eTraining Introduction

Specimen Preparation

Leica Ultracut UCT

Histology

JEOL JSM-6400 SEM

Hitachi S-4700 FE-SEM

Hitachi FB-2000A FIB

Microanalysis

Veeco Dim 3000 AFM

Fluorescence Microscopes

Support

JEOL JSM-6400 SEM

SEM Equipment Status

Basic Science

A Scanning Electron Microscope (SEM) forms a two-dimensional image on a cathode ray tube by moving a beam of focused electrons across a specimen and collecting the signals produced by it.

Form and Function

SEMThe Scanning Electron Microscope uses electrons to illuminate a sample, instead of visible light used in optical microscopy. Since the wavelength of electrons is much smaller than that of visible light, the SEM is capable of imaging at much higher magnification than light microscopes. The high image resolution of scanning electron microscopy allows users to focus on features of the sample in a range of millimeters down to a few nanometers.

The subject of this tutorial, the JEOL JSM-6400 SEM, has a tungsten filament that, when heated, emits electrons which are then accelerated down through the column. The column is lined with electromagnetic lenses that are used to direct the electrons down the column and, ultimately, onto the surface of a specimen. The group of lenses contained in the column have one main goal: focusing the electron beam so that by the time it hits the specimen, the "spot size" (the diameter of the electron beam at the specimen) has decreased from ~50 microns to ~10 microns. That’s a magnification of about 5,000 times!

SEM images are formed when the electron beam position on the sample surface is synchronized with the CRT, or cathode ray tube viewing screen. At each location where the electron beam strikes the sample, an electron signal is used to produce contrast in the CRT image. The process of tracing or sweeping the electron beam across the sample surface is called rastering. When the user zooms in using the magnification knob, the electron beam must raster a smaller section of the sample. This results in a smaller area being analyzed, while the size of the CRT remains the same. This is the basic principle of magnification.

SEMs along with all electron microscopes operate in a vacuum environment, which removes air and its contaminants from the electron column. The vacuum environment in the SEM precludes the direct examination of living things, liquids or anything that contains liquids, thus samples must be dried before use in the system. If the system is operated in air, the high-energy electron beam turns the air's contaminants into carbon and "pastes" the products onto parts in the column. This process eventually spoils the optical qualities of the electron microscope. Additionally, a vacuum provides the needed insulation to prevent "arching" or jumping of high voltages in the electron gun.

Commonly used vacuum systems consist of an oil filled roughing pump to produce low vacuum and an oil diffusion pump (DP) for high vacuum. The roughing pump evacuates the microscope column from atmosphere to low vacuum. The DP then creates the higher vacuum needed for the SEM to operate under standard conditions, which is why both pumps are needed to produce the operating vacuum. Maintaining a vacuumed environment in the SEM is tough: any small hole in the system results in a vacuum leak and then the whole system must be shut down. Additionally, the DP requires a cold water source for cooling. SEM vacuum systems (and therefore cold water systems) are operated continuously and the maintenance required to keep up a dependable 24/7/365 vacuum and water source is difficult.

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