lab 4 goals and objectives: ***use wax pencils (china marker): sharpie will wash off with alcohol!...
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Lab 4 Goals and Objectives:
***Use wax pencils (China Marker): Sharpie will wash off with alcohol! Be sure to resuspend broth cultures completely!Exercise 15: Gram Stain: Each pair make 2 slides each mix and stain:
1. Staphylococcus aureus & Pseudomonas aeruginosa: two loops of each broth mixed
2. Bacillus subtilis & Escherichia coli: two loops of each broth mixed***95% Ethanol for Gram Stain, Acid-alcohol for Acid Fast Stain!!!
2 loopseach organismmixed together
Wax
P
enci
l
Exercise 17: Acid Fast StainingEach pair make 2 slides with:
Staphylococcus aureus (use 2-4 loops of broth on slide first)Mycobacterium smegmatis (slant, use needle, get tiny amount and mix into
broth on slide very well: break up all clumps)***Make all smears on slides first so they can dry before heat fixing!!!Split the work to save time but make sure everyone gets to see all slides!
Crystal Violet = Primary Stain: stains all cell walls purpleIodine = Mordant: combines with crystal violet to create large
insoluble dark purple complexAlcohol = Decolorizer: solubilizes Gram negative outer
membrane flushing iodine/crystal violet complex out of thin wall, thick Gram positive cells resist decolorization
Safranin = Counterstain: stains cell walls pink (obscured by purple walls of Gram positive, but Gram negative, who had become colorless, now are pink)
Lab 4 Goals and Objectives:
***Use wax pencils (China Marker): Sharpie will wash off with alcohol! Be sure to resuspend broth cultures completely!Exercise 15: Gram Stain: Each pair make 2 slides each mix and stain:
1. Staphylococcus aureus & Pseudomonas aeruginosa: two loops of each broth mixed
2. Bacillus subtilis & Escherichia coli: two loops of each broth mixed***95% Ethanol for Gram Stain, Acid-alcohol for Acid Fast Stain!!!
2 loopseach organismmixed together
Wax
P
enci
l
Exercise 17: Acid Fast StainingEach pair make 2 slides with:
Staphylococcus aureus (use 2-4 loops of broth on slide first)Mycobacterium smegmatis (slant, use needle, get tiny amount and mix into
broth on slide very well: break up all clumps)***Make all smears on slides first so they can dry before heat fixing!!!Split the work to save time but make sure everyone gets to see all slides!
Lab 4 Goals and Objectives:
***Use wax pencils (China Marker): Sharpie will wash off with alcohol! Be sure to resuspend broth cultures completely!Exercise 15: Gram Stain: Each pair make 2 slides each mix and stain:
1. Staphylococcus aureus & Pseudomonas aeruginosa: two loops of each broth mixed
2. Bacillus subtilis & Escherichia coli: two loops of each broth mixed***95% Ethanol for Gram Stain, Acid-alcohol for Acid Fast Stain!!!
2 loopseach organismmixed together
Wax
P
enci
l
Exercise 17: Acid Fast StainingEach pair make 2 slides with:
Staphylococcus aureus (use 2-4 loops of broth on slide first)Mycobacterium smegmatis (slant, use needle, get tiny amount and mix into
broth on slide very well: break up all clumps)***Make all smears on slides first so they can dry before heat fixing!!!Split the work to save time but make sure everyone gets to see all slides!