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Microscopy

Microbiology

QuestionAnswer
Most common length units in microbiology are : micrometer (µm) nanometer (nm)
Compound light microscope uses: Visible light
Compound light microscope contains two lenses : Ocular • 10X (most common)
– Objective (in a revolver) 4X, scanning • 10X, low-power • 40X, dry high-power • 100X, Oil-immersion high power
4X scanning
10X low power
40X dry high power
100X oil-immersion high power
Magnification = ocular 10 x times objective
Resolution or (resolving power) is the ability to distinguish fine detail between two points
Resolution of brightfield microscope 0.2um
Human eye resolution 0.2 mm
• Resolution of electron microscope 0.5nm
– The shorter the wave length the______________. the higher the resolution
Long wevelength light
Short wevelength (electrons) electrons
field of vision for 4x
Long wevelength light
Short wevelength (electrons) electrons
Field of vision for 4x 5
Field of vision for 10x 2
Field of vision for 40x 0.5
Field of vision for 100x 0.2
Numerical aperture (NA) setting Match NA of objective lens in use with NA of iris diaphragm in condenser
Numerical aperture (NA) setting gives the specimen : Better resolution and contrast
Refraction is the: the change in direction of a wave (light) due to a change in its transmission medium (air, glass, water, etc.)
Without immersion oil most light is refracted and lost
Refractive index Light-bending ability of a medium
Electron beam is located on the electron microscope
The Light microscope 's highest magnification is : 1000 x
The Light microscope's Resolution is : 0.2um
The Light microscope's Radiation source is Visible light
The Light microscope's Lenses are: Glass
The electron's microscope's highest magnification is: >100,000
The electron's microscope's resolution is 0.5nm
The electron's microscope's radiation source is electrons
The electron's microscope's lenses are: electromagnet
Preparation of bacterial specimens for brightfield microscopy include: 1. Smear microorganisms onto slide 2. Fixing 3. Flood with stain 4. Washing 5. Drying 6. Microscope observation
Smear A thin film of a solution of microbes on a slide
What are the most common heat-fixing? Alcohol or heat
What does heat-fixing do? – Attaches microorganisms to slide – Kills microorganisms – Preserves structures with minimal distortion
Chromophore is the positive ion • Chromophore attaches to negatively charged surfaces Basic dyes
Chromophore is the negative ion • Chromophore attaches to positively charged surfaces Acidic dyes
The colored ion is named the chromophore
Positive chromophore the dye: penetrates the cell
Negative Chromophore the dye: does not penetrate the cell
Basic (positive chromophore) typse of dyes are: – Methylene blue – Fuchsin – Crystal violet – Safranin
Acid dyes (negative chromophore) types of dyes are: Nigrosin (China/India ink) – Eosin – Rose bengal
Simple stains are rarely used and: Highlight the entire microorganism Cell shape and Arrangement of cells
Only one dye (basic) is used: simple stain
Differential Stains Used to differentiate bacteria – Gram stain – Acid fast
T/F Different bacteria respond differently to different stains true
Christian Gram developed gram staining in 1884
not part of the original stain by Christian Gram) Counterstain
No lipid coat gram positive
Lipid coat present gram negative
Gram positive cells have a thick peptidoglycan coat that: –Prevents CV-I complexes from exiting the cell
Gram negative cells have a lipid coat and a thin peptidoglycan coat and Alcohol disrupts the lipids, and the thin peptidoglycan coat does not stop CV-I complexes from exiting
T/F Some bacteria stain poorly or not at all. true.
Why do older cultures of gram-positive bacteria give inconsistent results? because of the degradation of peptidoglycans
In acid fast staining Bacteria of the genus___________has a coat of waxy material. Mycobacterium
After staining with carbol-fuchsin (red), the red color persists after washing with acid-alcohol (acid fast)
Carbol-fuchsin is washed off from bacteria without the waxy coat (non-acid fast) Red color disapears – Cell stained with a conunterstain, usually methylene blue
What are the names of 3 type of special staining? Negative staining, Endospore staining,Flagella staining
Negative Staining: – Stains background, not cell
Endospore staining: Heat to drive malachite green into endospore
Flagella staining: – Use of a mordant to widen flagella
Created by: 1155187441193384