Disclaimer: The information provided in this

study guide and review session does not

replace the information provided by your
textbook, lab manual, or Professor Nguyen.
This is meant to be a supplement to the
aforementioned sources.
GO TO LAB
You will be tested on lab material
Almost NO make-up labs.
Be well prepared
Read ahead of time
Know the basic materials, procedures, and
biological processes
Complete your labs
Again, no make-up labs
Pay attention to your labs
You will be tested on them.
Make sure you record your results
Be mentally active
Think about what is going on during your lab
(theory)
Some things aren't explicit, you will need to think
critically
Take good notes in your lab journal
Pictures help too
Understand your lab before you leave
Unknowns
Procedures
Different tools, reagents, and types of media
used
The theory behind the experiment
Be able to identify different results
Be able to explain the results (both wrong and
right)
Be able to identify mistakes
Safety is usually fair game on quizzes and tests
in most science classes; as are the lab rules
Safety rules are there for you
Shoes
Hair
Lab rules are there to make sure you have a
proper lab to attend.
Not being safe and not following rules can lead
to a cut in participation points.
Mainly for you to learn how to use a microscope
properly
Essential skill for microbiology. You must know how
to use a microscope
Use proper procedures before, during, and after
microscope use
Still may be tested on the microscope
Parts, magnification.
Will benefit you for the rest of the semester.
If you do it well, your results will turn out
Unknown project
Key to remember is that microorganisms are
EVERYWHERE
Pay attention to morphology or it will haunt you
during your unknown.
Know why the stain works
Inside of cell is transparent and negatively charged
Basic stains are positively charged (chromogen)
Part that gives color is chromophore
Part that has a charge is auxochrome
Opposites attract
Heat-fixing
Pros: kills the cell; adheres cell to slide; coagulates
cytoplasmic protiens to make the more visible
Cons: distorts the cells; some organisms are too delicate to
withstand heat-fixing
Know types of basic dyes
EG: methylene blue, safranin, crystal violet
General rule, if dye stains the inside of the cell, the dye is
basic
Know the basic procedures, especially the bacterial smear
Fundamental staining procedure.
Also see section "Bacterial structure and Simple stains
Morphology of bacteria on a microscopic level as opposed
to the macroscopic colony morphology
Shapes, how many form, how they are clustered
Another skill you will
need to use throughout
the semester
You will have multiple
attempts
Unknown project
Acidic dye
Negative chromogen
EG: nigrosin, eosin
Used to identify bacteria that are too delicate to
withstand heat-fixing
One of the more important differential stains in
microbiology
Has clinical significance
Steps
Heat -fixed emulsion
Crystal Violet; rinse with DI water
Iodine; Rinse with DI water
Decolorize with ethanol or ethanol/acetone; Rinse
with DI water
Counterstain safranin; Rinse
How does the gram stain work?
Gram positive and gram negative cells have
structurally different cell walls
Gram positive: thicker, peptidoglycan layer with more
cross linking from teichoic acid
Gram negative: thinner, more lipids
Primary stain is CV
Can pass through both types
Iodine acts as a mordant
Binds to CV forming crystal violet-iodine complex; essentially a larger
molecule than CV alone
Decolorization
VERY IMPORTANT STEP
Must do properly to get good results
Alcohol/acetone removes lipids from cell walls; makes the wall more
porous
Gram negative: CVI washes out
Gram positive: CVI stays
Counterstain
CVI overpowers safranin so gram positive stays purple
Gram negative turns red