AS Practicals Flashcards
Calibrating an eyepiece graticule
Calibrating an eyepiece graticule:
1). Set up microscope to required magnification
2). Place a stage graticule on the stage
3). Line up the two scales (the stage and eyepiece graticule)
4). Count the number of divisions on the eyepiece graticule equivalent to each division on the stage micrometer
Purpose of staining
Staining provides contrast between different structures within a sample
Differential staining
Differential staining is when multiple stains are used, and each stain binds to a specific cell structure, staining each structure differently so the structures can be easily identified
Aecetic orcein stain
Aecetic orcein binds to DNA and stains chromosomes dark red
Eosin stain
Eosin stains cytoplasm dark red or pink
Iodine stain
Iodine stains starch blue/black
Iodine in potassium iodide solution as a stain
Iodine in potassium iodide solution stains cellulose yellow
Haematoxylin stain
Haematoxylin stains RNA/DNA a purple/blue colour
Methylene blue
Methylene blue is an all-purpose stain, used often to stain DNA blue
Wet mount technique
Wet mount:
-Use pipette to add a drop of water on the slide
-Use tweezers to place specimen in water
-Put cover slip on
-Add stain (drop on the edge of the cover slip)
-Put paper towel on the opposite edge to absorb stain, drawing it under the coerslip and staining the specimen
Dry mount technique
Dry mount:
-Slice specimen thinly
-Use tweezers to pick up and put on the middle of the slide
-Put cover slip ontop of it
How to use a light microscope
Using a light microscope:
1). Clip the slide onto the stage
2). Select the lowest-powered objective lens
3). Use the coarse adjustment knob to move the objective lens to just above the slide
4). Look down eyepiece, adjust focus by moving the lens away from the slide using the fine adjustment knob until clear image appears
How to measure size of structures with a microscope
Measuring size:
Structures can be measured by counting the number of eyepiece divisions (once eyepiece graticule has been calibrated)
-To find actual length, the number of divisions should be multiplied by the length of one division
Why is resolution limited by diffraction?
Resolution is limited by diffraction of light as it passes through samples (and lenses). Diffraction is the tendency of light waves to spread as they pass close to physical structures such as those present in the specimens being studied.
Heart dissection method (external examination)
Heart dissection method: (external examination)
1). Feel the differences in thickness of the walls using fingers - identify left and right atria + ventricles (left = thicker, ventricles = thicker)
2). Identify 4 main vessels attached to heart (arteries = thick + rubbery, veins = thinner)
3). Identify coronary artery on external surface and where it connects to the aorta
Heart dissection method (internal examination)
Heart dissection method (internal examination):
1). Cut along left ventricle and through left atrium wall
2). Open up atrium and ventricles, look for tendinous chords and observe how they area attached to the atrioventricular valve
3). Look for the AV alve and Sl valves
4). Use ruler to measure thickness of walls
Plant dissection method
Plant dissection method:
1). Cut cross-section of stem thinly (transverse + longitudinal)
2). Place transverse section in water for 2 mins (stops them from drying out)
3). Lift sections of stem onto watch glass containing a stain (Eg tuolodine blue)
4). Rinse off excess stain
5). Place under microscope on slide with drop of water and coverlip, on lowest mag
Effect toludine blue on plant tissue
Effect toludine blue on plant tissue: (stains non-lignified and lignified tissue to allow contrast between xylem and phloem)
Phloem - red
Xylem - green/blue
Parenchyma = red/purple
mm to micrometer
1mm = 1000 micrometeres
Micrometeres to nanometers
1 micrometer = 1000 nanometers
Typical stage micrometer sizes
Stage micrometeres are typically 2mm long with subdivisions that are 10 micrometers apart
External examination of a heart dissection
External examination of a heart dissection:
-Place fingers inside 4 chambers to feel the differences in thickness of walls -> left should be thicker than right, ventricles thicker than atria
-Identify main vessels -> arteries are thick and rubbery, veins are thinner
-Identify coronary artery on external surface -> see where it connects to the aorta
Internal examination of a heart dissection
Internal examination of a heart dissection:
-Cut along left atrium, cut down apex to left ventricle
-Look for tendinous heart strings that are attached to the AV valve
-Look for AV and SL valves -> AV has two flaps, SL has half-moon shape
-Examine wall and internal structures of right atrium
-Measure thickness of walls and compare