The Use Of Advanced Biological Microscope To Study Biological Materials
Inspite of many years of using the microscope, the advanced biological microscope remains one of the most useful and common instruments in biological research and teaching. A large portion of biological exploration involves imaging objects we cannot see with our naked eye. New technologies applied to light microscopy are today changing our ability to understand the behavior of cells and even molecules. This article is designed to explain some basic principles of advanced biological microscope light microscopy.
There are three major problems a biologist is confronted with when using an advanced biological microscope. These are the magnification, resolution and contrast of an advanced biological microscope. The first major problem is advanced biological microscope magnification. This is what one first thinks about when one sits down to use an advanced biological microscope. The lenses of the advanced biological microscope form images that are magnified versions of the objects placed at their focus. The magnification of an advanced biological microscope is simply the magnification of the eyepiece ocular times the magnification of the objective. Most commonly for a compound microscope as opposed to a dissecting microscope this will range from about 40X 50X up to 1000X. The biological microscope eyepieces typically have a magnification of 10X, the smallest objective usually magnifies 45X, and the most powerful objective typically magnifies 100X. Even the 1000X magnification in the very best advanced biological microscope does not enable us to see very tiny objects such macromolecules and atoms that biologists would like to see.
The information content of the magnified image from an advanced biological microscope refers to the microscope resolution. In particular, it refers to our ability to discern fine details in the specimen when viewed under the microscope. It is typically defined as the ability of advanced biological microscope to produce an image in which two objects separated by a short distance are shown as two objects rather than as single, blurred object. Owing to the diffraction of light of an advanced biological microscope that comes from each pointing the specimen, the laws of physics tell us that the resolving power of the advanced biological microscope is limited.
A different method of overcoming the limited contrast of biological specimens from an advanced biological microscope is to make use of interference microscopy. Light behaves as if it were composed of waves. While these waves are transmitted about equally well through different parts of advanced biological microscope specimens like, for example, if the amplitude of the waves are unaltered, these specimens do induce changes in the phase of light waves. That is, the timing relationship of the peaks and troughs of the wave can be altered. These phase differences arise because light assumes a different velocity as it passes through differing biological structures of differing refractive index, light passing through the nucleus of a cell is retarded compared with the light passing through the nearby cytoplasm of this same cell. When the light waves re emerge on the opposite side of the cell, the phase of the wave passing through the cytoplasm is different from that passing through the nucleus. While the eye is insensitive to these phase differences, interference microscopy for advanced biological microscope converts these phase differences to differences in intensity to which the eye is sensitive. The basis of this kind of microscopy is the interference that occurs between light waves of different phases.
Another method for producing contrast in biological specimens is by use of dark field microscopy. In dark field microscopy, the objectives lenses of the advanced biological microscope are the same as those used for bright field or for phase, but the advanced biological microscope condenser is special. The dark field condenser of an advanced biological microscope illuminates the preparation with an annulus ring of light, but, in contrast to phase microscopy, none of the undiffracted light enters the objective lens of the advanced biological microscope. The only light that enters the objective of an advanced biological microscope is that is diffracted by the specimen. As a result, in dark field, the specimen is dark except for structures in the specimen that scatter light into the objective lens of an advanced biological microscope. These structures appear bright against the dark background when viewed under the microscope. While dark field microscopy is not utilized in most laboratory exercises, it is an important technique of to be aware of when using advanced biological microscope.