Lecture 4 Flashcards
What are the two main types of microscopes
- Light
2. Electron
What are 3 types of electron microscopes?
- Transmission Electron Microscopy
- Scanning Electron Microscopy
- Scanning Tunneling Microscopy
What is a light microscope?
Any microscope that uses visible light to observe specimens
What are the two lenses that are used in a light microscope?
- Objective Lens
2. Ocular Lens
Where is a objective lens located and what does it do?
Closest to the slide
Magnifies the specimen 10-100x
Where is the ocular lens located and what does it do?
Within the eyepiece of the microscope
Magnifies specimen 10x
How do you figure out the magnification of a compound microscope?
Objective lens magnification x ocular lens magnification
What is resolution?
The ability to distinguish fine detail and structure, two 2 points a certain distance apart
In order for two objects to be seen as distinct from one another what must happen
Light must pass between
Short wavelengths= ________ energy =_______ resolution
Higher energy, higher resolution
What is the general principle of resolution
The shorter the wavelength of the light, the better the resolution to be
How do electron microscopes work?
Use a beam of electrons instead of light
Are electron waves longer or shorter than light waves?
Much shorter
Can electron microscopes achieve less or greater resolution compared to a light microscope
Much greater (up to 500,000x)
What is the smallest object visible with the human eye?
0.10 mm
How small can a compound microscopes view objects? Electron?
Electron- 0.2 nanometres
Compound- 0.2 um
What is a transmission electron microscope used to examine?
Internal Cell Structure
What is thin sectioning
Cell being cut in order to be viewed
Can transmission electron microscopes see through a cell?
no, the electron beams do not penetrate the thick cell wall
What is an example of a stain used when thin sectioning?
Uranium
Scanning electron microscopes view what?
Only the surface of an object
What is required to view something with a scanning electron microscopy?
The specimen must be coated with a thin find of heavy metal ex.gold
How magnified can a scanning electron microscope see?
15-100,000x
What is the most powerful electron microscope?
Scanning Tunneling Microscopy
What is a scanning tunnelling microscopy used to see?
Atoms
How does a scanning tunnelling microscopy work?
Uses a thin metal probe to scan specimens revealing surface irregulations
What must be used on the microorganism so that it is visible under a light microscope?
Stains
Stains are composed of what?
Negatively and positively charged ions
What is a chromophore?
The coloured negative or positive stain
What are the steps of the staining procedure for microorganisms before being viewed under a light microscope
- Thin film called smear containing the microorganism is put onto slide
- Sample is fixed by passing through a flame
- Stain is applied to sample
- Stain is removed by rinsing
- The stain is now ready to be viewed
The outer surface of bacteria has what type of charge?
Negative
How does a positive stain work?
Positively charged stain adheres to negatively charged bacterium therefore bacteria is coloured and background is clear
Whats an example of a positive stain?
Crystal violet
How does a negative stain work?
It is repelled by the negatively charged bacteria therefore bacteria appears clear and background appears coloured
What is an example of a negative stain?
Nigrosin (black)
What are two types of staining techniques?
Simple stains and differential stains
How does a simple stain work? What is sometime applied
Positively charged basic dye is applied therefore bacteria appears coloured, a mordant
What is a mordant?
Something used to increase the intensity of a stain
What is a differential stain used for?
To differentiate different types of bacteria because the stain react differently to different types of bacteria
What does a differential stain use to its advantage to be able to tell between cells?
It exploits differences in cells wall structure and composition
What are 2 examples of differential stains
- Gram stain
2. Acid Fast stain
What is the gram stain used to determine?
Whether a bacterium is gram positive or negative
When doing a gram stain, how can you tell which is gram negative and which is gram positive?
Gram postive- wil hold first purple stain
Gram negative- will not hold first stain but will hold second and appear pink
What occurs in a gram stain?
Two stains are used with a wash in between
Acid fast stains bind closely to..?
Bacteria that contain a waxy cell wall component
What is an acid fast stain used to identify?
All bacteria that fall within the genus Mycobacterium
What are two examples of Mycobacterium
Mycobacterium tuberculosis and Mycobacterium leprae
What occurs in an acid fast stain?
The waxy cell wall retains the carbo fuschin die that is added (pink on blue), a counterstain methylene blue leaves tissues and non acid fast bacteria blue
What are ‘special stains’ used for?
In order to visualize cell structures and specific parts of microorganisms
What are 3 types of special stains
- Capsular stain
- Flagella stain
- Endospore stain
What does a capsular stain do?
Reveals the presence of a thick polysaccharide layer outside the bacterial cell
What does a capsule presence indicate?
A bacterium with increased ability to cause disease
When doing a capsule stain, the background is coloured with what type of charged stain? The bacteria?
Negative (usually black), positive (pink)
What does the capsule look like after being dyed?
Remains colourless and appears as a halo around the bacterium
Why must special stains be used to detect the presence of endospores?
Ordinary stains cannot penetrate the bacterial cell wall
What occurs in an endospore stain?
Primary stain colours endospore green
A counterstain with safranin colours the rest of the cell pink
How does a flagella stain work?
The mordant and stain together work to increase the thickness of the flagella so it can be seen under a light microscope
