Lab Practical 1 Flashcards
Why are most college campuses equipped with compound light microscopes?
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Why is the microscope called a compound light microscope?
The term light refers to the method by which light transmits the image to your eye. Compound deals with the microscope having more than one lens. Microscope is the combination of two words; “micro” meaning small and “scope” meaning view
SCAN power
4x
total 40x
LOW power
10x
100x total
HIGH power
40x
400x total
Oil immersion
100x
1000x total
distance between the slide and the lens is called
working distance
magnificaion
the apparent increase in size of an object
total magnification
ocular power multiplied by objective power
located on the stage; this allows light from the lamp to penetrate the stage and illuminate the specimen
Aperture
What controls the amount of contrast the microscope specimen has
condenser and iris diaphragm
Which way do you move the diaphragm level to make the specimen darker
Right
Field of View
the area visible through the eyepiece
Most stains are charged…..
positive
Parfocal
the ability for an object to stay in relative focus when changing objectives
Why is oil used on oil immersion?
To decrease light refractivity entering the specimen.
Resolution
the fineness of detail that can be examined using a scope, the better the resolution the greater amount of detail that can be examined
What unit of measurement is usually used for measuring the size of cells
micrometer
Field of view SCAN
4.5 mm
field of view LOW
1.8 mm
field of view HIGH
0.45 mm
Field of view for OIL IMMERSION
0.18 mm
To determine average diameter of a cell
- Count # of cells that fit strait across field
- Estimate the size of cells by dividing field of view in mm by the number of cells that fit across the field of view.
- Convert number to micrometers by moving decimal point to the right 3 places
Protozoans
Protists
Single Cell
Motile
3 ways protoza move
Psudopodia
cilia
flagella
flagellated algae
protists
single cells
green
motile
filamentous algae
protists
cells from chains
non-motile
green
nonfilamentous and nonflagellated algae
protists
single cells
green
nonmotile
invertebrates
animals large multicellular motile (mosquito larve, rotifer)
diatoms
usually very detailed with 2 parts connected together
S.epidermidis
clusters of spheres
B. megaterium
chains of rods
negative stain
nigrosin or India ink (acidic) is placed on the slide and the bacteria is mixed into it and then spread out in a thin layer and allowed to air dry.
The stain in anionic so it stains the background rather than the cells.
Benefit of negative staining over simple or differential staining is…
there is no heat fixation needed which can shrink the cells.
Reason for heat fixing
to ensure cells are adhered to the slide so they aren’t washed off during staining and rinsing
and to ensure cell shrinking happens before staining so it doesn’t happen during staining which could result in distortion and artifacts
Simple Stain
positive charge (basic) Bacteria is stained bluish purple by the methylene blue
Gram Stain
a differential stain
Primary Stain - Crystal Violet (20 seconds)
Mordant - Gram’s Iodine (1 minute)
Decolorization - Ethyl Alcohol (10 -20 seconds)
Counter stain - Safranin (1 minute)
*rinse with H20 for 2 seconds between each step and blot dry when finished
Gram+
Stained bluish purple by crystal violet
B.megaterium & S. epidermis & S. aureus
Gram -
stain pinkish red
s. marcesens & E.coli
Acid fast stain
Differential stain
presence of mycobacterium (rod shaped w. special lipids in cell wall that are hard to stain - mycolic acid)
+ red - blue
2 different methods of acid fast stain
Ziehl Neelson method (same as Kinyoun but uses heat after phenol)
Kinyoun Method
Acid fast staining method
Primary - Carbolfuschin (red) - 5 minutes rinse w. water
Mordant - heat or chemical concentration
Decolorant - Acid-Alcohol (1 minute) rinse with water to stop
Counterstain - Methylene blue - 30 seconds
DO NOT BLOT DRY
Why is phenol important in acid fast staining
it is able to penetrate the wall of the mycobacterium so they can be stained.
Acid fast positive
Red/pink in color
ex. M. smegmatis
Acid fast negative
stained blue/purple in color
ex. S.epidermis S.aureus
Endospore stain
Stains endospore
Vegetative cell is Red
Endospore is Green (pale)
Method of endospore staining
Schaeffer-Fulton Method
heat is applied to ensure malachite green can adhere/penetrate the tough outer coating of the endospore
3 locations of endospores
Central, Terminal, and lateral
Why is knowing if endospores are present and their size and location important to a microbiologist?
It helps them identify what bacteria the endospore is from
Euglena
a photosynthetic protist
Green
motile
Yeast
show Brownian movement but not motile
grayish white in color look like long jelly beans
types of flagellar arrangement
monotrichous
lophotrichous
peritrichous
amphitrichous
