THE BASICS OF LIGHT MICROSCOPY
The microscope that will be utilized is a typical light microscope fixed with phase contrast objective lenses and a phase contrast turret condenser. Such design permits the microscope to be utilized as an ordinary light microscope and with some slight adjustments as a phase contrast microscope. Though majority of the students have former familiarity with detecting and focusing on specimens with a compound microscope, only a number of them have knowledge with the particular light adjustments required to utilize the microscope. It is very important that the student must know how appropriately regulate the light on these microscopes. Not being able to do so will lead to poor image attributes and if the light is very far out of being regulated the viewer might not be able to observe anything in any way evidently.
Kohler illumination is necessary every time optimum resolution of aspect in opaque or colored objects is being wanted. It might be considered as the standard illumination for a compound microscope. It is necessary for phase contrast microscopy, microspectrophotometry, interference microscopy, photomicrography and cinematography, polarization microscopy, and fluorescence microscopy.
There are important reminders when using the microscope. First, check the voltage of the outlet as well as the voltage to be used for the functioning of the microscope, both must synchronize with each other to avoid short circuit. Never switch on the preset light intensity toggle switch located on the right side of the base. Such button overrides the voltage control and makes it not possible to regulate the brightness of the lamp. Never confuse the one condenser focusing knob located underneath the stage on the left side with the two sets of coarse and fine focusing knobs situated on both sides of the lower arm. Put the microscope slide with specimen on the stage, and arrange it so the specimen is above the light beam.
Make use of the two knobs hanging below the right side of the stage to rotate the slide around. Focus the microscope on the specimen. Lower the condenser gradually by turning the condenser focusing knob until an image of the edges of the field diaphragm is brought into sharp focus. When the condenser is being lowered, the beam of light will become smaller and sharper neighboring the edges. Gradually open the field diaphragm by turning its dial clockwise and the decagon will increase in size as seen above. Regulate the light intensity to a comfortable level by means of the sliding brightness control.
In order to observe a transparent specimen or a thin specimen of low visibility, Kohler illumination may possibly be so intense. The transparent specimen may vanish. Living cells or cell organelles are tricky to view in the traditional refracting microscope. In majority of the cells there is small or no colored substance at hand to add contrast to the image by light absorption at visible wavelengths. In cases of such specimens, phase contrast microscopy is the excellent choice.
The phase contrast microscope is a refracting microscope with two essential additions. Underneath the condenser lens is mounted an annular ring, which is a piece of black glass having a transparent ring that sends out a hollow cylinder of light to the condenser. There is mounted an annular phase plate in the objective. The ring on this plate is developed by means of covering the glass so the ring is not transparent. The shape and size of the phase ring is exactly the same to the image of the annular ring. Inside the condenser assembly underneath the stage is a rotating turret containing five various condenser lenses. Every lens is tagged with a number which can be seen on the phase contrast selection wheel through the small notch in the front of the assembly. The zero position is utilized with any objective lens once phase contrast is not desired. When utilizing the phase contrast, the viewer will continuously turn the phase contrast selection wheel so that its number and the equivalent condenser lens corresponds the magnification of the objective the viewer is using.
