- Researchers develop a new technique to make optical microscope more accurate while imaging nanoparticles.
- They put an optical microscope under a microscope to achieve precision with high accuracy.
Optical microscopes are one of the simplest and oldest design of microscope developed in the 17th century. Over the past 20 years, researchers have made several improvements in conventional optical microscope design, making it suitable for detecting, tracking and imaging elements much smaller than their usual limit – about a half micrometer.
Optical microscopy is widely used in cryogenic physics, cell biology, microelectromechanical systems and nanoscale fabrication. It is also used for medical diagnosis when dealing with tissue fragments. Binocular microscopes are quite common in industrial use.
A new research [development of super-resolved fluorescence microscopy], which won the Nobel Prize in Chemistry 2014, has allowed scientists to analyze the motion of miniature motors at nanoscale-level, visualize formation of electrical connections between brain nerve cells and trace proteins in fertilized eggs. In short, this research has brought optical microscopy into the nanodimension.
Now, scientists at the National Institute of Standards and Technology has developed a technique to measure objects at these nanometer-scales with a whole new accuracy-level. The two key factors to localization microscopy are precision and accuracy. In order to obtain greater accuracy they have put an optical microscope under a microscope.
Before 2014, optical microscopes were not used for observing nano-scale objects, and that’s why they don’t have enough calibration to obtain accurate data at that particular scale.
An optical microscope might be precise, yet it could be exceptionally inaccurate at the same time, i.e. it might indicate the same location for a molecule, but because of some internal or external errors, this location could differ within a billionth (or even millionth) of a meter.
New Calibration Process
Calibration of microscope and correction of localization data ensure precision with accuracy
To deal with this issue, the researchers have built a new calibration process to closely analyze and fix imaging errors. To implement this process, they used reference materials with stable and well-known properties, which can be mass-produced and widely distributed to different labs.
Reference material is built using ion milling and electron beams, which create a pinhole aperture array through a thin platinum sheet. This allowed researchers to put the apertures five thousand nanometer apart, to within one nanometer of accuracy. This way, they created a way to measure accuracy of the aperture positions.
Aperture array with a spacing of 5,000 nm ± 1 nm | Credit: NIST
The array of apertures forms a point array to image objects. Since the lenses used in microscopes aren’t perfect, we see errors during imaging. These errors are responsible for altering apparent point-positions, which makes the apertures seem to be smaller and larger than the true spacing.
However, if one knows about the actual spacing, he can correct image errors and calibrate microscope to measure the position of objects with greater accuracy across a large field of view.
Why It’s Important?
Optical microscopes have become a common instrument in almost every lab, where it’s used to magnify a wide range biological specimens and particles. With current advances, they are becoming more capable and economical scientific equipment of the lights.
This research would allow scientists to implement calibrations in their own lab. According to the authors, the aperture arrays could enhance the microscopes’ ability [of accurately locating nanoparticles] by 10,000 fold.
In the future, researchers will be investigating aperture arrays in other kind of critical dimension metrology. They will integrate the aperture arrays with different samples, and fabricate other reference materials for localization microscopy.