Introduction to Microscopy - Part Two

por **Admin** Ter Nov 05, 2019 10:55 am

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Functions of the Major Parts of a Optical Microscope [1-3]

• Light source and Condenser: project a parallel beam of light onto the sample for illumination - Condenser gathers light and concentrates it into a cone of light that illuminates the specimen with uniform intensity over the entire view field [6].

• Sample stage with X-Y movement: sample is placed on the stage and different part of the sample can be viewed due to the X-Y movement capability

• Fine/ Focusing knobs: since the distance between objective and eyepiece is fixed, focusing is achieved by moving the sample relative to the objective lens

• Objectives: does the main part of magnification and resolves the fine details on the samples (mo ~ 10 – 100)

• Eyepiece: forms a further magnified virtual image which can be observed directly with eyes (me ~ 10)

• Beam splitter and camera: allow a permanent record of the real image from the objective be made on film (for modern research microscope).

Magnification

• To visualize any tiny object 0.1mm there is limit of unaided Human eye.

• To see micro organism much smaller than 0.1mm a system required convex lenses.

• When parallel rays pass through convex lens get converged at point called as Focal length (f) of the lens.

• Hence, a lens with shorter focal length will have higher magnification power.

• In compound microscope it will be i.e 10X, f=16mm; 40X, f=4 mm; 100X, f= 1.8mm.

• Image produced by objective lens falls on the eyepiece lens serve as object.

• Image formed in the eyepiece is perceived by one eye. The overall magnification is given as the product of the lenses and the distance over which the image is projected: M = (D *M1 * M2)/ 250 mm – (1) Where, D = projection (tube) length (usually = 250 mm); M1, M2 = magnification of objective and ocular. 250 mm = minimum distance of distinct vision for 20/20 eyes.

Resolution

• The ability to see two close objects as two distinct objects called resolution.

• The limit of resolution is the closest distance between two points at which the points still can be distinguished as separate entities.

• Magnification should be coupled with good resolution to visualize small micro organisms, else magnification alone will produce an inconclusive or blurred image.

• The resolution (Fig. 2.a-c) can be calculated as shown in Eqn. 2. Maximum resolution: R = (0.061λ)/ N.A – (2) Where, 0.61 is a geometrical term, based on the average 20-20 eye, λ = wavelength of illumination, N.A. = Numerical Aperture - The N.A. is a measure of the light gathering capabilities of an objective lens. N.A. = n sin α – (3) Where, n = index of refraction of medium, α = < subtended by the lens ra

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Introduction to Microscopy - Part Two Numeri11

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Introduction to Microscopy - Part Two Prof_p13