Week 3: Staining and observing microorganisms Flashcards
why is staining important? (2)
- increase the contrast to see the sample better
- differentiate between bacteria/genus
why is staining bad?
kills the cells so that we cannot view live organisms
what type of stain is gram staining?
a differential stain
steps of gram staining?
- CV
- Iodine
- Alcohol
- Safranin
what’s a common mistake of gram staining?
decolorize a smear for the incorrect amount of time
what happens if you over-decolorize?
a gram positive cell will appear pink after counterstaining
what happens if you under-decolorize?
gram negative cell appears purple
tips for gram staining? (3)
- make more than one slide of my sample
- don’t use phase contrast setting on stained samples
- move around the sample to find a good cell, not clumps
what is negative staining?
staining the background and the cell while the capsule stands out as being colorless
what is negative staining used for?
when water-soluble capsule of some bacterial cells is often difficult to see by standard staining procedures
what is the importance of capsules in negative staining?
they’re the area that’s not stained
three reasons why gram staining is bad:
- some bacteria don’t stain properly
- old samples don’t stain well
- cells are dead
three reasons why gram staining is good:
- differentiates based on cell wall
- narrows down the list of possible identities
- cheap, quick, easy to use
describe the steps of gram staining:
- CV: stain cells purple
- Iodine: binds to dye, both cells are purple
- Alcohol: in G+, binding tightens and stay purple. in G-, thin peptidoglycan layer is damaged and holes are punctured in the outer membrane, and are now colorless.
- Safranin: both cells are stained, but G+ stays purple and G- takes on a pink color.
