micro-lab quiz

aseptic technique

procedures designed to prevent contamination of cultures by unwanted microbes

inoculation

act of placing bacteria into/on culture media

contaminant

unwanted microbes present in culture media

inoculating loop/needle

metal wire used to transfer microbes

incinerator

heat source that is used to kill any unwanted microorganisms on the inoculating tools

broth

a liquid medium
helps grow large amounts of microbes and/or cultivation of wide variety of microorganisms

agar slants

a solid, angled medium
helps with long term storing of pure cultures
gives greater surface area in test tube and minimize water loss

agar deeps

semi-solid medium
help grow bacteria that require less oxygen than is present on the surface of medium
can also determine motility of bacteria

petri dish

solid medium on flat surface (usually containing a type of Agar)
best to observe colony morphology and growth

staining for light microscopy

using dye to color emphasize specific structures

simple stains

uses one type of dye (ex: methylene blue)
can see size, shape, and arrangement of cells
can't differentiate between types of bacteria

differential stains

2 or more dyes are used
can differentiate between organisms or cell structures
two types: gram stain ad acid-fast stain

gram staining

differentiate between gram-positive bacteria and negative bacteria
gram positive=purple (thicker peptidoglycan layer)
gram negative=pink (thinner peptidoglycan layer)

acid-fast staining

uses a red dye and a blue dye
used to differentiate between gram positive bacteria that have waxy cell walls (ex: mycobacterium tuberculosis)
acid-fast cells will retain a red color
non acid-fast cells will appear blue

special staining

used to color unique structures of microbes that include endospore, flagella, and capsule

smear

a thin layer of bacteria on a clean slide
smears have to be heat-fixed to kill bacteria, preserves them from autolysis, and fix cells to slide

staining technique

1. prepare a heat-fixed smear
cells should be air dry before heat fixing
don't add too much heat
2. flood slide with selected stain
3. rinse with distilled water
repeat if necessary
4. blot dry with paper towel
don't add pressure

steps to gram staining

1. add primary stain (crystal violet dye) to heat- fixed smear (1 min)
2. add mordant (iodine) to help fix the dye to the cell walls (1 min)
3. use decolorizing agent (ethanol) to penetrate through gram negative cell walls to wash dye away (10-12 seconds)
4. add counterstain (safranin) to stain gram negative cells (45 seconds-1 min)