Microscopy PDF

Microscopy Learning Outcomes • Identify the components of the light microscope • Describe basic microscopy terms ➢ Magnification ➢ Resolution • Compare the types of microscopes Practical • Operate the microscope Light Microscope Simple LM Compound LM Light Microscope Parts of the Microsco...

Microscopy Learning Outcomes • Identify the components of the light microscope • Describe basic microscopy terms ➢ Magnification ➢ Resolution • Compare the types of microscopes Practical • Operate the microscope Light Microscope Simple LM Compound LM Light Microscope Parts of the Microscope Remove dust cover Parts of the Microscope • Eyepiece: – May also be called the ocular lens – Magnifies by 10X, 12X, or .. 20X – Sometimes contains a pointer that can be seen as you look into the eyepiece. – Has an adjustment for inter-pupillary distance X X • Objectives: – Magnify material being examined – 4X • magnifies 4 times – 10X • magnifies 10X – High dry • magnifies 40X – Oil immersion • Magnifies, 60X or 100X Total magnification = magnifying power of eyepiece X magnifying power of objective Objectives • Nose piece: – Part of microscope to which the objectives are attached – Rotates to allow for the changing of objectives to increase or decrease magnification • Arm: – Part of microscope to which the nosepiece is attached – A secure part of the microscope to hold on to when the microscope is being carried. Stage: Platform on which the slide rests Slide holder: Secures slide on stage Stage control knobs: Move the slide on stage Objective Lens 100x Coverslip Specimen Slide Stage Oil Coarse focusing knob: Coarse focusing Fine focusing knob: Precise focusing Substage condenser - concentrates light & directs it through opening in microscope stage towards the specimen Iris diaphragm: - regulates the amount of light passing through the condenser - must be opened Iris diaphragm lever: ❑ adjusts the iris diaphragm to increase or decrease microscope resolution Condenser control knob: ❑ raises & lowers condenser Illuminator Light source Illuminator control: Controls the intensity of light Serves as the on/off switch for some microscopes Base: provides support for microscope Resolution (Resolving Power) ❑ Defined as the minimum distance by which 2 points must be separated to be seen as 2 separate points rather than a fused image ❑ This is a measure of the clarity of the image ❑ A microscope with a high resolving power will allow 2 small objects which are close together to be seen as 2 distinct objects Ability to distinguish (resolve) two close-together points as separate High resolving power Actual Low resolving power Magnification • The magnification refers to the ratio of image size to object size • The total magnification is the eyepiece magnification multiplied by the objective magnification • The maximum magnification of a light microscope is x1500 Multi-head Microscope Virtual Microscopy Virtual Microscopy https://histologyguide.com/ Focusing the microscope 1. Rotate the nosepiece so that the 4X objective clicks into place. 2. Place the slide on the stage. Pull out the slide clip, slip the slide in place and gently release the slide clip. The slide should be held under tension in place. 3. Center the specimen over the aperture. 4. Using the coarse focus to raise the stage slowly until you reach a stop (while using 4x). (Watch from the side to make sure that the objective does not smash into the slide!) 5. Look through the right eyepiece only and lower the stage using the coarse focus until the specimen is focused. Focusing the microscope 6. Adjust with the fine focus knob until specimen is sharp and clear. 7. Now adjust the binocularity by changing the distance between eyepieces so that they match the distance between your eyes (IPD) and you see one circle with both eyes. 8. Cover (or close) your left eye and confirm the specimen is in sharp focus with the fine focus knob. 9. Cover (or close) your right eye and use the left eye to adjust the focus by rotating the telescoping knob on the eyepiece. 10. Your specimen should be in focus for your both eyes.