Lecture 4 Flashcards Preview

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Flashcards in Lecture 4 Deck (54):
1

2 main types of microscopes

-light microscopes
-electron microscopes

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Light microscope

any microscope that uses visible light to observe specimens

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Compound light microscope uses 2 lens to observe specimens

-objective lens
-ocular lens

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Objective lens

located closest to specimen, magnifies specimen, 10x-100x

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Ocular lens

lens located w/in the eye piece, magnifies the specimen 10x

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Total magnification

objective lens X ocular lens

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Example of calculating magnification for compound microscope

ocular lens= 10x
objective lens= 100x
total magnification = 10 x 100 = 1000

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Resolution

ability to distinguish fine detail and structure

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If a microscope has a 6nm resolving power then it can distinguish two points if they are at least ...

6nm apart

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In order for 2 objects to be seen as distinct from on another...

light must pass in between them, and be short enough wavelength otherwise only seen a 1 objects

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General principle of resolution

shorter the wavelength go light the better resolution will be (inverse relationship)

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Electron microscopes

uses beam of electrons, waves of electrons are much shorter than light waves, achieves greater resolution, also higher magnification

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Smallest object visible w/ human eye

0.10 mm

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Compound microscope can be used to view an object as small as..

0.20μm

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Electron microscope can be used to view objects as small as

0.20nm

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Trasmission electron microscope (TEM)

used to examine internal cell structure

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Electron beams of TEM do not penetrate thick cell wall which requires..

cell to be cut using process called thin sectioning, the thin sections must be stained before viewed to improve contrast

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Scanning electron microscopy (SEM)

only be used to view surface of object, specimen must be coated w/ thin film of heavy metal, wide range of magnification to be achieved

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Scanning tunneling microscopy (STM)

most powerful, visualize atoms, uses thin metal probe to scan specimens revealing surface irregularities

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Chromophore

stain composed of -'ve or +'ve charged ions and one type of either +'ve or -'ve is coloured

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1st step in staining procedure

thin film of material called smear which contains the microorganism of interest is 'smeared' on slide

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2nd step of staining porcedure

sample is then find by passing though flame

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3rd step of staining procedure

stain is then applied to the sample

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4th step of staining porcedure

stain is remover from sample by rising

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Final step in staining procedure

stained sample is now viewed under microscope

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How to stains work

outer surface of bacteria carries net -'ve charge, -'ve charge attracts +'ve charge and repels -'ve charge which strains use to manipulate

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Positive stains

+'ve charged stains will agree to -'ve changed bacterium (basic), bacteria will appear color of stain while background is clear

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Negative stain

-'ve stain will be repelled by -'ve charged bacterium (acid) bacteria will appear clear, background will appear colored

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Simple stains

single colored basic dye used

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Basic dye

when the coloured ion is the ='ve ion which then binds to the organism making it appear in the color of dye (mordant used sometimes ti increase intensity)

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Differential stains

used to differentiate in between different yes of bacteria because the stains react differently with different types of bacteria

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Differential stains are very important for..

diagnostics because it exploits different in cell wall structure and composition

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Examples of differential stains

gram stain and acid fast

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Gram stain dtermines

whether a bacterium of interest is gram +'ve or -'ve

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2 primary types of bacteria based on among of peptidoglycan in the cell wall

-gram positive
-gram negative

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Gram positive have a ____ layer of peptidoglycan then gram negative

thicker

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Anything gram positive = ____ and gram negative = _____

purple; pink

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Any eukaryotic cells in gram stain will be ...

pink because negative membrane

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During gram stain __ stains are used with wash(s) between them

2

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If a bacteria is gram positive then..

the thicker layer of peptidoglycan will hold the first purple stain

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If the bacteria are gram negative them..

the thin peptidoglycan will not hold the first stain w/ violet and it will be washed away, resulting in second stain (safranin) ticking and making bacteria pink

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Acid fast stain

stain binds strongly bacteria that contain waxy cell wall component.

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Acid fast stain is used to identify all bacteria that fall under the genus..

mycobacterium

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Waxy cell wall of mycobacterium retains the..

carbol fushin die that is added

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Counterstain for acid fast

methylene blue leaves tissues and non-acidfast bacteria blue

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3 types of special stains

-capsular
-flagella
-endospore

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Capsule stain

reveals presence of thick polysaccharide layer outside of bacterial cell

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Capsule presence indicates ..

bacterium w increase virulence or increase ability to cause disease in comparison to same bacteria that lacks capsule

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During capsule stain background is coloured with..

a negative stain such as nigrosin which makes it black

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During capsule stain bacterial cell is stained with..

a positive cell stain such as safranin making it colored

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During capsule stain capsule ..

doesn't take up dye so remains colorless with halo around cell

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Endospore stain

primary stain colors endospore green, counterstain safranin colors rest pink

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Example of endospore forming bacteria is

bacillus anthracis

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Flagella stain

flagella are too small to beset under light microscope w/o stain, a mordant and stain together works to increase thickness of flagella to it is visible