 Learning objectives include various types of microscopes that include dark field, microscopy, phase contrast, microscopy, fluorescent microscopy, transmission electron, microscopy, and scanning electron microscopy. There are many others also, but we would go through only a few of these. In dark field microscopy, it is a variation of light microscope, where the light source is used with a different condenser. It is a special condenser that bends the light and throws the light through the specimen in a way that light basically does not, if there is no specimen here, the light would not pass into the object. So when the specimen contains the organism there, this organism would basically bend the light, and that bent light would enter into the objective lens. And the advantage of this dark field microscopy is that the field, other than the organism would appear as dark. So when the light strikes the organism, the organism would be lit. It would be looking like white against a totally black background. And that is the reason we call it dark field microscopy. The advantage of using this is that we don't have to stain the cells. Sometimes we use such microscopes like dark field, and there is another one, phase contrast, without staining the cells, because staining with chemicals kills the organisms. If we want to capture them alive or without disturbing their morphology, this provides a better tool. Another one is phase contrast microscopy. It also does not use any stain, and we can study the cells in vitro as live cells. The principle behind this phase contrast microscopy is that wave light rays, they have the light travels in waves. If the two waves, they follow each other in all aspects, like this is the peak of the wave, and this is the trough of the wave. So a trough follows the trough, a peak follows the peak. So what we call these two waves are in phase with each other. Here the two waves, you see the trough of this wave is almost superimposed on the peak of this wave. So these two waves are at 180 degree out of phase with each other. The same principle of this phase change is used in phase contrast microscopy. The organism, when the light rays passes through the organism, it is retarded by almost one fourth of the wavelength. And then in this microscopy, which is phase contrast, we have a special plate, which is called phase plate, where you could see that the depth or the size of the plate is not exactly the same at all places. Here you see it is smaller than this portion. So the light that passes through the specimen, when it passes through thicker part of this phase plate, it is retarded even more. And here, because it has to pass through smaller distance of the plate, it is not retarded as much. So the trick is basically in this phase plate, which creates a contrast because of the variation in the phases of these two waves. So this wave, when this passes through the specimen and through the phase plate is out of phase, one half wavelength. And this difference creates the contrast. And the advantage again is that we can use this microscope without staining the cells. Another version of the light microscopy is a fluorescent microscope, where a dye, a special dye that what we call them fluorochromes, they are used. These are substances that absorb short wavelengths of light but then emit longer, which is visible, wavelength of light. As you can see here, this is the image taken from a fluorescent microscope. Fluorochromes, which are dyes or substances, they are attached to antibodies. As here, these are antibodies. They're attached with these antibodies. And then these antibodies, because they are specific against different organisms, those microbes could be stained specifically by these antibodies. And as these antibodies have these tags, these dyes associated with them, when we visualize these images under the microscope, they would fluoresce. They would emit different lights, colored lights. Here is the diagrammatic presentation of a fluorescent microscope. It, of course, uses a UV light, ultraviolet rays, which is kind of blue light. It passes through a filter. And then there's a special mirror, which does not allow a shorter wavelength, which is here. This is a UV light. And when it strikes this mirror, it gets into the specimen. And here is the fluorochrome lying here. And when it strikes the fluorochrome, it gets converted into longer wavelength. And then this longer wavelength can pass through this mirror. And then the image could be seen by the viewer or by the person who is looking through the eyepiece. Here is, again, the same thing. There's a source of light, a UV light, of course, passes that light through. And then there's a dichromatic lens mirror, which allows longer wavelengths to go through. And the observer can see that image through the eyepiece. Another version, as I mentioned, that resolution basically depends on the wavelength. Electron microscopy has way higher resolution power. Because electrons have very, very short wavelength compared with the light. But the principle of electron microscopy is exactly the same as light microscopy. So there is a source of light. There is a source of electrons. So this is the wavelength. This is the wavelength. The difference is that this wavelength is much shorter. And resolution, as a result, is much higher. Electrons pass through electromagnets, which basically act like lenses in the light microscope. So they focus the electron beam. And the electron beam then passes through the specimen. And then this image is created here. Because this is electron microscope, we cannot see the electrons with our eyes. We need a phosphorous screen, like a TV screen. So the image basically is created on a screen by the bombardment of these electrons. The advantage is that the image is much, much bigger. We can resolve very small structures. And we can see them in detail. Another version is scanning electron microscopy. Here the source of short wavelength is, again, the electrons. The only difference is that the electron, when it strikes the specimen, they are reflected off the specimen. And these are reflected off rays of electrons, or waves of electrons, or electrons. They are collected by a detector, special detector, and amplified through the electronic circuitry of the scanning electron microscopy. And the image is found, is a kind of three-dimensional image. This is the advantage of scanning electron microscope. So you can see the organism in 3D. Here is the image of a stephalorius. These are spherical cells. And as you can see, they look like beads, beautiful beads. In summary, there is a host of microscope available for us to use or to see different organisms at different magnifications, some with staining and some without staining.