- greater resolving power and magnification - 10,000x to 100,000x - detailed view of bacteria, viruses, ultrastructure, and large atoms

Chapter 4 Microscopy, Staining, and Classification Flashcards by jessica quintos

Principles of Microscopy (4)

1 wavelength
2 magnification
3 resolution
4 contrast

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Wavelength

length between 2 crests of light

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visible light spectrum

(purple, short) 400nm - 700nm (red, long)

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Magnification

process of enlarging something in appearance

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Resolution

how close 2 points in the image can be before they are no longer seen as 2 separate points

clarity of an image

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Contrast

difference of intensity bw 2 objects or an object and its background

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Simple Bright-Field Microscope

light microscopy
single magnifying lens
similar to magnifying glass
leeuwenhoek used simple microscope

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Compound Bright-Field Microscope

light microscopy
series of lenses for magnification
light passes thru specimen into objective lens
oil immersion lens increases lenses
1 or 2 ocular lenses
may have condenser lights

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Total Magnification

(objective lens)x(ocular lens)

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Dark-Field Microscopes

light microscopy
best for observing pale/unstained objects
only light rays scattered by specimen enter the objective lens
specimen appears light against dark background
increases contrasts and enables observation of more details
ideal for visualizing spirochetes

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Electron Microscope

greater resolving power and magnification
10,000x to 100,000x
detailed view of bacteria, viruses, ultrastructure, and large atoms

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Light Microscopes

-cannot resolve images closer than 200nm

2 types: bright-field and dark-field

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2 types of Electron Microscopes

1 Transmission Electron Microscopes (TEM)

2 Scanning Electron Microscopes (SEM)

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Transmission Electron Microscopes (TEM)

2D images

- cross sectional

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Scanning Electron Microscopes (SEM)

3D images

- topographies

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Probe Microscopy

-magnifies more than 100,000,000x (one hundred million)

2 Types of Probe Microscopy

1 Scanning Tunneling Microscopes

2 Atomic Force Microscopes

Principles of Staining

staining increases contrast and resolution by coloring specimens with stains/dyes
smear of microorganism (thin film) made prior to staining
microbiological stains contain chromophore
acidic dyes stain alkaline structures
basic dyes stain acidic structures

3 types of Staining

1 Simple Stains
2 Differential Stains
3 Special Stains

4 types of Differential Stains

1 Gram Stain
2 Acid-Fast Stain
3 Endospores
4 Histological Stains

2 types of Special Stains

1 Negative (Capsule) Stain
2 Flagellar Stain

Gram Stain Procedure

5 steps

(+)...............................(-)
(...)......heat fix.......(...)
(pu)..crystal violet..(pu)
(pu).gram's iodine.(pu)
(pu)....decolorize....(...)
(p)red counterstain(pi)

Acid-Fast Stain

acid-fast organisms have wax-like, impermeable cell walls

- mycobacterium i.e. tuberculosis

Endospore Stain

staining and visualizing endospores
endospores can survive even the harshest conditions
allows us to see if endospores are present
ex) anthrax, tetanus

Histological Stains

- hematoxylin and eosin (H&E) -purp | - gomor methenamine silver (GMS) - blue

Negative (Capsule) Stain

- "special stain" - detects the presence of bacterial capsules - see bacterium right away

Flagellar Stain

- "special stain" | - flagella is normally too thin/small to be seen w normal staining technique

Staining for Electron Microscopes

- requires stains that contain heavy metals - helps absorb and reflect electrons - stains may bind moles in specimens or backgrounds - heavy metal will kill specimen

Linnaeus

- classified organisms based on common characteristics - "species" - "binomial nomenclature - only proposed 2 kingdoms (now there's 5)

Species

- created by Linnaeus | - organisms that can successfully interbreed

Taxonomic identifying characteristics

``` 1 physical characteristics 2 biochemical tests 3 serological tests 4 phage typing 5 analysis of nucleus acids ```

Taxonomy order

``` 8 Domain 7 Kingdom 6 Phylum 5 Class 4 Order 3 Family 2 Genus 1 Species ```

Agglutination Test

a type of serological test - neg result = antibodies won't clump - pos result = clumping of antibodies and bacteria

Phage Typing

- a bacterial lawn and a bacteriophage is applied to see if the bacteria is susceptible to phage - if there are plaques (holes on the lawn), then the bacteria is susceptible