BIS104 Edwards F06 MT1 - Page 5

BIS104 Edwards F06 MT1 - Page 5 - calculate the limit of...

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Page 4 Name ________________________ 2. The formula for calculating resolution is: d = (0.61) (wavelength) / (n) (sin alpha). a. Give a one sentence definition of d (resolution) as it applies to microscopy. The resolving power of a microscope lens refers to the minimum distance of separation between two objects in a specimen so that they may be seen as two distinct objects. (small “d” = high resolution) b. Suppose that you have in your research lab a microscope outfitted with ocular (eyepiece) lenses capable of 10x magnification and 2 objective lenses, one at 10x and one at 40x (oil immersion). What is the maximal achievable magnification with this microscope? (10x mag from oculars) x (40x mag from the highest power objective ) = 400x (400 diameters of mag) c. Now suppose your microscope light source has a wavelength of 450 nm. Using your 40x objective lens (numerical aperture = 1.4), and a drop of immersion oil between the objective lens and the specimen,
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Unformatted text preview: calculate the limit of resolution of this microscope. d = (0.61) x (450 nm) / 1.4 = approx 190 – 200 nm (so, two objects closer together than 200 nm would appear as one) d. The set up described above provides a certain magnification and resolution, but what about contrast ( visibility)? Describe two different technical approaches that can be used with the light microscope to improve specimen contrast . (1 ) Alter the specimen by treating it with selective stains or vital dyes that will bind to and highlight specific organelles and cell components. Increase in contrast increases visibility. (2) Alter the optics of the light microscope. Use phase- contrast or differential interference contrast (DIC) microscope to convert diffracted light passing through the specimen to light vs dark contrast. This may provide good contrast (visibility) in living specimens without adding dyes. Demo...
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