1	Using the Microscope ANHB 2212 – 2009 Avinash Bharadwaj
2	"This PowerPoint was originally created..." This PowerPoint was originally created for Human Structure and Development ANHB 2212, a second year science unit. Most of the students enrolled for this unit have a basic knowledge of histology but have not used a microscope before. This PowerPoint is not an exhaustive resource on the subject. In fact, some technical aspects like setting the illumination system have not been discussed in detail. Students outside ANHB 2212, whose area of study involves extensive and serious use of the light microscope and other varieties of microscopes are advised to refer to other resources. These instructions are specific for the Leica students’ microscope used in the histology lab in the School of Anatomy and Human Biology. The descriptions of controls may not be accurate for other models / makes of microscopes.
3	Know Your Microscope Your microscope has : a heavy base which also houses the lamp for illumination the condenser which directs a controlled beam of light to the slide which is placed on the… ‘stage’ the optical system. The optical system includes a set of objective lenses and two eyepieces. (There is more to the optical system, but it is not seen and you do not have anything to do with it!) Be careful! The microscope is delicate, with moving / detachable parts. If you must lift it, hold it firmly by the curved column (x) and support the base.
4	Below the Stage… Make sure the main power cord is connected to a power outlet and the switch is on! The Power switch (1) on the microscope is a red wheel on right side of the base. It slides with a click to turn on, and you can rotate it further to control the intensity of light. The condenser can be raised or lowered by turning a small black knob (2). You can gently turn the knob to try this.
5	Focusing Knobs
6	Objectives
7	Objectives
8	Eye pieces
9	The Rotating Head
10	Ready to Use it? Turn on the power – check the power cord and the main switch, then turn on the red button at the base. You will see a glow through the hole in the stage. Always begin with lowest power objective – 4x. Turn it into position if it is not already so. (Which also means that someone was careless! The previous user should have brought the 4x into position at the end of his/her work). Using the coarse adjustment, lower the stage.
11	Further… Check the condenser. We shall not go into the details of adjusting the condenser and centering it. Hopefully these arrangements will not be disturbed. (These adjustments are critical in advanced microscopy). For our purpose: low power (magnification) = low condenser high power (magnification) = high condenser. Now peep through the eyepieces. Adjust your eye position (distance between the eyes and the eyepieces) until you see a single circle of light when both eyes are open. It is very natural to close one eye to escape this little exercise, but these bad habits are hard to kill! Adjust the distance between the eyepieces. You may need to check the iris diaphragm if the light is very dim.
12	Mounting the Slide Select a slide from the box. One side of the slide has labels. This is the ‘upper’ surface. Observe it carefully – you will see a very thin piece of glass (called the coverslip) on this surface. If you are careful, you can feel it. When placing the slide on the stage the coverslip must face up! Failure to do so will result in damage to the slide and the objective when the high power objective is used! Secure the slide with the spring clip on the stage. Move the stage with the two screws and watch from one side (not through the tube!) the hole in the stage. A faint coloured blob will be seen in the glowing circle of light when the section on the slide is in the light path.
13	Observation Always begin with the scanner (4x). Keep the condenser low. Take the stage low down and raise it slowly as you see through the eyepieces. Stop when you see a reasonably focused image. Then use the fine adjustment to bring the image in sharp focus. Adjust the iris diaphragm if necessary until you have a good image – not too bright, not too dark. Study the slide at low magnification.
14	10x Objective Without disturbing the focus, bring the 10x objective into position. Do NOT hold the objective lens! Most likely, you will only need fine adjustment to bring the section in focus. If coarse adjustment is necessary, do this very carefully. Most likely, you will also need to reduce light by using the iris diaphragm. Notice that the ‘field of view’ is now smaller, and what you see is the central part of the field that was seen with the 4x lens.
15	40x Objective With the 10x lens select the area of interest and using the stage movement, bring it roughly in the centre of the field. Raise the condenser. You may also need to open the iris diaphragm. Very carefully, rotate the nose piece so that the 40x objective clicks in place. Be very careful – watch how close the lens is to the slide. Do NOT touch the coarse adjustment knob! Use only fine adjustment to get a sharp image. If you face difficulty in obtaining a sharp image, call a demonstrator! When you are done with the 40x lens return the setting to the 10x or 4x objective. Never take the slide off the stage with a 40x objective in place.
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