Re: What are the limitations of a light microscope?

In a light microscope---or any optical device, including telescopes, eyeballs, or even windows---you encounter something called diffraction. Visible light behaves like a wave, with a wavelength of about 300 to 900 nanometers. We say that light "diffracts" when its wavelike behavior makes it bend around obstacles, instead of traveling in straight lines. To visualize this, think of ocean waves. They don't travel strictly in straight lines: they will wash through a pier and go around the pylons (the pylons don't cast "shadows"), or come through a narrow gap in a sea wall and "spread out" into the bay. If the gap (or obstacle) is much larger than the wavelength of the incoming wave, the spreading-out will be smaller. If the gap is smaller than the wavelength, then the spreading-out will be very large.

Now, in a light microscope, the light waves will spread out whenever they pass through a lens, an aperture, or any sort of obstruction. The primary lens in most microscopes is of course much bigger than 300-900 nanometers! So the angle by which light waves diverge is really quite small. However, it is not zero---the light waves do spread out a little bit, and the result is that the visual field is always a bit blurry. It is impossible for an ordinary light microscope to avoid this problem, so they can never see structures smaller than about 500 nm.

With an electron microscope, you have the same issues---electrons are waves also---but the wavelength is much much smaller. An electron microscope can have a resolution of 0.1 nm!

Since electrons are charged particles, you can't just shoot them through the air like you do with light; an electron microscope must operate in a vacuum chamber. The way it takes an image is a bit different than a light microscope: you scan an electron beam across a surface, and measure how many electrons bounce off to reach a detector. Electrons bouncing off of a high, protruding feature are more likely to reach the detector, so these features appear "bright"; elecrons bouncing off of a pit or crevice will not reach the detector, so these areas appear dark. (That's very different than the bright and dark spots in an optical image, which depend on light absorption rather than on 3D structure). In order for the electrons to interact properly, the surface of the object must be conductive. In order to image an insect or a cell, it must be dried out and coated with gold or carbon. This is one of the main disadvantages of the electron microscope: samples must be killed, dried, and prepared. They're also expensive and delicate.

There are other differences between light microscopes and electron microscopes. Although the resolution of a light microscope is limited, it is good enough to look at cells and at tissues. Since it can see properties like color, polarization, and opacity, a light microscope can sort of see chemical substructure of a cell, while the electron microscope mostly sees shapes and densities. For example, a very useful procedure in cell biology is to try to manipulate a cell's DNA to make it produce "green fluorescent protein". Under a light microscope, the cell's color tells you whether the protein is there or not.

Hope this is helpful! There are many places online to read about optical microscopes and electron microscopes. There is a good page about the scanning electron microscope at the Boston Museum of Science---but check your local science museum or university, they may have their own! You might also be interested to learn more about diffraction---try this page to get started, or any intro physics textbook.

-Ben