Cheatography
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Microscopes and Laboratory Basics
This is a draft cheat sheet. It is a work in progress and is not finished yet.
Bacterial Stains
Simple Stain |
-Single basic dye e.g. Methylene blue
- All bacteria take the color of the dye |
Differential Stain |
Primary Stain |
_ First dye used in the staining process
-Will initially stain all cells and then be removed from a subset |
Mordant |
Improves the ability of the primary stain to bind cells |
Decolorizer |
Removes the primary stain from a subset of cells |
Counterstain |
Second dye that stains decolorized cells |
Positive stain = purple color, negative charge |
Negative stain = pink stain, negative charge |
Types of Microscopes
Light Microscope (Bright-field) |
Unstained
Passes light directly through specimen
unless cell is naturally pigmented or artificially stained, image has little contrast
Stained
Staining with various dyes enhances contrast, but most staining procedures require that cells be fixed (preserved) |
1.Iris diaphragm
2.Condenser
3.Specimen
4.Objective lense
5.Eye
|
Dark-field Microscope |
-Light is projected at an angle to the surface, causing any variations to deflect light up into the camera
-Nothing is seen by the vision system if there are no aberrations on the surface
-dark background makes the organism glow |
1.Light source
2.Dark Field Patch Stop
3.Condenser Lens
4.Sample
5.Direct Illumination Block
6.Objective Lens
7.Eyes
|
Phase-contrast microscope |
Enhances contrast in unstained cells by amplifying variations in refractive index within specimen
-especially useful for examining living, unpigmented cells |
1.Light from source
2.Annular ring
3.Condenser
4.Specimen
5.Objective
6.Deflected Light
7.Phase Ring
8.Eye |
Fluorescent microscope (UV light) |
-Shows the location of specific molecules in the cell
-Fluorescent substances absorb UV radiation and emit visible light
-the fluorescing molecules may occur naturally in the specimen but more often are made by tagging the molecules of interest with fluorescent dyes or antibodies
Fluoresceins=Apple Green
Rhodamines=Orange red |
1.HBO lamp house
2.Excitation filter
3.Objective
4.Specimen
5.Dichroic beam splitter
6.Barrier Filter
7.Eyepiece |
Electron microscope |
-Uses electrons to scan specimens to create and magnify images
-Produces clear/detailed 3D images
-Specimen has to be held in place
-Unable to view living specimen
-up to 50,0000x magnification |
1.Electron
2.Condenser
3.Specimen
4.Objective lens
5.First image
6.Final image
7.Fluorescent screen
8.Eye |
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Bacterial Cell Structures
Outer Layers |
1. Glycocalyx (capsule,slime layer) |
2. Cell Wall |
Gram Positive Cell Wall
-Peptidoglycan
-teichoic acid and lipoteichoic acid
Gram Negative Cell Wall
-Outer membrane
--LPS, Lipoprotein, Porin protein
-Peptidoglycan
-Periplasmic space |
Cell Membrane |
Phospholipid Bilayer |
-fatty acids
-glycerol |
Cytoplasm |
Chromatin |
Ribosomes |
Cell Shapes |
Cocci |
-Circles |
Bacilli |
-Rods |
Spirilla |
-Spirals |
Types of Differential Stains
Stain Type |
Specific Dyes |
Purpose |
Outcome |
Gram Stain |
Uses crystal violet, Gram's iodine, ethanol (decolorizer) and safranin |
Used to distinguish cells by cell-wall type (gram-positive, gram-negative) |
Gram-positive cells stain purple/violet. Gram-negative cells stain pink. |
Acid-fast Stain |
After staining with basic fuchsin, acid-fast bacteria resist decolorization by acid-alcohol. Non acid-fast bacteria are counterstained with methylene blue. |
Used to distinguish acid-fast bacteria such as M. tuberculosis, from non-acid fast cells |
Acid-fast bacteria are red; non-acid fast cells are blue. |
Endospore Stain |
Uses heat to stain endospores with malachite green, then cell is washed and counterstained with safranin |
Used to distinguish organisms with endospores from those without; used to study the endospore. |
Endospores appear bluish-green; other structures appear pink to red. |
Flagella Stain |
Flagella are coated with tannic acid or potassium alum mordant, then stained using either pararosaline or basic fuchsin |
Used to view and study flagella in bacteria that have them |
Flagella are visible if present |
Capsule Stain |
Negatie staining with India ink or nigrosin is used to stain the background, leaving a clear area of the cell and the capsule. Counterstaining can be used to stain the cell while leaving the capsule clear |
Used to distinguish cells with capsules from those without |
Capsules appear clear or as halos if present |
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