Biol2230 Lab Wk 10 Lab 9

Front 1

What are the steps in the gram stain we made:

Back 1

1. 1 loop water, Staph epi and E coli, smear, air dry, heat fix

2. Crystal violet for 20 sec; rinse

3. gram's iodine for 1 min; rinse

4. acetone alcohol for 10 secs; rinse

5. safranin for 20 secs

6. rinse, blot dry, observe

Front 2

What is the primary stain we used in the gram stain? How longs?

Back 2

crystal violet

20 secs

Front 3

After staining Staphylococcus epidermidis and E. coli with crystal violet in a gram stain, what would the slide look like?

Back 3

purple coccus (staph epi) and

purple bacillus (E. coli)

Front 4

What was the mordant we used in the gram stain? How long?

Back 4

Gram's iodine

1 minute

Front 5

What is a mordant?

Back 5

helps color adhere

Front 6

After using the Gram's iodine mordant in our Gram stain of Staph epi and E coli, what would the slide look like?

Back 6

purple coccus (staph epi) and

purple bacillus (E coli)

Front 7

What was the decolorizer we used in our gram stain? How long?

Back 7

acetone alcohol

10 secs

Front 8

After applying the acetone alcohol in the Gram stain of Staph epi and E coli, what would slide look like?

*** verify this info***

Back 8

you would see purple coccus (Staph) but wouldn't be able to see any bacillus (e coli) because it was decolorized?

Front 9

Why does the decolorizer only decolorize E coli and not Staph epi?

Back 9

Because E coli is gram negative, therefore it has peptidoglycan AND a lipid layer but the peptidoglycan layer is thin, therefore it decolorizers faster

***verify this info****

Front 10

What was the counterstain we used in the gram stain? How long?

Back 10

Safranin (red)

20 secs

Front 11

After applying the counterstain in our Gram stain, what would the slide look like?

Back 11

purple coccus (staph- gram positive) and

red bacillus (E coli- gram negative)

Front 12

What two differential stains did we do in lab 9?

Back 12

Gram stain

Acid-fast stain

Front 13

What color will gram negative bacteria stain?

Back 13

pink-red

(think stop- negative)

Front 14

What color will gram positive bacteria stain

Back 14

PPPPurple

(PPPurple= positive)

Front 15

What is the composition of gram positive bacterial cells?

Back 15

thick layer upon layer of peptidoglycan

Front 16

What is the composition of gram negative bacterial cells?

Back 16

thin layer of peptidoglycan and phospholipid layer (with LPS with exotoxins)

Front 17

When did we have to use goggles during lab nine?

Back 17

with acid-fast stain and endospore stain

-or with any stain that is being heated, as they might explode

Front 18

What are the steps in acid-fast stains

Back 18

1. loop water, Mycobacterium nonchromogenicium and Staph epi, smear, air dry, heat fix

2. add carbolfuchsin to paper towel covering smear in an aluminum weigh boat on a hot plate for 5 minutes

3. cool down, remove paper towel, rinse

4. acid-alcohol (decolorizer) for 3-5 secs; rinse

5. counterstain with Methylene blue for 30 secs

Front 19

What will the acid-fast stain look like after primary stain of carbolfucshin is applied to Mycobacterium nonchromogenicium and Staphylococcus epidermidis?

Back 19

red bacillus (tiny) (Mycobact. non.)

red coccus (Staph epi)

Front 20

What was the primary stain we used in acid-fast stain? how long?

Back 20

carbolfuschsin

5 minutes

Front 21

What decolorizer did we use in the acid-fast stain? How long

Back 21

acid-alcohol

3-5 seconds

Front 22

After using the decolorizer on the acid-fast stain, what would our slide look like? Why?

Back 22

Red Mycobacterium nonchromogenicium but would not see coccus (Staph epi) because it was decolorized

Mycobacterium is acid-fast, just like Cheer detergent is color-fast

Front 23

What counterstain did we use in the acid-fast stain? How long?

Back 23

Methylene blue

30 seconds

Front 24

What was the final result of our acid-fast stain?

Back 24

Red bacillus Mycobacterium nonchromogenicium

Blue coccus- Staphylococcus epidermidis

Front 25

What are the 2 types of acid-fast bacteria; according to LAB (not lecture!)

Back 25

Mycobacterium

Nocardia

Front 26

What are the steps in endospore staining?

Back 26

1. loop water, Bacillus megaterium, smear, air dry, heat fix

2. alumininum boat, paper towel, hot plate to 4, and Malachite green for 3-5 minutes

3. remove from heat, remove paper towel, cool down use Water as Decolorizer

4. counterstain with Safranin for 30 seconds

5. rinse, blot dry, observe

Front 27

What is the end result of the endospore stain?

Back 27

red bacillus and green endospores

Front 28

What is the primary stain in the endospore stain? How long?

Back 28

malachite green

3-5 minutes

Front 29

What is the decolorizer in the the endospore stain?

Back 29

Water

Front 30

What is the counterstain in the endospore stain?

How long?

Back 30

Safranin

30 seconds

Front 31

What would the slide look like in the endospore stain after applying the primary stain?

Back 31

both spores and bacillus would be green

Front 32

What would the slide look like in the endospore stain after using the decolorizer?

Back 32

Spores would be green but bacillus would not be visible because they were decolorized

Front 33

What is the end result of our endospore stain

Back 33

red Bacillus

green endospores

Front 34

What was the primary stain in the gram stain, acid-fast stain and endospore stain?

Back 34

gram stain- Crystal violet (20 secs)

acid-fast stain- carbolfuchsin (red) (5 mins heat)

endospore stain - malachite green (3-5 mins heat)

Front 35

Which stain used a mordant and when was it used?

Back 35

The gram stain used a mordant. It was used after applying primary stain (crystal violet- 20 secs) and before applying decolorizer (acetone alcohol - 10 secs)

Front 36

How long was the mordant applied and what was it called?

Back 36

Gram's iodine

1 minute

Front 37

Which stain did we use for Staph epi?

Back 37

Gram stain and acid fast stain

Front 38

Which stain did we use for E coli?

Back 38

Gram stain

Front 39

Which stain did we use for Mycobacterium nonchromogenicium?

Back 39

acid-fast

Front 40

What microorg did we use with the endospore stain?

Back 40

Bacillus megaterium

Front 41

What was the decolorizer used in gram stain, acid-fast stain and endospore stain?

Back 41

gram stain - acetone alcohol (10 secs)

acid-fast stain - acid-alcohol (3-5 secs)

endospore stain - water (as needed)

Front 42

What was the counterstain we used in gram stain, acid-fast stain and endospore stain?

Back 42

gram stain - safranin (20 secs)

acid-fast stain- Methylene blue (30 secs)

endospore stain - safranin (30 secs)

Front 43

Which stained microorg can take up to 8 weeks to grow in a culture?

Back 43

????acid-fast???

??endospore??/