Microscopy Flashcards
Light Microscopy
any type of microscope using visible light
Compound light microscopy
uses light and two or more lenses to obtain higher magnification
Total magnification
objective lens x ocular lens
Ocular lens
look through it
Stage
holds glass slide
Objective lens
magnification power of 4x, 10x, 40x, 100x
Condenser
lens, focuses light into a fine point
Diaphragm
controls amount of light entering the condenser
Light Source
underneath the stage and lens
Course/Fine Focus Knob
course first focuses, then fine focus for optimal clarity
Resolving power
(at least 0.4 nm) can distinguish between two points
-Shorter wavelengths of light provide greater resolution
-Infrared (red), pixelated resolution
Refractive Index
Measure of the light bending ability of a medium, refracts after passing through a specimen
Immersion oil
used to keep light from refracting
Brightfield Illumination
-White background
-Most commonly used
-Dark objects are visible against a bright background
Darkfield
-Black background
-Light objects are visible against a dark background
-opaque disk placed in condenser
-Only light reflected off of the specimen enters the objective lens
Phase Contrast
-Internal structures/texture
-inside topography of the specimen can be seen
Fluorescence
-Uses UV light (short wavelength)
-Absorb UV light and emit a longer wavelength (visible light)
-Cells may be stained with fluorescent dyes
Confocal
-Cells are stained with fluorochrome dyes
-Blue light (short wavelength) excites a single plane of a specimen
-Sliced into sliver, better solution
-illuminated, 3D image is constructed
Transmission Electron Microscopy (TEM)
-Beams of electrons passes through ultra thin sections of a specimen, passing through an electromagnetic and projector lens
-May be stained with heavy metal salts for contrast
-Magnifies 10,000 to 10,000,000x; resolution of 10pm
Scanning Electron Microscopy (SEM)
-An electron gun produces a beam of electrons that scans the surface of an entire specimen
-Secondary electrons emitted from the specimen produce a 3D image
-Magnifies objects 1,000 to 500,000x; resolution of 10 nm
Simple Stain
use of a single basic dye
Gram-positive
thick peptidoglycan layer
-purple (picks up lots of dye)
Gram-negative
thin peptidoglycan layer
-red (less dye)
Acid-Fast Stain
Binds only to bacteria that have a waxy material in their cell walls
Special Stains
Capsule Stain
Endospore Stain
Flagella Stain
