2) Observing The Microbial Cell

Question 1

Resolution

Answer 1

the smallest distance by which two objects can be separated and still be distinguished

Question 2

Which part of the human eye can the finest resolution of two separate point be perceived

Answer 2

Fovea

Question 3

Fovea

Answer 3

The portion of the retina where the photoreceptors are packed at the highest density

Question 4

Detection

Answer 4

The ability to determine the presence of an object

Question 5

Magnification

Answer 5

The increase in the apparent size of an image to resolve smaller separations between objects

Question 6

Eukaryotic Microbes

Answer 6

Protozoa, algae, fungi

10 - 100 um

Can be seen under a light microscope

Question 7

Prokaryotes

Answer 7

Bacteria, archaea

0.4 - 10 um

Subcellular structures too small to resolve by light microscopy

Question 8

Wavelength of Visible Light

Answer 8

400 - 750 nm

Question 9

For electromagnetic radiation to resolve an object 3 conditions must exist

Answer 9

1) Contrast between object and its medium
2) Wavelength smaller than the object
3) Magnification

Question 10

Absorption means

Answer 10

The photon’s energy is acquired by the absorbing object

Question 11

Reflection means

Answer 11

That the wavefront bounces off the surface of an object

Question 12

Refraction means

Answer 12

The bending of light as it enters a substance that slows its speed

Question 13

Scattering occurs

Answer 13

When the wavefront interacts with an object smaller than the wavelength of light

Question 14

Magnification requires the

Answer 14

Bending of light rays (refraction)

Question 15

How does refraction accomplish magnification

Answer 15

Refraction magnifies an image when light passes through a refractive material shaped so as to spread its rays

Question 16

Bright-Field Microscopy

Answer 16

Generates a dark image of an object over a light background

Question 17

To increase resolution

Answer 17

• Use shorter wavelength light
• Reduce contrast
• Use immersion oil
• Use wider lens closer to specimen
• Higher numerical aperture (NA)

Question 18

Compound microscope

Answer 18

A system of multiple lenses designed to correct or compensate for aberration

Question 19

Total magnification

Answer 19

Magnification of the ocular multiplied by that of the objective

Question 20

Wet mount

Answer 20

Placing a drop of water on a slide with coverslip

Advantages - Observation of cells in natural state

Disadvantages - Little contrast between cell and background
- Sample may dry out quickly

Question 21

Fixation

Answer 21

Cells are made to adhere to a slide in a fixed position

Question 22

Staining

Answer 22

• Cells are given a distinct color
• Most stains have conjugated double bonds or aromatic rings, as well as one or more positive charges

Question 23

The detection and resolution of cells under a microscope are enhanced by

Answer 23

Fixation and Staining

Question 24

Different kinds of stains

Answer 24

Simple stain and differential stain

Question 25

Simple Stain

Answer 25

Adds dark color specifically to cells, but not to the external medium or surrounding tissue (Methylene blue is the most commonly used stain)

Question 26

Differential Stain

Answer 26

Stains one kind of cell but not another (Most famous differential stain is the Gram stain)

Question 27

Gram Stain devised

Answer 27

In 1884 by Hans Christian Gram

Question 28

Gram-positive bacteria Gram-negative bacteria

Answer 28

- retain the crystal violet stain because of their thicker cell wall - bacteria do not

Question 29

Fluorescence microscopy

Answer 29

The specimen absorb light of a defined wavelength and then emits light of lower energy, thus the longer wavelength

Question 30

Excitation wavelength

Answer 30

The specimen absorbs light of a specific wavelength

Question 31

Emission wavelength

Answer 31

Emits light at a longer wavelength

Question 32

First to demonstrate the possibly of single-molecule tracking of fluorescent proteins in bacteria

Answer 32

William Moerner

Question 33

Dark-field Microscopy

Answer 33

Enables microbes to be visualized as halos of bright light against darkness

Question 34

Phase-contract microscopy (PCM)

Answer 34

Exploits difference in refractive index between the cytoplasm and the surrounding medium or between different organelles

Question 35

Differential Interference Contrast Microscopy (DIC)

Answer 35

Enhances contrast by superimposing an image of the specimen onto a second beam of light that generates interference fringes

Question 36

Electron microscopy (EM)

Answer 36

The foremost tool for observing the shapes of macro molecular structures

Question 37

Scanning probe microscopy

Answer 37

Images the contours of live bacteria

Question 38

X-ray crystallography

Answer 38

The tool of choice for atom-level detail of a macromolecule

Question 39

Two major types of electron microscopy

Answer 39

Transmission Electron Microscopy (TEM) Scanning Electron Microscopy (SEM)

Question 40

Transmission Electron Microscopy

Answer 40

- Electrons pass through the specimen - Reveals internal structures

Question 41

Scanning Electron Microscopy (SEM)

Answer 41

- Electrons scan the specimen surface - Reveals external features in 3D

Question 42

Cryo-electron microscopy (cryo-EM)

Answer 42

High-strength electron beams now permit low-temperature -Specimen does not require staining - Specimen must be flash-frozen

Question 43

Scanning probe microscopy (SPM)

Answer 43

Enables nanoscale observation of cell surfaces

Question 44

Atomic force microscope (AFM) (example if an SPM)

Answer 44

- It measures the van der Waals forces between electron shells of adjacent atoms of the cell surface and the sharp tip - It can be used to observe live bacteria in water or exposed to air

Question 45

Bright-field: Fluorescence: Dark-field: Phase-contrast:

Answer 45

- employs various stains - employs fluorophores for labeling - detects unresolved objects - exploits differences in refractive indices

Question 46

TEM: provides SEM: provides

Answer 46

- Internal details in 2D - External details in 3D

Question 47

Scanning probe microscopes (SPMs) include the atomic force microscope (AFM)

Answer 47

- Allow observation of living cells in water or in air

Question 48

Molecules can be visualized by

Answer 48

X-ray crystallography

Question 49

Which of the microscopes allows the best view of bacterial flagella during motility

Answer 49

Dark-field microscope

Question 50

Wavelength is the

Answer 50

Distance between one peak of a wave and the peak

Question 51

The height of each peak (or depth)

Answer 51

Amplitude

Question 52

Frequency of the wave

Answer 52

The rate of vibration of the wave or the number of wavelengths within a specified time period

Question 53

The variety of staining techniques (used for light microscopy)

Answer 53

Gram staining, acid-fast staining, capsule staining, endospores staining and flagella staining

Question 54

Commonly used light microscopes

Answer 54

Brightfield, darkfield, phase-contrast, differential interference contrasts fluorescence, confocal and two-photon microscopes