5-phase contrast and other types light microscopy

phase contrast microscopy

allows us to view live cells in natural form

colourless, unstained, untreated

principle of phase contrast

cellular components have differing RI

amplify subtle difference in RI between components into difference in amplitude and brightness

phase shift modification

condenser annulus and phase plate ring

condenser annulus

added to sub stage condenser

allows hollow cone of light to pass through condenser to specimen

phase plate ring

corresponding absorption ring added to objective

absorbs light from condenser annulus

phase shift application

view live cells, urinalysis, cell counting

darkfield microscopy

blocks direct central light rays so only light from specimen enters objective lens

only peripheral light focused onto specimen

darkfield modification

substage condenser with central stop/opaque disc

causes light waves to cross specimen rather than pass through

darkfield application

live unstained motile bacteria or protozoa, blood cells and tissue culture

best for observing external detail and edges, smooth reflective surfaces, High RI

polarizing microscopy

light rays vibrate and travel in one direction

vibrates only in one plane

polarizing modification

analyzer and polarizer

polarizer lens

between the light and specimen

analyzer lens

between specimen and oculars

polarizing microscopy application

for birefringent objects → crystals, fibers, microorganisms,

birefringent

object has 2 different refractive index, produces 2 images from refracted rays

fluorescent microscopy modifications

refined darkfield, exciter filter (selective for short wavelengths of light)

inverted microscopy

light source above specimen

condenser above specimen

objectives below specimen

inverted microscopy application

cell agglutination, cell or tissue cultures

stereomicroscopy

consists of 2 separate lens systems → 3d magnification

produces 2 images → one for each eye

used for dissection of small tissue