Microscopy Classification Flashcards
(32 cards)
what are the 4 main principles of microscopy
wavelength of radiation, magnification, resolution, contrast
the wavelength of visible light is
400-700 nm
the wavelength of waves of moving electrons depends on
the voltage of an electron beam
what is magnification
is an apparent increase in size
what is resolution
the ability to distinguish objects that are close together
what is contrast
differences in intensity between 2 objects or an object and its background
what sort of light increases contrast
light that is in phase
how is total magnification found
magnification of objective lens * magnification of ocular lens
what are the 3 kinds of microscopy
- bright field
- dark field
- phase: for living organisms
differential contrast microscope - nomarski for 3D
how do fluorescent microscopes work
• Direct UV light source at specimen
• Specimen radiates energy back as a longer, visible wavelength
• UV light increases resolution and contrast
• Some cells are naturally fluorescent; others must be stained
• Used in immunofluorescence to identify pathogens and to
make visible a variety of proteins
how is mycobacterium tubercolosis stained
with fluorescent dye auramine O
how do confocal microscopes work
• Use fluorescent dyes
• Use UV lasers to illuminate fluorescent chemicals in a
single plane
• Resolution increased because emitted light passes
through pinhole aperture
• Computer constructs 3-D image from digitized images
how does an electron microscope work and what are the 2 kinds
Much higher resolution because of much shorter wavelength of electrons: 10,000-100,000 X
magnification (viruses, cell’s ultrastructure). Staining with heavy metals.
- Transmission Electron microscopes (TEM): ultramicrotome. Dead organisms.
- Scanning Electron Microscopes (SEM): whole specimen, coated with platinum or gold. It uses magnetic lenses to focus a beam of primary electrons
Dead organisms.
what is probe microscopy
it magifies more than 100 M times, done by scanning tunneling microscopes for surface details. And atomic force microscopes
bright field microscope is used for
observe stained killed specimens
dark field is used for
living colorless unstained
phase contrast is used for
living organisms
Nomarski is used for
living organisms
fluorescence is used for
as a quick diagnostic tool
confocal microscopy is used for
detailed structures of cells
electron microscopes transmission is used for
observation of viruses and small bacteria
electron microscope scanning is used for
to see surface structures
give the general concepts of staining
Dyes used as stains are usually salts
Chromophore is the colored portion of the dye
Acidic dyes stain alkaline structures
Basic dyes stain acidic structures
More common since most cells are negatively charged
name 3 simple stains
crystal violet, safranin, methylene blue
