The following pictures show an ancient and modern type of microscopes:
Aim:
To investigate a typical compound light microscope throw:
- Identifying its parts and specifications
- Practically testing its functions
- Preparing slides and testing them
- Focusing and preparing images
- To use an oil immersion objective
- Learning how to take care of it
Equipment:
- Typical light microscope
- Slides
- Cover slip
Method:
The method used to investigate the typical light microscope is a following:
General method
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Make sure all backpacks are out of the aisles before you get a microscope. You always carry the microscope with one hand on the Arm and the other hand on the Base. (Carry it close to your body).
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Remove the cover of the microscope, and plug the microscope in, then place the excess cord on the table. You always have to be careful with placing the excess cord so it doesn’t hang over the edge, and then your knee may get caught on it. Also so it doesn’t fall down and crash. Before placing the slide you have to make sure that the ocular and the objective lenses are cleaned by using a lens tissue. (The following picture shows us out to handle the microscope and how to place it on the table).
- Turn the illuminator on and open the diaphragm.
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Look through the ocular. Starting with the scanning objective as close to the stage as possible. (Always start and end with Low Power. Then we can go ahead and place the slide on the stage. Then we can use the coarse adjustment knob until speciemen is on sharp focus. (The following picture shows a specimen on a slide).
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Readjust the light intensity directly on specimen directly over the center of the glass circle on the stage. Then find the specimen as soon as you look through the eyepiece. Note: If you see a dark line that goes part way across the field of view, try turning the . That dark line is a pointer that will be very valuable when you want to point out something to your lab partner, teacher etc. (The following picture shows some slides under microscope that are under different types of magnifications.
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If, you are on lower power, then lower the objective lens to the lowest point, and then focus using first the coarse knob, then the fine adjustment knob. The specimen will be in focus when the lower power objective is close to the lowest point, so start there and focus by slowly raising the lens. If you can’t get a view of any thing at all, then focus using the coarse knob, then switch to the fine focus knob to specify the image.
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Adjust the Diaphragm as you look through the eyepiece, and you will see that more details are visible when you’re in less light. Too much light will give the specimen a washed-out appearance.
8) Once you have found the specimen on Low Power (100x), center the specimen in your field of view, then, without changing the focus knobs, switch it to High Power. If you don’t center the specimen you will lose it when you switch to High Power.
9) Once you have it on High Power remember that you only use the fine focus knob. The High Power Objective (430x) is very close to the slide. Use of the coarse focus knob will scratch the lens, and crack the slide.
Oil immersion objective
If you use an oil immersion lens. You have to use oil to lubricate the lens. Also, the oil is needed to help gather enough light to actually see through the lens. In a more detailed way we can start with racking down the stage slightly, then move the objective aside and place one drop of the correct type of oil on the center of the cover slip. Rotate into position the oil immersion objective and while looking to it from one side of the microscope, move the course adjustment control until the objective touches the crop of oil on the slide. Then rack up the condenser to close below the slide. Use the fine adjustment control to bring the objective into focus. Then adjust the aperture of the diaphragm until suitable illumination is obtained. Finally wipe of the oil with the lens tissue.
Making Slides
- Prepare a clean peace of slide, the latter a drop of water on the slide.
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Gather a thin slice/piece of whatever your specimen is. If your specimen is too thick, then the cover slip will wobble on top of the sample like the following picture:
- Place one drop of water directly over the specimen. If you put too much water over the specimen, then the cover slip will float on top of the water, making it harder to view the specimens as they float past the field of view.
4) Place the cover slip at a 45-degree angle (approximately), with one edge touching the water drop, and let go.
Taking care of the microscope
The worst enemies of the microscope are careless users, for example air, grease from fingers, liquid and chemicals from slides, oil, etc. So to avoid this there are certain ways to clean up the microscope. Starting with regularly cleaning the lenses with (xylene) and lens cleaning tissue. Always do not screw the lens components. You also have to carry the microscope carefully (with two hands one, one holding the arm and the other hand on the base. Wiping off split liquids from the stage immediately, and if contaminated liquids are split then use strong disinfectant solution (Lysol).
Results:
An image of a typical “Compound Light Microscope”:
The following table is shows different types of adjustments:
The following pictures are cropped pieces from a microscope:
Discussion of Results:
The compound microscope uses lenses and light to enlarge the image and is also called an optical or light microscope (vs. an electron microscope). The simplest optical microscope is the magnifying glass and is good to about ten times (10X) magnification. The compound microscope has two systems of lenses for greater magnification, 1) the ocular, or eyepiece lens that one looks into and 2) the objective lens, or the lens closest to the object. Before purchasing or using a microscope, it is important to know the functions of each part. This information is presented below. Links will take you to additional information and images.
Eyepiece Lens (ocular lens): the lens at the top that you look through. They are usually 10X or 15X power.
Tube: Connects the eyepiece to the objective lenses.
Arm: Supports the tube and connects it to the base.
Base: The bottom of the microscope, used for support.
Illuminator: A steady light source used in place of a mirror. If the microscope has a mirror, it is used to reflect light from an external light source up through the bottom of the stage.
Stage: The flat platform where you place your slides. Stage clips hold the slides in place. If your microscope has a mechanical stage, you will be able to move the slide around by turning two knobs. One moves it left and right; the other moves it up and down.
Revolving Nosepiece or Turret: This is the part that holds two or more objective lenses and can be rotated to easily change power.
Objective Lenses: Usually you will find 3 or 4 objective lenses on a microscope. They almost always consist of 4X, 10X, 40X and 100X powers. When coupled with a 10X (most common) eyepiece lens, we get total magnifications of 40X (4X times 10X), 100X, 400X and 1000X. To have good resolution at 1000X, you will need a relatively sophisticated microscope with an Abbe condenser. The shortest lens is the lowest power; the longest one is the lens with the greatest power. Lenses are color-coded and if built to DIN standards are interchangeable between microscopes. The high power objective lenses on all National microscopes are retractable (i.e. 40XR). This means that if they hit a slide, the end of the lens will push in (spring loaded) thereby protecting the lens and the slide. All microscopes have achromatic, par centered, par focal lenses.
Rack Stop: This is an adjustment that determines how close the objective lens can get to the slide. It is set at the factory and keeps students from cranking the high power objective lens down into the slide and breaking things. You would only need to adjust this if you were using very thin slides and you weren't able to focus on the specimen at high power.
Condenser Lens: The purpose of the condenser lens is to focus the light onto the specimen. Condenser lenses are most useful at the highest powers (400X and above). Microscopes with in stage condenser lenses render a sharper image than those with no lens (at 400X). If your microscope has a maximum power of 400X, you will get the maximum benefit by using a condenser lenses rated at 0.65 NA or greater. 0.65 NA condenser lenses may be mounted in the stage and work quite well. A big advantage to a stage-mounted lens is that there is one less focusing item to deal with. If you go to 1000X then you should have a condenser lens with an N.A. of 1.25 or greater. All of our 1000X microscopes use 1.25 Abbe condenser lens systems. The Abbe condenser lens can be moved up and down. It is set very close to the slide at 1000X and moved further away at the lower powers.
Diaphragm or Iris: Many microscopes have a rotating disk under the stage. This diaphragm has different sized holes and is used to vary the intensity and size of the cone of light that is projected upward into the slide. There is no set rule regarding which setting to use for a particular power. Rather, the setting is a function of the transparency of the specimen, the degree of contrast you desire and the particular objective lens in use.
