Micro
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| 4 General Principles of Microscopy | 1. Wavelength of radiation
2. Magnification
3. Resolution
4. Contrast
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| Wavelength | Smaller wavelengths of radiation result in enhanced microscopy
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| Magnification | Is the apparent increase in size of object
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| Resolution | *Is the ability to distinguish objects that are close together
*Smaller wavelengths = increased resolution
*Resolution distance = 0.61 x wavelength/numerical aperture
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| Contrast | Differences in intensity between two objects or between an object and its background
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| Resolution | increased resolving power
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| Bright-Field Microscopes - Simple | Contain a single magnifying lens – similar to magnifying glass & Leeuwenhoek’s microscopes
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| Bright-Field Microscopes - Compound | *Series of lenses for magnification
* Light passes through specimen into objective lens
* Oil immersion lens increases resolution
*1 or 2 ocular lenses
* Total magnification = obj len × ocul lens
* Most have condenser lens
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| Dark-Field Microscopes | * Best for observing pale objects
* Only light rays scattered by specimen enter objective lens
* Specimen appears light against dark background
* Increases contrast/enables observation of more details
* Useful for examining small or colorless cells
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| Phase Microscope | Used to examine living organisms or specimens that would be damaged/altered by attaching them to slides or staining
2 types:
Phase-contrast microscope (good 4 cilia & flagella)
Diff interference contrast microscope (creates a 3D shadowed appearance)
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| Fluorescence Microscopes | * Direct UV light source at specimen
* Specimen radiates energy back as a longer, visible wavelength (fluorescent)
* UV light increases resolution/contrast
* Used in immunofluorescence to identify pathogens and to make visible a variety of proteins
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| Confocal Microscopes | Use fluorescent dyes
Use UV lasers to illuminate fluorescent chemicals in a single plane
Resolution is increased because emitted light passes through pinhole aperture
Computer constructs 3-D image from individual digitized images
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| Electron Microscopy | * Transmission electron microscopes TEM (2D images of internal cell structures)
* Scanning electron microscopes SEM (3D images of cell surface features)
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| Probe Microscopy | 100,000,000×
2 types
Scanning tunneling microscopes STM
Measures electron flow (tunneling current) to&frm the probe&spec surf
Resolution of 0.01nm
Atomic force microscopes AFM
Pass probe on the spec surf
viruses, bacteria, proteins, amino acids.
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| Principles of Staining | Dyes used stains are usually salts tht are composed of a + charged cation & a – charged anion
Acidic dyes stain alkaline structures (neg stains)
Basic dyes stain acidic structures (simple stains)
Most cells/ bacteria are neg charged
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| Chromophore | colored portion of the dye
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| Differential Stains | Use more than one dye
Used to distinguish between diff cells, chemicals, structures
Common differ stains:
Gram
Acid-fast
Endospore
Histological
Bacillus & Clostridium produce endospores
*Highly resistant to desiccation, heat, chemicals
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| Special Stains | Simple stains used to identify specific microbial structures
Special stains include:
* Negative stains: are negatively charged and repelled by a negatively charged bacterium
*Flagellar stains
*Fluorescent stains
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| Taxonomy | consists of classification, nomenclature, and identification
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| Carolus Linnaeus | His system classified organisms based on common characteristics- 1st name ALWAYS cap
Grouped organisms that can successfully interbreed into categories called species - 2nd name ALWAYS lc
Used binomial nomenclature (latin)
Eg; Streptococcus pneumoniae
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| 5 taxonomy kingdoms | Animalia, Plantae, Fungi, Protista, and Prokaryotae
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| 3 domains as determined by ribosomal nucleotide sequences: | Eukarya, Bacteria, and Archaea
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| Taxonomic and Identifying Characteristics | Physical characteristics
Biochemical tests
Serological tests (Blood)
Phage typing (Virus)
Analysis of nucleic acids
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| MicroScan | All bacteria on 1 sheet for testing - quicker results
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| Phage Typing | Bacteriophages (phages) are viruses that infect bacterial cells
Phages are specific for the hosts they infect
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| Analysis of Nucleic Acids | Nucleic acid sequence can be used to classify and identify microbes
Prokaryotic taxonomy now includes the G + C content of an organism's DNA
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| Dichotomous keys | Series of paired statements where only one of two "either/or" choices applies to any particular organism
Key directs user to another pair of statements or provides name of organism
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| Which of the following is smallest? a. Decimeter B. Millimeter C. Nanometer D. Micrometer | Nanometer
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| A nanometer is __________ that a micrometer. a. 10 times larger b. 10 times smaller c. 1000 times larger d. 1000 times smaller | 1000 times smaller
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| Resolution is best described as the __________. a. ability to view something that is small. b. ability to magnify a specimen c. ability to distinguish between 2 adjacent objects d. difference between 2 waves of electromagnetic radiation | Ability to distinguish between 2 adjacent objects
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| Curved glass lenses __________ light. a. refract b. bend c. magnify d. both a and b | both a and b
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| Which of the following factors is important in making an image appear larger? a. thickness of the lens b. curvature of the lens c. speed of the light passing through the lens d. all of the above | all of the above
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| Which of the following is different between light microscopy and transmission electron microscopy? a. magnification b. resolution c. wavelengths d. all of the above | all of the above
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| Which of the following types of microscopes produces a three-dimensional image with a shadowed appearance? a. simple microscope b. differential interference contrast microscope c. fluorescence microscope d. transmission electron microscope | b
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| Which of the following microscopes combines the greatest magnification with the best resolution? a. confocal microscope b. phase-contrast microscope c. dark-field microscope d. bright-field microscope | confocal microscope
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| Negative stains such as eosin are also called __________. a. capsule stains b. endospore stains c. simple stains d. acid-fast stains | capsule stains
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| In the binomial system of nomenclature, which term is always in lowercase letters? a. kingdom b. domain c. genus d. specific epithet | specific epithet
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| different objective | 4x scanning
10X low power
40X high dry
100X oil immersion
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| If an object magnifies 40X, and each binocular lens magnifies 15X, the total magnification of the object being viewed is ___________. | 600x
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| Immersion oil __________ the numerical aperture, which ___________ resolution because _________ light rays are involved. | increases, increases, more
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| What refers to differences in intensity between two objects? | contrast
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| What are Koch's Postulates? | A series of steps that must be taken to prove the cause of any infectious disease
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| The type of fixation developed by Koch for bacteria is | heat fixation
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| Catonic chromophore said such as methylene blue ionically bond to | Negatively charged chemicals such as DNA and proteins
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