2018 Biol1262 Tutorial 1 Week 2 Microscope Techniques.docx

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BIOL1262 – LIVING ORGANISMS 1 – SEMESTER 1 WRITTEN TUTORIAL DUE WEEK 4 during tutorial session. TUTORIAL 1 –WEEK 2 - Exploring with microscopes – MISCONCEPTION THE MAGNIFICATION AT WHICH THE SPECIMEN IS VIEWED USING MICROSCOPE IS THE MAGNIFICATION OF YOUR DRAWING. Demonstrator leads the discussion The scale of magnification or the magnification of you drawing    

is determined by dividing the size of your drawing by the actual size of the object drawn. Scale of Magnification of magnification = Size of Drawing / size of the specimen The size of your drawing can easily be determined by using a ruler to measure the drawing on your page. How is the size of the object (cell) viewed using a light microscope determined? To achieve this the light microscope must be calibrated. The two methods are explained below.

Field of view (FOV) method – The field of view is defined as the area that you see when you look through the eyepiece of the microscope. As magnification is increased, the field of view becomes smaller. You effectively see fewer cells, but each cell is more magnified. It should be noted that the size of the object is not changing, but it is size of the FOV that is changing. The diameter of the FOV is calculated for each magnification, using a stage micrometer or a ruler. Exercise: To measure /calibrate diameter of the FOV – will be done in the lab either week 3 or week 4.        

Use a micrometer slide – 1 mm = 1000 micrometers µm Place a stage micrometer on the microscope stage. Beginning with objective 4 – bring the scale on the stage micrometer into sharp focus Move the stage mechanism to position the 0 on the scale on the left edge of the field of view. Determine the diameter – How many 1 mm or 1000 µm are needed to cover the diameter at x10. Repeat for objectives X40 and 100 The micrometer slide can then be removed and replaced by a specimen slide. To determine the size of a cell viewed at X10 objective, estimate how many cells of a similar size is needed to cover the diameter of the field of

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view. If your estimate is 5 and your field of view 2 mm then the size of the cell is 2000 µm/5 = 400 µm. Repeat for each objective

Objective lens

Diameter of the field of view (µm)

Area of the scale of view (µm2)

X10 X40 X100

Ocular graticule method: The size of each division on the eyepiece (ocular) graticule which is located in the eyepiece of the microscope) is calibrated against the stage micrometer. After the ocular calibration is made for each objective the eyepiece graticule can then be used as a measure (or measuring stick) to calculate the size of the object viewed. Calibrating the microscope: WRITTEN TUTORIAL DUE WEEK 4 during tutorial session. Name: ID No: Tutorial date and time: Tutor: Answer questions 1-9 1. 2. 3. 4. 5.

Why are microscopes important to the study of biology? Define scale of magnification or magnification of the drawing. Define magnification and resolution so as to distinguish between them. What are the limitations of the light microscope? In a table format describe the function four parts of the microscope (you can choose any four parts. 6. Identify t least 4 things which can be done when using and storing the microscope to ensure that the microscope remains in good condition. 7. What is meant by the term par focal as it refers to the microscope or objective lens. Are the student microscopes in our lab par focal? 8. Describe the field of view method used to calibration of the light microscope so as to allow for the determination of the size of the object viewed at each objective lens. 9. Why is it necessary to calibrate at each objective? 10.Is the object changing in size as you go from one objective to the next? Explain what is happening.