Left: Images of fluorescent particles that are above, at and below (top to bottom) the vertical position of best focus of a microscope. Calibrating the effects of lens aberrations on the apparent shape and position of the particle images enables accurate measurement of the position in all three spatial dimensions using an ordinary optical microscope. Right: Tracking and combining information from many fluorescent particles on a tiny rotating gear tests the results of the new calibration and elucidates the motion of a complex microsystem in all three dimensions.
Credit: NIST
Conventional microscopes provide essential information about samples in two dimensions — the plane of the microscope slide. But flat is not all that. In many instances, information about the object in the third dimension — the axis perpendicular to the microscope slide — is just as important to measure.
For example, to understand the function of a biological sample, whether it is a strand of DNA, tissue, organ or microscopic organism, researchers would like to have as much information as they can get about the three-dimensional structure and motion of the object. Two-dimensional measurements yield an incomplete and sometimes unsatisfying understanding of the sample.
Now researchers at the National Institute of Standards and Technology (NIST) have found a way to convert a problem affecting nearly all optical microscopes — lens aberrations, which cause imperfect focusing of light — into a solution that enables conventional microscopes to accurately measure the positions of points of light on a sample in all three dimensions.
Although other methods have enabled microscopes to provide detailed information about three-dimensional structure, these strategies have tended to be expensive or require specialized knowledge. In one previous approach to measuring positions in the third dimension, researchers altered the optics of microscopes, for instance by adding ..
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