can apply to many forms of infection. The authors concluded that their “results elucidate the structural basis for the binding and neutralization of EV71 by the broadly neutralizing antibody D5, thereby enhancing our understanding of antibody-based protection against EV71 infection.”
Some medically relevant studies use other imaging technologies to analyze particles. In protein-based medicines, aggregates of the drug and microscopic particles, such as metal pieces from production equipment, can reduce the effectiveness. “Microflow digital imaging (MDI) has become a widely accepted method for assessing subvisible particles in pharmaceutical formulations.”2 Using the FlowCam VS from Fluid Imaging Technologies (Scarborough, Me.), the scientists studied microscopic particles combined with a vaccine adjuvant called AddaVax. The scientists reported, “The instrument was capable of imaging all particle types assessed (polystyrene beads, borosilicate glass, cellulose, polyethylene protein aggregate mimics and lysozyme protein aggregates) at sizes greater than 5 μm in concentrations of AddaVax up to 50% (vol:vol).”
Benjamin Spaulding, laboratory manager at Fluid Imaging Technologies, says, “Compared to previous models, the FlowCam 8000 series of instruments offers an easier self-rinsing and cleaning system, a faster camera and a quick-connect flow-cell holder.” This platform can be used in many ways. “The FlowCam 8100, for example, is a valuable tool for drug researchers for analyzing particles during drug research and development,” Spaulding says. “In the testing phase, FlowCam provides the exact size and shape of biological material and compounds, making it an important tool for stability studies.” The FlowCam 8100 can be combined with the FlowCam ALH autosampler system. “The FlowCam ALH system includes programmable pre-analysis sample preparation, including mixing, heating and cooling. Additionally, the system can automatically analyze up to 96 separate samples without operator intervention,” Spaulding adds. “Finally, its vertical setup also saves bench space.”
Sizing up the software
“Lots of the recent advances come in the software,” Noga says. “With taking pictures, the camera technology and the processing power of the computer allow more parameters to be compared, and you can look more deeply into the data.”
The software can also provide in-process controls over manufacturing. “You can set up alerts for when the particles on a production line become, say, flaky or are just starting to vary and you need a change in the process,” Noga explains. “This lets the customer be more proactive.”
Rawle also points to computers as a big advance in the technology. “This has allowed more complex calculations to be carried out more quickly,” he says. “In light scattering, for example, the sizes and shapes of water crystals in the upper atmosphere can be assessed using models such as the discrete dipole approximation and T-matrix.” This kind of modeling can be used in weather prediction, quantifying global warming and, as Rawle points out, “interplanetary studies, including the composition of Saturn’s rings and asteroids.”
Adding more advanced algorithms to particle analysis through computation plus new methods of data collection can help researchers explore the universe and save lives.
References
- Ye, X.; Fan, C. et al. Structural basis for recognition of human enterovirus 71 by a bivalent broadly neutralizing monoclonal antibody. PloS Pathology 2016, 12, e1005454.
- Frahm, G.E.; Pochopsky, A.W. et al. Evaluation of microflow digital imaging particle analysis for sub-visible particles formulated with an opaque vaccine adjuvant. PLoS One, 2016, 11, e0150229.
Mike May is a freelance writer and editor living in Ohio. He can be reached at [email protected].