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Name the 4 reagents used in a gram stain in order of use:CV,GI,D,S
- crystal violet
- gram's iodine
- decolorizer
- safranin
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A substance that aids in the chemical reaction between the dye and bacteria. Making the dye absorb better.
mordant
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Name the mordant
gram's iodine
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What is made of acetone or alcohol?
decolorizer
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Name the counter stain
safranin
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What is made of carbol fuschin?
safranin
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Fill in the steps
1.place specimen on glass slide
2.air dry
3.
4.flood slide with crystal violet for 1 min
5.
- 3.heat fix
- 5.rinse slide carefully under slowly running water
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When heat fixing what should you be sure to do?
pass the slide through the flame several times
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When flooding the slide with crystal violet for 1 minute what should you be sure to do?
cover the entire area with stain
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Fill in the steps
6.flood stain with gram's iodine for 1 minute(cover entire slide with stain)
7.
8.
- 7.rinse slide again slowly
- 8.decolorize slide carefully with alcohol
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How long should decolorizer run?
until the slide runs clear
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Fill in the steps
9.
10.flood slide with safranin(counter stain) for 30 sec
11.rinse
12.
- 9.immediately rinse once the slide runs clear after decolorizing
- 12.blot slide carefully with paper towl
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How long is crystal violet applied to the slide?
1 minute
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How long is iodine applied to the slide?
1 minute
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How long is decolorizer applied to the slide?
until it runs clear
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Should decolorizer be left to set on the slide?
NEVER
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How long is safranin applied to the slide?
30 seconds
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What powers are used under the microscope?
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Why do gram + and gram - cells stain differently?
the amount of peptidoglycan in their cell walls
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Peptidoglycan provides strength, but what is it made of?
sugar and amino acids
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What cells stain purple?
gram + bacteria and yeast
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Why do gram + cells stain purple?
they have 5x's the amount of peptidoglycan and this absorbs the crystal violet
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What cells stain pink?
gram - bacteria and host cells
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In short name the steps of gram staining for review:F-C-R-I-R-D-R-S-R-B
- fix
- crystal violet
- rinse
- iodine
- rinse
- decolorizer
- rinse
- safranin rinse
- blot
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What bacteria has the thicker cell wall?
gram + because of the peptidoglycan
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What outer layer do gram - bacteria have that gram + bacteria do not?
LPS
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What does LPS stand for?
lippolysaccharide layer
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Where is the LPS?
it sits on top of the cell wall and is a second membrane
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What advantage do gram - bacteria have since they have the LPS?
harder to kill because it increases their resistance to antibiotics
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Name the 3 forms of bacteria
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GPCOC
gram positive cocci
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GNCOC
gram negative cocci
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What form of bacteria is rare?
spirillum
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What is the leading cause of cystitis?
gram - enteric bacilli
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gram + grape like cluster
staphyococci
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gram + chain
streptococci
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What 3 things can bacillus species look like?
- long/wide
- long/narrow
- branched
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Mixed populations are harder to kill what should be used?
broad spectrum abx (clavamox,baytril)
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Yeast can be what 2 shapes
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What are yeast called when they branch?
hyphae
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