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cell wall properties; differentiates between gram negative and gram positive (violet=gram positive, pink=gram negative)
gram stain (procedure)
1. smear prep, air dry, heat fix
2. stain with crystal violet (2 min)
3. add grams iodine or mordant (1 min), rinse with water
4. use ethyl alcohol for 5-10 seconds and rinse
5. add safranin (1 min) rinse and blot dry
used to see bacterial cell morphology, size, capsules. bacteria appear bright in a dark background
negative staining (procedure)
1. put inoculum on a slide
2. add one drop of nigrosin and mix it with a loop
3. use another slide to make a smear
4. dry and observe, no rinsing
**no heat applied
endospore staining schaeffer fulton
endospores will appear green and the vegetative cells red/pink
**heat is applied so that it can help the malachite green to penetrate the thick endospore
endospore staining schaeffer fulton (procedure)
1. prep smear, air dry, heat fix
2. put the slide on a beaker containing boiling water
3. add malachite green cont. for 5 min by putting a piece of paper towel on the slide
4. remove the paper towel and add safranin for 1 min
5. dry and observe
acid fast, ZN or kinyoun stain
mycobacteria will appear pink in a blue background
**can not perform gram stain as mycobacteria has mycolic acid which is waxy (lipid substance) the phenol in carbol fuchsin is lipid soluble
acid fast, ZN or kinyoun stain (procedure)
1. smear prep (with saliva)
2. add carbol fuchsin for 5 min (no heat)
3. rinse with acid alcohol
4. add methylene blue
4 basic requirements for growing microorganisms
3. growth factors
based on chemical composition
synthetic or defined media, complex or rich media (chemically undefined)
mannitol salt agar; supresses growth of all bacteria except Staph; 7.5% mannitol salt; phenol red to yellow; ph drops; color changes; S. spp makes yellow colonies and ferments mannitol; S. epi grows but does not ferment mannitol so does not make media yellow
selective for gram negative and differential for lactose fermentors; inhibitor is bile salts; indicator is neutral red (ph drop around colony causes color change); pink or white colonies with or without a zone of precipitation if lactose fermenters; if lactose nonfermenters they are clear colonies
thermal death time; the minimum length of time required to kill a bacterial suspension at a give temperature
main advantage is you can see viable cells to make spread plate; we use a technique called serial dilution
erroneously called blue green algae; found in aquatic lakes and oceans; some fix nitrogen in heterocysts (important fertilizer providers); oxygenic photosynthetic (in thylakoids) bacteria; unicellular; colonial; filamentous;
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