Microscopy Flashcards
what is the smallest object you can see with an objective lens
.3um
who invented the gram stain
by the Dutch
physician Hans Christian Gram
do Gram-positive retain the crystal violet stain?
bacteria retain the crystal violet
stain because of their thicker cell wall
do gram-negative bacteria retain the crystal violet stain
bacteria do not retain the crystal violet stain
what is Fluorescence Microscopy used for?
Used to view marine and pathogenic bacteria
how does Fluorescence Microscopy work?
The specimen absorbs
light of a defined wavelength, and then emits light of
lower energy, thus longer wavelength; that is, the
specimen fluoresces
Excitation and Emission
he specimen absorbs light of a specific wavelength
(the excitation wavelength), then emits light at a
longer wavelength (the emission wavelength)
what is a fluorophore
a fluorescent chemical compound
- its cell specificity can be determined in four ways:
- Chemical affinity
- Labeled antibodies
- DNA hybridization
- Gene fusion reporter
what did fluorescence microscopy reveal? (this isnt asking about the marine and pathogenic bacteria)
the presence of symbiotic
Bacteroides fragilis bacteria
Chemical imaging microscopy
uses mass spectrometry (analysis of
molecular fragments by mass) to visualize the distribution of
chemicals within living cells or microbial communities.
what is a high resolution method for chemical imaging
nanoscale secondary ion spectrometry (NanoSIMS)
how does nanoscale secondary ion spectrometry (NanoSIMS) work
NanoSIMS combines fluorescence in situ hybridization (FISH)
with mass spectral analysis of bacterial components labeled with
heavy isotopes.
- Allows detailed observation of the gut microbiome
which organism is responsible for syphilis
treponema pallidum
Phase-Contrast Microscopy
Superimposes refracted light and transmitted light
shifted out of phase
what does phase contrast microscopy reveal
- Reveals differences in refractive index as patterns of
light and dark - Can be used to view live cells and cellular organelles
Electron Microscopy
two major types:
- Transmission electron microscopy (TEM)
- Scanning electron microscopy (SEM)
- Transmission electron microscopy (TEM)
- Electrons pass through the specimen
- Reveals internal structures
Scanning electron microscopy (SEM)
- Electrons scan the specimen surface
- Reveals external features in 3D
The specimens for electron microscopy can be
prepared in several ways
- Embedded in a polymer for thin sections
- Microtome is used to cut slices
- Sprayed onto a copper grid
Tomography
In cryo-EM, or electron cryomicroscopy, the
specimen is flash-frozen
Cryo-electron tomography, or electron
cryotomography
avoids the need to physically slice
the sample for thin-section TEM
Scanning probe microscopy (SPM) enables…
nanoscale observation of cell surfaces
what is an example of SPM
The atomic force microscope (AFM)
what does the AFM measure
measures the van der Waals forces between
electron shells of adjacent atoms of the cell surface and the sharp tip
-It can be used to observe live bacteria in water or exposed to air (unlike electron microscopy)
