Submitted to: Scanning
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: February 28, 1997
Publication Date: N/A
Interpretive Summary: Attempting to understand many biological problems in agriculture requires observation of cells from crops, animals or pathogens at very high magnifications in a special instrument known as a transmission electron microscope (TEM). To observe the cells with this instrument, they must be cut into very thin sections (less than 1/25,000 of an inch). As a result of having to observe these thin sections, reconstructing the three dimensional nature of the cell and understanding how it is being affected is very difficult. To solve this problem, researchers developed an instrument called a high voltage TEM which allows them to observe sections as thick as 1/5,000 of an inch. Unfortunately the cost of this instrument is in excess of $ 2,000,000 and very few of them exist in the U.S. We have developed a technique that allows us to use a different type of microscope, known as a field emission scanning electron microscope, to observe thick sections of biological tissue. Use of this instrument, which is considerably less expensive and found in many research laboratories, will allow researchers to examine thick sections of biological tissues and thereby increase our understanding of how cells are affected by pathogenic or climatic factors.
A field emission (FE) scanning electron microscope (SEM) equipped with the standard secondary electron detector was used to image thin (70 to 90 nm) and thick (1 to 3um) sections of biological material that were chemically fixed, dehydrated and embedded in resin. The preparation procedures and subsequent staining of the sections were identical to those commonly used to prepare thin sections of biological material for observation with the transmission electron microscope (TEM). The results suggested that the heavy metals, namely osmium, uranium and lead, that were used for postfixation and staining of the tissue provided an adequate secondary electron signal that enabled imaging of the cells and organelles that were present in the sections. The FESEM was also used to image sections of tissues that were selectively stained using cytochemical and immunocytochemical techiques. Futhermore, thick sections could also be imaged in the secondary electron mode. Stereo pairs of thick sections were easily recorded and provided images that approached those normally associated with high voltage transmission electron microscopy.