TABLE Eye Versus Instrument Resolution1.3 Distance Between Resolvable Points Human eye 0.2 mm Bright-field microscope 0.2 m SEM 2.5 nm TEM Theoretical 0.05 nm Tissue section 1.0 nm Atomic force microscopy 50.0 pm
The resolving power of the human eye—that is, the distance by which two objects must be separated to be seen as two objects (0.2 mm)—is determined by the spacing of the photoreceptor cells in the retina. The role of a microscope is to magnify an image to a level at which the retina can resolve the information that would otherwise be below its limit of resolution. Table 1.3 compares the resolution of the eye with that of various instruments.
Resolving power is the ability of a microscope lens or optical system to produce separate images of closely positioned objects.
Resolution depends not only on the optical system but also on the wavelength of the light source and other factors such as specimen thickness, quality of fixation, and staining intensity. With light of wavelength 540 nm (see Table 1.1), a green-filtered light to which the eye is extremely sensitive, and with appropriate objective and condenser lenses, the greatest attainable resolving power of a bright-field microscope would be about 0.2 m (see Folder 1.4, page 12 for method of calculation). This is the theoretical resolution and, as mentioned, depends on all conditions being optimal. The ocular or eyepiece lens magnifies the image produced by the objective lens, but it cannot increase resolution.
Various light microscopes are available for general and specialized use in modern biologic research. Their differences are based largely on such factors as the wavelength of specimen illumination, physical alteration of the light coming to or leaving the specimen, and specific analytic processes that can be applied to the final image. These instruments and their applications are described briefly in this section.
The microscope used by most students and researchers is the bright-field microscope.
The bright-field microscope is the direct descendant of the microscopes that became widely available in the 1800s and opened the first major era of histologic research. The bright-field microscope (Fig. 1.9) essentially consists of:  a light source for illumination of the specimen (e.g., a substage lamp), a condenser lens to focus the beam of light at the level of the specimen, a stage on which the slide or other specimen is placed, an objective lens to gather the light that has passed through the specimen, and an ocular lens (or a pair of ocular lenses in the more commonly used binocular microscopes) through which the image formed by the objective lens may be examined directly.
cussed earlier are used.
Examination of a Histologic Slide Preparation in the Light Microscope
Organs are three-dimensional, whereas histologic sections are only two-dimensional.
As discussed in the earlier “Tissue Preparation” section, every tissue sample prepared for light microscopic examination must be sliced into thin sections. Thus, two-dimensional sections are obtained from an original three-dimensional sample of tissue. One of the most challenging aspects for students using the microscope to study histology is the ability to mentally reconstruct the “missing” third dimension.
For example, slices in different planes through an orange are shown in Figure 1.10. Note that each cut surface (indicated by the dotted line) of the whole orange reveals different sizes and surface patterns, depending on the orientation of the cut. Thus, it is important when observing a given section cut through the orange to be able to mentally reconstruct the organization of the structure and its component parts. An example of a histologic structure—in this case, a kidney renal corpuscle—is shown as it would appear in different sectional planes (Fig. 1.10). Note the marked difference in each section of the renal corpuscle. By examining a number of such two-dimensional sections, it is possible to create the three-dimensional configuration of the examined structure.
Artifacts in histologic slides can be generated in all stages of tissue preparation.